BrummellBlog JournalClub

Lacelle et al. 2010

2011-08-25T16:20:00.000-04:00

Lacelle D, Radtke K, Clark ID, Fisher D, Lauriol B, Utting N, Whyte LG. 2011. Geomicrobiology and occluded O2-CO2-Ar gas analyses provide evidence of microbial respiration in ancient terrestrial ground ice. Earth and Planetary Science Letters 306: 46-54.

These authors compared the gas composition of ice from massive ground ice bodies (e.g. 30 000-year-old buried snowbanks) to atmospheric gas concentrations and gas contents from glacial ice. They also cultivated some microorganisms collected from within the ice, and used some non-culture-dependent methods to examine the diversity of those organisms.

Loss of O2, as determined by comparison of the ratio of O2 to Ar in samples, as well as changes to the 13C-CO2 contents indicate heterotrophic microbial activity in the ice, most likely by organisms living in high-salt brine channels in cracks in the ice, using dissolved organic carbon as both energy and C source.

This paper provides an overview of methods for extracting and measuring the gases trapped in microscopic bubbles in permafrost and ground ice, as well as interesting findings about microbial activities.

Lacelle et al. 2008

2011-08-25T11:10:00.000-04:00

Lacelle D, Juneau V, Pellerin A, Lauriol B, Clark ID. 2008. Weathering regime and geochemical conditions in a polar desert environment, Haughton impact structure region, Devon Island, Canada. Canadian Journal of Earth Science 45: 1139-1157.

These authors examined the soils and waters near the Haughton crater on Devon Island, to determine the importance of chemical and mechanical weathering in this polar desert environment. They examined dissolved material in streams, lakes, snow, and groundwaters, and the size distribution, shape, and chemical composition of particles from soils in several different local landforms and parent materials.

Despite low temperatures, low precipitation, and very low vegetation presence, significant chemical weathering was found. Signs of chemical weathering, rather than mechanical, include rounded surfaces and pits in sand particles and the concentrations of Ca2+, Mg2+, and HCO3- in waters. Signs of mechanical weathering were also found, including sharp fracture lines in particles.

A gradient of increasing chemical weathering and decreasing mechanical weathering was found from the surface to the permafrost table. Thermal buffering reduces the frequency of frost-driven forces and thermal expansion from daily at the surface to annually at the permafrost table. The permafrost acts as a barrier to water movement, creating relatively wet condtions at depth that allow aqueous chemistry including the dissolution of dolomite to proceed.

Xu et al. 2009

2011-08-24T16:45:00.000-04:00

Xu C, Guo L, Ping C-L, White DM. 2009. Chemical and isotopic characterization of size-fractionated organic matter from cryoturbated tundra soils, northern Alaska. Journal of Geophysical research 114, G03002.

These authors examined the isotopic composition and organic matter distribution in soil horizons and particle size fractions from two soils in Alaska, a moist acidic tundra and a moist non-acidic tundra. The organic matter quality and quantity in the deeper part of the active layer and down into the permafrost, material estimated to be between 3000 and 7000 years old indicated high susceptibility to microbial activity, that is, decomposition to CO2 and subsequent release to the atmosphere.

I read this paper as part of my background reading to understand the potential uses of a Picarro field-portable carbon isotope analyser; this paper includes a description of the δ13C values of organic matter throughout the soil / permafrost profiles of these Alaskan soils. These values, and the associated discussion of signatures of microbial activity, suggest it is quite possible to distinguish the source of CO2, permafrost-SOM, deep-SOM, shallow/autotrophic, based on the 13C content of effluxing CO2 and / or CO2 at various depths within a soil profile. Table 5 is particularly valuable in this regard.

Lin et al., 2009

2011-08-11T15:00:00.001-04:00

Lin X, Wang S, Ma X, Xu G, Luo C, Li Y, Jiang G, Xie Z. 2009. Fluxes of CO₂, CH₄, and N₂O in an alpine meadow affected by yak excreta on the Qinghai-Tibetan plateau during summer grazing periods. Soil Biology and Biochemistry 41: 718-725.

I read this paper in an attempt to gain a better understanding of the methods used to compare greenhouse gas fluxes between ecosystems or treatments, and between gases, particularly the use of CO₂-equivalents when estimating total global warming potential contributions by ecosystems that may be simultaneously sources and sinks for the greenhouse gases CO₂, CH₄, and N₂O. In addition, this is one of a small number of studies I have been able to find that draw conclusions about global warming potentials based only on growing season measurements, rather than whole-year or growing season plus “cold season” (often what the tourist industry might call the shoulder seasons, spring and fall, though sometimes including winter as well).
These authors studied the effects of yak (Bos grunniens) excreta, dung and urine, on soil emissions of the three greenhouse gases. Excreta were hypothesized to increase GHG emissions because both are rich in nitrogen, especially inorganic forms of nitrogen such as urea, ammonia, and nitrate, contain sufficient water to stimulate microbial activity in dry soils, and, in the case of dung, are rich sources of labile organic carbon compounds and large microbial populations already present in the material. Furthermore, because production of CH₄ by grazing mammals is strongly linked to their digestive systems, fresh dung may contain considerable dissolved CH₄ that will be emitted quickly upon excretion.
The main results of this study were that while fresh dung did significantly shift a patch of meadow from a weak sink for CH₄ to a source, this difference was not sufficient to render the larger meadow area a net source because the spatial distribution of dung patches, as well as the duration of the CH₄ emission from dung, were relatively small. Urine application did not significantly increase CH₄ emission, which is surprising considering the high N concentration and rapid, large addition of water represented by urination by a yak; both factors are expected to increase methanogenesis.
Emissions of CO₂ were increased by dung, but not by urine when considering a longer, cumulative set of emissions. Interestingly, urine produced a significant pulse of CO₂ nearly immediately upon application to the soil, though whether this CO₂ is the result of hydrolysis of urea ((NH₂)₂CO + H₂O --- 2NH₃ + CO₂) or increased microbial respiration is not clear.
Emissions of N₂O were increased by both dung and urine application relative to untreated controls. However, the magnitude of the increase was less than predicted by IPCC (2001) guidelines for calculating the effects of grazing mammals on grasslands; those guidelines were based primarily on temperate low-altitude grasslands, not the high-altitude alpine meadows studied here. In general, patches of yak excreta accounted for an increase in N₂O emissions of up to about 10% compared to ungrazed and untreated control meadow, while total CO₂-equivalents emissions increased by about 1%, largely due to the small total areal extent of excreta patches.

Van Groenigen et al., 2005

2011-07-26T16:50:00.000-04:00

Van Groenigen JW, Zwart KB, Harris D, van Kessel C. 2005. Vertical gradients of δ¹⁵N and δ¹⁸O in soil atmospheric N₂O – temporal dynamics in a sandy soil. Rapid Communications in Mass Spectrometry 19: 1289-1295.

These authors examined the production and consumption of N₂O in soil profiles using stable-isotope analysis. Enzymatic processes in the microbial nitrification and denitrification pathways all strongly fractionate isotopes of both N and O, with products of reactions significantly depleted in the heavier isotopes. This allows identification of regions of soil producing or consuming N₂O by distinguishing between local production and diffusion through the soil layer.

Similar to my own studies, gas concentrations were measured using diffusion wells, in this case probes buried in the soil and sampled by syringe; gas samples were measured by GC-IRMS. Internationally-recognized isotope samples for N₂O are not available, so standards for analysis were prepared from commonly available laboratory chemicals and abiotic chemical reactions to completion to avoid fractionation (e.g. reduction of N₂O to N₂ over copper at 600ºC).

The largest concentrations of N₂O were found at the deepest sampling position, 90cm. There was a negative logarithmic relationship between the δ¹⁵N value of N₂O in the soil and depth, consistent with a relatively large single source of N₂O at 90 cm and consumption shallower in the soil combined with vertical-upwards diffusion of N₂O; reduction of N₂O to N₂ enriches the remaining N₂O pool for ¹⁵N.

Davidson et al. 1991

2010-12-03T12:05:00.000-05:00

Davidson EA, Hart SC, Shanks CA, Firestone MK. 1991. Measuring gross nitrogen mineralization, immobilization, and nitrification by 15N isotopic pool dilution in intact soil cores. Journal of Soil Science 42: 335-349.

These authors evaluated the use and limitations of the isotope-pool dilution technique when studying nitrogen dynamics in soil. Because addition of inorganic nitrogen compounds (NH4+, NO3-) can stimulate microbial activity in N-limited systems such as most soils, estimating the rate of these processes by tracking 15N through a system will almost certainly overestimate these rates. The isotope-pool dilution method, on the other hand, measures the dilution of enrichment in the nitrogen pool at the end of a particular process, relying on the assumption that additional product of metabolism will have negligible effects on the magnitude of that metabolism. In this study, immobilization of nitrogen was the main focus of investigation, comparing 15N isotope dilution in pools of either 15NH4+ or 15NO3-.

There are three key assumptions for the isotope-pool dilution method in this context. 1. Microorganisms do not discriminate between 15N and 14N; 2. rates of processes measured remain constant over the incubation period; 3. 15N assimilated during the incubation period is not remineralized. Previously, these assumptions had been evaluated for well-mixed soils, but not for unmixed field-collected soil samples. While fractionation by biological processes certainly does result in discrimination between isotopes of nitrogen, it is of negligible importance when injected solutions are very highly enriched and incubation periods are relatively short; in this case, injections were more than 90% 15N and incubations ran for 24 hours. Rates of measured processes will change if the population and / or activity of microorganisms changes, but again, over a 24-hour incubation period under controlled conditions this is unlikely. Highly enriched injections allow the use of small injection volumes, limiting the impact of nutrient enrichment. These authors were able to measure the remineralization of immobilized 15N, and estimated that between 1.0 and 1.6% of injected 15NH4+ appeared in the 15NO3- pool after 24 hours; they consider this an insignificant amount, but caution that longer incubations would almost certainly result in much more problematic amounts of remineralization.

This paper is clearly a major part of the basis of the project I am currently engaged in with Katherine from our 2009 field season at Alexandra Fjord. I probably should have read this paper long ago. The three main conclusions stated by these authors at the end of their paper I think can be quoted verbatim as justification for both why I (should have earlier) read this paper, and as a reminder to myself to include this paper in the methods & materials section of the eventual manuscript.

"Three points should be considered when applying the isotope dilution method.
1. Accurate estimation of both 14N and 15N initial pool sizes is important. Abiotic consumption of label, such as by clay fixation, can cause significant errors. A subset of intact cores may need to be destructively sampled directly after adding 15N for estimation of initial pool sizes.
2. Homogeneity of 15N enrichment throughout a soil sample is not possible, and perfectly uniform distribution of added label is not necessary. However, significant errors can arise from a bias in 15N distribution that is concurrent with a non-random
distribution of microbial processes. Distribution of label should, therefore, be as uniform as possible.
3. In situ gross immobilization rates may be overestimated by isotope dilution methods and underestimated by chloroform fumigation methods, depending on which (if any) kN factor is applied to the latter. Gross mineralization and gross nitrification estimates from isotope dilution are more reliable because these rates should not be affected by addition of 15N label in the form of the process products."

Miller et al. 2008

2010-12-02T10:05:00.000-05:00

Miller MN, Zebarth BJ, Dandie CE, Burton DL, Goyer C, Trevors JT. 2008. Crop residue influence on denitrification, N2O emissions and denitrifier community abundance in soil. Soil Biology & Biochemistry 40: 2553-2562.

These authors conducted a factorial experiment using packed soil cores to examine the influence of varying levels of available carbon and nitrogen on the process of denitrification. The treatments consisted of addition of glucose at three levels and KNO3 at four levels in experiment 1, and additions of either red clover or barley straw crop residues with or without additional KNO3. They measured soil chemistry, including extractable organic carbon and NO3- concentration based on K2SO4 extractions, as well as N2O production, the molar ratio of N2O (N2O : (N2O + N2)), and a handful of bacterial genes by qPCR.

The experimental setup was very similar to what we used in SLSC 802 (Special Topics) in the fall of 2010; cylindrical soil cores with gas-exchange holes in the sides were filled with soil at 1 g cm-3 bulk density and a water content of 70% and placed in 1 L canning jars with lids fitted with a perforable septum for gas sampling. One important difference between this experiment and what we have done is that in this experiment, gas measurements were of total cumulative gas production, whereas we flushed each jar with ambient air after each sampling event. Presumably this difference will have important effects on the formation of anaerobic conditions and microbial consumption of N2O previously produced under less anoxic conditions.

Not surprisingly, minimal denitrification activity was found in treatments without added NO3-. Starting NO3- concentrations were 3 mg NO3--N kg-1 soil, and fell in all treatments without added NO3- to less than 1 mg NO3--N kg-1. Once this supply of readily available nitrate was used, it appears the bacteria ceased denitrification activity, or at least it was reduced.
The red clover had a much lower C:N ratio than the barley straw, 13:1 and 45:1, respectively, and more labile carbon. This difference appears to have driven the observed difference in denitrification activity, in a manner that reflects the results of the simple-C-source experiment 1. In general, more labile C and more available N leads to stronger denitrification activity and greater production of N2O; in sealed jars such as these, strong respiration under these conditions leads to anaerobic conditions and a fall in the molar ratio of N2O as nosZ-equipped microbes consume N2O as a terminal electron acceptor.

Extractable organic carbon (EOC) was a relatively poor predictor of denitrification, compared to respiration as measured by CO2 production. EOC is a measure of the instantaneous size of the pool of labile C, while respiration represents carbon that has already passed through a microbe’s metabolism. The distinction here may be between two different pools of carbon, as well as between an instantaneous snapshot measure and a series of measurements readily convertible to an estimate of the rate of a process.

In conclusion, these authors reiterate their finding that available C and available N (especially as NO3-) are strong predictors of denitrifying activity, across a range of C and N sources. I read this paper for the class SLSC 802 in the fall of 2010, but the portions describing denitrification physiology and especially the qPCR information will be generally useful to my other projects.

Delgado et al. 2010

2010-09-29T18:15:00.000-04:00

Delgado JA, Del Grosso SJ, Ogle SM. 2010. 15N isotopic crop residue cycling studies and modeling suggest that IPCC methodologies to assess residue contributions to N2O-N emissions should be reevaluated. Nutrient Cycling in Agroecosystems 86: 383-390.

These authors reanalyzed two recent reviews of measuring nitrous oxide emissions from agricultural systems and used a model to simulate N2O emissions and NO3 leaching associated with cropping practices in Colorado and Iowa. In general, use of crop residue instead of or in addition to synthetic fertilizers significantly altered patterns of N loss, whereas IPCC recommendations assume no difference between these N sources in regards to N2O emissions. Microbial immobilization of nitrogen, particularly associated with residues with high C/N ratios, is a major factor in these differences, and these authors provide supporting arguments for their suggestion of revisions to IPCC recommendations and modeling.

Huang et al. 2004

2010-09-28T17:45:00.001-04:00

Huang Y, Zou J, Zheng X, Wang Y, Xu X. 2004. Nitrous oxide emissions as influenced by amendment of plant residues with different C:N ratios. Soil Biology & Biochemistry 36: 973-981.

These authors examined the role of residue quality, in the form of C:N ratio and a range of crop residues, on N2O emissions from soils. They also measured CO2 emissions, and found strong correlations between organic-matter decomposition and respiration, and nitrogen cycling.

Gas fluxes of CO2 and N2O were highly correlated across all incubations. To ensure only respiration-derived CO2 was measured, the CO2 released by urea breakdown in urea-treated treatments was calculated and subtracted; respiration in the urea-only treatment was similar to that in the untreated controls. Both gas emissions were negatively correlated with residue C:N ratios. Finally, residue C:N ratios were negatively correlated with dissolved organic carbon concentrations.

Overall, higher C:N ratios in residues seem to result in slow decomposition of mainly recalcitrant organic matter, and low CO2 and N2O emissions. Addition of urea in conjunction with crop residues produces a range of N2O emissions depending on the C:N ratio of the residues.

This short paper may serve as a model for the work I will be doing in the special topics class in soil science, fall 2010.

Trinsoutrot et al. 2000

2010-09-28T11:35:00.002-04:00

Trinsoutrot I, Recous S, Mary B, Nicolardot B. 2000. C and N fluxes of decomposing 13C and 15N Brassica napus L.: effects of residue composition and N content. Soil Biology and Biochemistry 32: 1717-1730.

These authors studied the decomposition process by soil microorganisms when isotope-labelled crop residues were added to soil. The crop used, oilseed rape Brassica napus (also known as canola) varies its nitrogen content of tissues, and the C:N ratio, depending on levels of N inputs by fertilization. This allows variation in input organic matter quality by manipulation of growing conditions; in this experiment, both carbon and nitrogen inputs to the plant included stable-isotope labels, in the form of 13C-CO2 and 15N-KNO3. Plant residues were added to soils and incubated for 168 days.

Initial C:N ratio and especially the labile-C fraction of organic-matter inputs are major controls of both the rate of decomposition and fate of matter through the system. Additionally, temperature, particle size of residues, and water content in the soil also strongly influence decomposition processes.

Here, N mineralization (the formation of NO3- and NH4+ pools in the soil from organic-N precursors) occurred in two phases. In the early phase, up to about 3 weeks, the N cycle resulted in net mineralization. Later, mineral N pools were depleted and N was immobilized, that is, incorporated into the tissues of microbial cells.

Carbon dioxide release during the experiment occurred through two pathways. The more direct route was rapid mineralization of organic matter, which I interpret as non-incorporation of organic matter by microbes, consuming such material but metabolizing it rapidly through respiration. The second, presumably slower route was through metabolization of material after incorporation into cells through respiration. Either way, the ultimate fate of much of the organic-C in the residues was release as CO2.

Differences in the N-content of residues affected decomposition rates early in the experiment, but by about 4 months the differences between high-N and low-N residues had evened out. Only a small fraction of labelled N from residues ended up in soil mineral-N pools; the majority was either immobilized into microbial cells or remained in recalcitrant organic matter fractions. Immobilization of unlabeled, SOM-derived N was enhanced by the addition of C through a substitution effect.

These authors conclude that 15N labelling was fraught with difficulties, and both under- and overestimated some pools and processes. However, the use of their model, named NCSOIL, improved their ability to trace the fate of added material through the system. This paper represents a study similar in some ways to our planned course activity in the special topics in soil science course, fall 2010.

Smith et al. 2003

2010-09-27T17:15:00.000-04:00

Smith KA, Ball T, Conen F, Dobbie KE, Massheder J, Rey A. 2003. Exchange of greenhouse gases between soil and atmosphere: interactions of soil physical factors and biological processes. European Journal of Soil Science 54:779-791.

These authors present a broad review paper of the role of soil physical factors, mainly temperature and water-filled-pore-space, in controlling soil emissions of the greenhouse gases CO2, CH4, and N2O. The paper’s goal is stated to be to expose a variety of researchers to the links between soil physics and soil biology, as well as the importance of these fields to current research in many disciplines on global warming.

All three gases are produced and consumed in soil primarily by microorganisms, which respond to variation in soil physical parameters in different ways. In general, both temperature and WFPS impact GHG production. Higher temperatures almost always result in increased production of gases, though the Q10 values (measuring the magnitude of response to a change of 10º of temperature) vary widely in the literature for all three gases. The effect of WFPS is different, involving upper and lower bounds, though in the middle range increasing WFPS generally promotes increased gas production. Microbes are limited in their tolerance of dry conditions, such that gas production falls rapidly below some critical WFPS value; for CO2 this threshold is near 20%. WFPS is also indirectly important, through its effects on soil diffusivity. Higher WFPS as well as higher bulk density are associated with lessened CH4 oxidation, due to reduced diffusivity of both CH4 and atmospheric O2. Very high WFPS values are associated with reduction of N2O to N2, partly by limiting O2 supplies and creating larger anaerobic microsites, and partly by preventing the escape of N2O gas into rapid-diffusion pathways; it is trapped in the vicinity of microbes capable of using it as an electron acceptor.

There are other factors controlling net GHG emissions, such as the relationship between plant productivity and water table position, which will change the relationship between rates of soil organic matter oxidation to CO2 and the removal of CO2 from the atmosphere by plants; trees in particular can lower local water tables, increasing SOM oxidation while simultaneously consuming more CO2 than the previous wetland vegetation community.

I read this paper on the suggestion of my coworkers in the special topics class of fall 2010, but it applies well to the general area of my research. The reference list includes multiple interesting papers addressing particular specialties within this large topic.

Pennock et al. 1987

2010-08-23T17:45:00.000-04:00

Pennock DJ, Zebarth BJ, De Jong E. 1987. Landform classification and soil distribution in hummocky terrain, Saskatchewan, Canada. Geoderma 40: 297-315.

These authors present a method of analysing irregular terrain for the purposes of examining important soil parameters such as soil depth and soil hydrology. Three variables (profile curvature, plan curvature, and gradient) can be calculated from a matrix of elevation data; taken together for a given point or area, these variables can be used to classify an area into one of seven landforms. These landforms are level (for summits or bottom lands), shoulders, backslopes, and footslopes; the three non-level forms come in “divergent” (convex plan) and “convergent” (concave plan) varieties. Shoulders have convex profiles, backslopes have flat profiles (i.e. constant gradient when looking up or down the slope), and footslopes are concave. Divergent landforms shed water laterally; convergent landforms tend to collect water. Where water collects and is moving slowly, rates of infiltration will be highest, and erosion will tend to deposit, rather than remove, material at these places.

The variables required to calculate profile and plan curvature and gradient are relatively easy to calculate from a matrix of elevation data, using an interpolating topographical software package and some differential calculus. The seven categories of slope elements can be estimated in the field by eye, making for a useful method for field studies.

This paper was on the recommended reading list for SLSC 834; in addition, the course instructor is Dr. Pennock, lead author of this study. The study site, near Hafford, Saskatchewan, is perhaps 1 to 1.5 hours drive away from Saskatoon, suggesting this area may be the destination of one of the day trips scheduled for the week of August 30, 2010.

Hayashi et al. 1998a

2010-08-20T14:15:00.000-04:00

Hayashi M, van der Kamp G, Rudolph DL. 1998. Water and solute transfer between a prairie wetland and adjacent uplands, 1. Water balance. Journal of Hydrology 207: 42-55.

These authors report water balance data and analysis in the first of a pair of papers about the hydrology of wetlands in the prairie pothole region of central Saskatchewan. The second paper concerns dissolved material movements in the same example study system.

Through a series of measurements of water and water-related properties of soil as well as precipitation, total water balance and total water movements within the wetland were estimated. Direct precipitation and evaporation accounts for a small fraction of the total volume of water entering and leaving the pond in the middle of the hectare-scale wetland and surrounding catchment of uplands. Most of the water entering the pond is in the form of snowmelt during spring, and most of the water leaving the pond is in the form of horizontal groundwater flow, relatively shallowly (i.e. above a redox discontinuity at about 5-6m depth), driven strongly by evapotranspiration through the “willow ring” surrounding the pond and through crops (principally wheat during this experiment) on the uplands. Only a small amount of water is exported down to the aquifer; of a total input of 780mm into the pond, only about 3mm goes to recharging the underlying aquifer. Figure 3 provides definitions of the terminology, and Figure 12 the estimates of water balance.
This paper was on the suggested reading list for SLSC 834 in August 2010.

Christiansen 1979

2010-08-19T17:30:00.000-04:00

Christiansen EA. 1979. The Wisconsinan deglaciation of southern Saskatchewan and adjacent areas. Canadian Journal of Earth Science 16: 913-938.

This author describes in considerable detail the process of deglaciation that occurred at the end of the last glacial maximum from about 17 000 years ago, as it occurs to Saskatchewan. Major geological features and patterns of melt-water drainage led to the inclusion of nearby parts of Alberta, Manitoba, Montana, and North Dakota in the analysis. Patterns during the deglaciation were identified by glacial landforms, many of which are apparent only from aerial photographs, with sample analysis in the lab including radio-carbon dating.

The period from 17 000 years to 10 000 years ago is divided into 9 phases corresponding to periods of rapid glacial retreat or temporary stasis or glacial advance. By 10 000 years ago the ice sheet that had covered nearly the entire province had retreated towards Hudson Bay and covered only a small part of the north of Saskatchewan. Glacial lakes formed from meltwater and from water flowing from the west (presumably sourced from glaciers in the Rocky mountains), sometimes reaching enormous sizes; these lakes were bordered by the glacier’s edge, and connected to each other via spillways that often carved large channels from the plains; the east-west valleys of southern Saskatchewan such as the Qu’Appelle valley are the remains of such spillways. Modern river systems such as the Saskatchewan River and the Churchill River formed during the glacial retreat, occupying low areas and spillway remnants.

The rate of deglaciation varied considerably over the studied 7 000 years, generally accelerating from about 150 m / yr to around 275 m / yr, though with frequent pauses, occasional re-advances, and variation across the glacial edge. The evidence from glacial lake-edge movements and depth patterns suggests the ice sheet melted fastest initially, but with the slowest retreat at that time indicating the sheet first thinned, and then retreated, especially around newly-uncovered Nunataks where the underlying land formed highlands.

This paper was on the suggested reading list for the course SLSC 834 in August 2010, but it is also personally interesting in describing Pleistocene events in areas I visit during Sunday drives across the center of the province.

Wilkinson et al. 2009

2010-08-17T16:00:00.000-04:00

Wilkinson MT, Richards PJ, Humphreys GS. 2009. Breaking ground: Pedological, geological, and ecological implications of soil bioturbation. Earth-Science Reviews 97: 257-272.

These authors describe the process of bioturbation in soils, and the roles of various major groups of organisms, especially animals, responsible for bioturbation. Through mixing and surface-mounding, bioturbation sorts, buries, and transports soil and soil components such as organic matter. Bioturbation also interacts with abiotic processes, particularly those involving surface water inputs and movements, to create net effects on soil movement and sorting.

There is a great deal of detail in the description of various sub-processes and rates, as well as some urging for more detailed studies of these phenomena. This paper is required reading for the course SLSC 834, in August 2010.

Johnson et al. 2005

2010-08-16T14:45:00.000-04:00

Johnson DL, Domier JEJ, Johnson DN. 2005. Reflections on the nature of soil and its biomantle. Annals of the Association of American Geographers 95: 11-31.

These authors advocate a new paradigm to underlie studies of soil science and related fields, based on increased recognition of processes occurring in and responsible for the production of the “biomantle”, the upper layer of soil composed of and formed by the actions of organisms. Many of these processes are based on movements of soil and soil components, and so are dominated by animals, especially active burrowers that are responsible for large vertical movements of soil in some environments. The biomantle is defined as being composed of “biofabric”, or materials that owe their existence to the actions of organisms, from the bodies of these organisms themselves, to the materials released by the organisms, to the minerals created by biological processes, to the voids created by their movements and the gases filling those voids released by their metabolisms.

These authors trace their ideas from the writings of Darwin, particularly his final work involving the activity of worms in “vegetable mould”, a late-18th century term for what we now call soil.
A biomantle layer, residing chiefly in the A horizon (or topsoil) can be more easily recognized in some landscapes than others. Humid tropical soils especially may show very thick and distinct biomantles, in two layers. The upper, thicker layer is composed of relatively fine materials, resting on a basal layer of coarser material; this basal layer is referred to here as the stonelayer. Below the stonelayer is non-biomantle, typically a B horizon (or subsoil). The hypothesized process creating this two-layer biomantle is the action of “conveyor belt” animals, especially termites that carry small particles upwards but are unable to move larger stones, thus eventually sorting the soil mineral material.

In other soils, such as loess-derived sandy soils without a large component of gravel and larger stones, such a two-layer biomantle may not form, or may be very weakly developed and difficult to identify as such. Nonetheless, bioturbation activity by burrowing animals is usually apparent, for example in the form of “krotovina”, in-filled animal burrows.

Besides advocating for a view of soils and their processes with an animal-based, biomantle point of view, these authors spend some time dismissing subaqueous soils (e.g. marine sediments) as simplistic, uncomplicated places lacking many of the key (and very complex) processes that occur in subaerial soils. Their list of such processes near the end of the paper, taken as a kind of justification for their uncited and unsupported dismissal of subaqueous soils, is composed entirely of those processes relating to changing water amounts in terrestrial soils, such as groundwater flow and wetting and drying events. I found their argument unconvincing, as they do not describe any aqueous-only processes such as changes in dissolved-O2 concentrations or the sorting action of water currents, and their blithe disregard for marine biodiversity in statements about how much more diverse the life in terrestrial soils must be, is the proverbial icing on the insult cake. Johnson et al.: please cite some evidence to support such sweeping generalizations.

Stark and Hart 1996

2010-04-20T10:15:00.000-04:00

Stark JM, Hart SC. 1996. Diffusion technique for preparing salt solutions, Kjeldahl digests, and persulfate digests for nitrogen-15 analysis. Soil Science Society of America Journal 60: 1846-1855.

These authors evaluated the recently-developed Teflon-encased-acid-trap technique for collecting nitrogen from water samples for analysis, especially stable isotope (15N) analysis. Earlier acid trap techniques, which rely on the chemistry of ammonium/ammonia in solutions of varying pH, were based on suspended small vials containing acid solutions above a larger volume of sample solution at high pH. Ammonium present in the sample is converted to ammonia by high pH, and escapes solution into the gas phase. It is captured in acid solution; in a closed container with a separate acidic region, ammonium will migrate from the alkaline sample to the acid trap. Suspended trap designs are vulnerable to a number of problems, including vulnerability of the acid trap to contamination by alkaline solution and subsequent loss of captured ammonium. Teflon will pass gases but not liquids, and therefore provides an ideal barrier between the sample and the acid trap that will permit ammonia to enter the trap.

These authors conducted a set of six experiments to evaluate the utility of Teflon acid traps. First, blanks were evaluated, to determine the sensitivity of the technique to nitrogen derived from contamination in the various materials of the experiment (filter paper, Teflon, plastic bottles, etc.). There was some nitrogen detectable from these non-sample sources, but a correction factor could be constructed based on including a few blanks in sample sets. The second experiment examined the recovery of ammonium from 2M KCl solutions. These solutions are very similar to the solutions extracted from soils of Alexandra Fjord in 2009. As might be expected, incubation time correlated with recovery of ammonium; in addition, Teflon traps performed better over shorter incubations (up to about 6 days) than did suspended traps, though the difference disappeared at the longest (8 days) incubations.

The third experiment was of greatest interest to me. They examined the open-bottle time necessary to eliminate residual ammonium from samples used in experiment 2, and the incubation conditions to collect NO3- -derived nitrogen using Devarda’s alloy to reduce NO3- to NH4+. Residual ammonium left in solution at the end of the first reaction would contaminate NO3- examination and lead to overestimation of NO3- contents; this is especially true when the two forms of nitrogen have been previously enriched with 15N to different degrees. There is a trade-off between open bottle time (ranging in these experiments from one to five days) and recovery of NO3- from samples; long open-bottle times appear to lead to an unknown chemical change in samples, which these authors speculate, may involve sorption of atmospheric CO2. Long ammonium-collection times (i.e. experiment 2) do tend to collect the great majority of ammonium present, recovering on average more than 97% of ammonium from known-concentration samples. This suggests to me a long and potentially nitrate-losing open-bottle incubation is not necessary if ammonium collection has proceeded for at least six days. Agitation of samples, in these experiments often once per day, resulted in a strong improvement in nitrogen capture over at least the shorter (24-96 hour) incubations; our use of continuous agitation for seven days therefore seems likely to capture nearly all ammonium present in samples.

Experiments 4 through 6 were of little interest to me, as I am unfamiliar with Kjeldahl digests and persulfate digests and do not plan to use these techniques in my studies. From what I could glean based on my limited knowledge, the Teflon-acid-trap technique appears to work well with Kjeldahl digests and persulfate digests.

Overall, the Teflon-acid-trap technique performed very well, and had significant ease-of-use and contamination-avoiding advantages over earlier methods. One caution put forth by these authors is regarding the H2 gas evolved during Devarda’s alloy incubations; strong build up of gas can cause leaks, potentially allowing NH3 to escape and leading to underestimates of sample nitrate contents. They suggest storing bottles upside-down during such incubations, as leaks will then include sample solution, and leaking bottles can be easily identified.

Complete recovery of nitrogen was not possible on a routine basis using this technique, but 15N isotope ratios do not rely on 100% recovery if ammonium and / or nitrate concentrations have been measured by an independent method, such as the widely-used colorimetric techniques. The two isotopes of nitrogen do not migrate differently between alkaline sample and acid trap, thus even at relatively inefficient recovery rates, isotopic ratios are maintained. Blanks are important, but in general this technique is robust, reliable, and easy to conduct.

Palmer et al. 2010

2010-04-15T09:35:00.000-04:00

Palmer K, Drake HL, Horn MA. 2010. Association of novel and highly diverse acid-tolerant denitrifiers with N2O fluxes of an acidic fen. Applied and Enironmental Microbiology 76: 1125-1134.

These authors examined soils from an acidic fen in southern Germany, and discovered novel denitrifiers that are apparently adapted to local conditions and contribute to the cycling of nitrogen within the fen. Methods employed included measurement of soil parameters, microcosms to examine denitrification rates (both total denitrification and net production / consumption of N2O), cell counts of cultured organisms, and phylogenetic analysis using narG and nosZ sequences and RFLP.

Palmer et al. 2009

2010-04-06T17:30:00.000-04:00

Palmer K, Drake HL, Horn MA. 2009. Genome-derived criteria for assigning environmental narG and nosZ sequences to operational taxonomic units of nitrate reducers. Applied and Environmental Microbiology 75: 5170-5174.

These authors compared the sequences of narG and nosZ genes to corresponding sequences of 16s rRNA genes, using in-silico analysis of sequences downloaded from GenBank. While similarities above 97% are commonly used for species- or genus-level taxonomic delineation for 16s sequences, this analysis found much lower threshold similarities for such delineation using the structural genes.

This paper is confusing to me. One part of the text appears to contradict itself, when the authors state that the Nar operon in Pseudomonas stutzeri A1501 is putatively alien in origin (i.e. recent horizontal transfer), then go on to state in the same paragraph that it appears unlikely that the Nar operon was horizontally transferred in any species. I may just be misunderstanding the meaning of the term “putatively alien” in regards to a bacterial gene sequence.

A greater puzzle is presented by the list of nosZ sequences. These authors downloaded 85 such sequences, where my own attempts to extract data from GenBank resulted in only 42 unique nosZ sequences. The list in a supplementary table includes several cases of multiple accessions of the same species but of different PD. The paper these clusters of PD-sequences are derived from is Dandie et al. (2007); a quick scan of this paper did not reveal what the distinction “PD” indicates.

Tuomivirta et al. 2009

2010-04-01T18:30:00.001-04:00

Tuomivirta TT, Yrjälä K, Frize H. 2009. Quantitative PCR of pmoA using a novel reverse primer correlates with potential methane oxidation in Finnish Fen. Research in Microbiology 160: 751-756.

These authors developed novel primers for studying the methanotrophs of Finnish peatland fens. The existing primer pairs widely used to study methanotrophs, based on the gene pmoA, did not consistently amplify useful products in PCR using template DNA from these fens.

The novel primer pair was tested on 114 samples from two Finnish fens, representing the northern and southern peatlands of Finland. While A189f/A682r failed to provide useful products, the new primer A6821r in conjunction with A189f did produce strong bands. In qPCR, these primers produced results correlated with measured methane oxidation potential, further supporting their utility in these systems.

Schmidt et al. 2008

2010-04-01T10:25:00.000-04:00

Schmidt SK, Reed SC, Nemergut DR, Grandy AS, Cleveland CC, Weintraub MN, Hill AW, Costellow EK, Meyer AF, Neff JC, Martin AM. 2008. The earliest stages of ecosystem succession in high-elevation (5000 metres above sea level), recently deglaciated soils. Proceedings of the Royal Society of London, Series B 275: 2793-2802.

These authors describe the microbial community and soil parameters of a chronosequence at the foreground of a receding glacier high in the Peruvian Andes. From a combination of aerial photography and previous work at this site, a series of sites of soils of increasing ages from zero to 79 years old was established. No surface plants, even lichens, are present on any of these new soils, and soil nutrient levels (carbon, nitrogen) are very low; the only organisms present are microorganisms.

Two previous hypotheses had been proposed to explain the dynamics of very early primary succession on new soil. Organic matter has been observed to accumulate slowly in new soils; the source of this material is either aeolian deposits (i.e. wind-borne plant detritus and pollen) or in-situ fixation of CO2 and N2. These are not mutually exclusive hypotheses, but the relative contributions of each are explored in this study.

The methods used here cover an extensive list of soil parameters. Three sets of soil samples were collected: for microbiological analysis, N-fixation measurement, and all other chemical analyses. The other chemical analyses include photosynthetic pigment extraction, soil total and mineral nitrogen, pyrolysis for identifying sources of carbon compounds (i.e. microbial-autotroph, microbial-heterotroph, plant), enzyme assays for common and informative microbial enzymes, and soil stability analysis of the resistance of these new soils to erosion forces such as water runoff.

These authors focused on the cyanobacterial fraction of the microbial community in this study; some details of other components of the biota are described in an earlier paper, Nemergut et al. (2007). Cyanobacteria are autotrophs also capable of fixing atmospheric nitrogen, thus they are ideal primary colonizers of new soil as they require little more than a source of moisture and air. Analysis of the community included the use of the P-test (Martin 2002); note that as in Nemergut et al. (2007), he is one of the authors of this study. The analytical approach is very similar to that employed in the earlier study, with a comparison of discovered sequences to published sequences from around the world. In this study, cyanobacterial sequences from zero and 4-year-old soils were similar to sequences from an extremely broad sample of habitats, including Antarctic lake ice, marine subseafloor sediments, urban aerosols, forest soils, and oil-polluted soils.

The soil chronosequence showed a clear pattern of stages of primary succession at every level of analysis. The soil microbial community became both more abundant and more diverse through time, soil nutrients increased, the chemical environment included increasing amounts and diversity of complex organic molecules, key enzyme pathways became established, and soil stability increased as soils aged. N-fixation showed a peak, with increasing N-fixation activity from the zero to 4-year-old soils (by two orders of magnitude), then declining by about half in the 79-year-old soils. This mirrors and precedes a widely-observed pattern in plant primary succession, in which nitrogen-fixing plants are among the first colonizers, but decline in abundance at later stages of succession. Enzyme and organic molecule patterns were consistent with a total absence of heterotrophs in the extremely young soils, increasing occurrence of organisms capable of decomposing plant matter in the 4-year-old soils, and a molecular ecology qualitatively similar to a mature plant-associated soil in the 79-year-old soil.

The list of procedures and level of detail of analysis in this paper is impressive. Many, though certainly not all, of these techniques will be models for my own work, especially in the summer of 2010. The molecular-diversity techniques pioneered by Martin (Martin 2002, Nemergut et al. 2007, this paper) as well as the techniques of analyzing soil pigments and soil nutrients are all very interesting.

Sørensen et al. 2006

2010-03-31T16:10:00.001-04:00

Sørensen LI, Holmstrup M, Maraldo K, Christensen S, Christensen B. 2006. Soil fauna communities and microbial respiration in high Arctic tundra soils at Zackenberg, Northeast Greenland. Polar Biology 29: 189-195.

These authors sampled soil animals from three sites at Zackenberg station, Greenland, over three days in mid-summer. Two of the sites were considered mesic heath, with a mix of Cassiope tetragona and other High Arctic species of plants, while the third site was dominated by Dryas spp. and was considered dry heath; snow melts from the dry heath up to 20 days earlier than from the mesic heaths. Soil samples ranging down to about 6cm depth were collected, stored at 5ºC for up to two weeks, and analyzed by a range of methods in the laboratories in Europe.

Different groups of soil animals were extracted by varying methods. Soil microarthropods, a diverse group dominated by Collebola and Acari, were extracted by modified MacFadyen funnels into Benzoic acid. Enchytraeids and dipteran larvae were extracted in Baermann wet funnels with heating of the samples, into tap water. Protozoa were washed from soils in water and grown on media plates in the dark at 10ºC. Nematodes were collected by the Blender-Cotton wool method of Schouten and Arp (1991). Soil microbial respiration was measured in serum bottles, with the CO2 concentration in the headspace measured at zero, 5 and 25 hours, with a fully factorial design of nutrient amendments of C, N, and P. Soil pH and soil organic matter content, but not moisture content or other nutrient concentrations were determined using methods not clearly described, though presumably these procedures were similar to standard methods.

Once abundance and biomass data was collected, comparisons between plots were made using multivariate analysis and a software package named PRIMER 5.0. My understanding is the species counts were (log+1) transformed to reduce the influence of very abundant species, then analyzed using an approach similar to Principal Components Analysis. The result of this analysis was a clear difference between the dry heath and the two mesic heaths, while the two mesic heaths were not different from each other in parameter-space. A Bray-Curtis similarity matrix was also involved, though I’m not certain I understand how.

Different taxonomic groups were identified to different taxonomic levels; 19 species of Collembola and 7 species of Enchytraeids were found, for example, but Acari were identified to suborder (Cryptostigmata (oribatids), Prostigmata, Mesostigmata) and nematodes and protozoans were counted at those high taxonomic levels. While the two mesic heath sites were only marginally significantly different from each other, there was a clear increase in abundances in the dry heath site. For collembola at least, the dry heath site was also dominated by two highly abundant species, which differed from the majority of species in the mesic sites by being unpigmented and associated with sub-surface, rather than soil-surface, regions in the soil. The higher abundance of probably bacteria-eating nematodes at the dry heath strongly suggests higher turnover of microorganisms as well as generally higher biological activity from the higher populations of most soil animals.

These authors suggest higher organic matter decomposition rates at the dry heath, which seems reasonable given the higher animal populations there. However, their attribution of higher soil pH there to higher respiration levels seems like more of a stretch, absent supporting mineralogical and soil-nutrient data.

This paper provides an excellent example of the data that can be collected and analyzed from a brief but intensive study of soil invertebrates at a High Arctic site. In addition, meaningful information about differences in biodiversity between locations can be derived from studies of organisms not identified to fine taxonomic levels.

O'Neill et al. 2010

2010-03-31T16:00:00.001-04:00

O’Neill KP, Godwin HW, Jiménez-Esquilín AE, Battigelli JP. 2010. Reducing the dimensionality of soil microinvertebrate community datasets using Indicator Species Analysis: Implications for ecosystem monitoring and soil management. Soil Biology & Biochemistry 42: 145-154.

These authors used a dataset of soil microarthropods to evaluate a method for identifying indicator species for ecosystem monitoring. The method centres on the Indicator Value (IV) of a species, a number that integrates the degree of uniqueness to a place of a species and the abundance of that species within a given habitat. A high IV value indicates both high information content and a high probability of being sampled. The IV is apparently robust to differences in site number and species absolute abundances, and provides a single value for evaluating observed or expected changes in an ecosystem. Indicator species, furthermore, integrate habitat conditions over their lifespans, in contrast to measures of chemical and physical parameters that capture a snapshot of an ecosystem.

The basic evaluation approach here was to identify indicator species along a clear environmental gradient from meadow to forest in West Virginia. The habitat was divided into three zones, with an edge patch between the open meadows and closed-canopy forest. Near-surface soil cores were collected from each zone every month from April 2004 to April 2005 (n = 180), using the top of the mineral soil as the reference depth. Microarthropods were extracted in a modified Macfadyen funnel with a strong and increasing temperature gradient, into 70% ethanol.

Diversity measures, including Simpson’s and Shannon indices, were based on counts of individuals identified to family level (suborder for Acari). Differences between sites were analyzed by 2-way repeated measures ANOVA and Principle Components Analysis, after rare taxa (those that occurred in less than 10% of samples) were removed; rare taxa are extremely unlikely to be identified as indicator species.

Calculating IV for each taxon, regardless of the taxonomic resolution, provides large advantages in labour time and taxonomic expertise, as many microfauna are very difficult to identify to genus or species. These authors state that enumeration of a single sample required more than 1 hour of a trained taxonomist’s time. In studies such as this one, there are further advantages of IV associated with its robustness in the face of many zero measurements (i.e. taxa absent from samples) and the general messiness of these kinds of datasets. However, the ISA approach is intended for 2-stage studies, where an intensive initial survey identifies indicator species (taxa), and later long-term monitoring ignores other species. For studies specificially designed to address biodiversity, such as my own, excluding rare taxa would not be beneficial, and there may be no easy escape from time-consuming morphotaxa sorting.

I have spoken with Dr. Battigelli, the trained taxonomist in this study. He has indicated that while this IV-based approach may not be appropriate for my own work, it nonetheless demonstrates the types of analyses that can be conducted with soil invertebrates identified to middle taxonomic levels. He has assured me I could probably be trained to identify Collembola to Family and Acari to Suborder in a matter of a few days, and he would be interested in futher studies of collected soil invertebrates based on interesting patterns that emerge at these taxonomic levels.

Harding et al. 2001

2010-03-25T10:10:00.000-04:00

Harding RJ, Gryning S-E, Halldin S, Lloyd CR. 2001. Progress in understanding of land surface/atmosphere exchanges at high latitudes. Theoretical and Applied Climatology 70: 5-18.

These authors review and discuss the implications of studies based in two international projects in northern Europe. WINTEX was a large study examining the effects of snow cover and long nights in winter on high-latitude ground-atmosphere exchange processes, while LAPP was an independent but complementary study examining most of the same processes in a range of high latitude sites during spring and summer.

Snow cover plays a major role in Arctic exchange processes. The high albedo of snow reflects much of the incident solar radiation, and insulates the frozen ground below, prolonging the period of snow cover to upwards of 9 months in the year in many places. Where vegetation is tall, such as in the boreal forest, the low solar angle reduces the effective net albedo of the landscape, allowing sunlight to warm the dark trees and speed springtime melting. This study mentions the importance of snow-surface aerodynamics, though it appears there is little solid information on this complex topic.

Snow melt is the major hydrological event of the year in much of the Arctic. The combination of frozen soils, very low evaporation rates, and often flat terrain means much of the Arctic is very wet or saturated while annual precipitation rates are consistent with arid or semi-arid conditions. These areas are the classic tundra systems, with abundant shallow lakes and ponds and very wet high-organic soils.

Differences in snow-surface dynamics and the timing of snowmelt create an extremely heterogeneous landscape, particularly in the vicinity of the northern treeline. There are often very large temperature and air-flow differences between patches of trees and adjacent lakes or clearings, which greatly complicate attempts to model the carbon dioxide emissions (for example) of such areas. Much of this paper is a series of evaluations of some of the models that have been applied to this region. In general, more sophisticated models that can take some of the extreme variability into account perform better than models that cannot account for differences in snow depth or insulating properties. However, this paper makes it clear that current modelling efforts still leave much to be desired in terms of predicting Arctic heat budgets and biological responses.

Water storage is also very difficult to model, and has large and variable impacts on other parts of the system. There appears to be large and unpredictable year-to-year variation in water storage and transport at the scale of catchments and basins, and the importance of soil water in controlling biological processes such as the decomposition of organic matter is large. Runoff matters, even on very gentle slopes.

This paper provides a useful overview of large-scale processes and attempts to understand these processes in the Arctic.

Martin and Rygiewicz 2005

2010-03-11T16:55:00.000-05:00

Martin KJ, Rygiewicz PT. 2005. Fungal-specific PCR primers developed for analysis of the ITS region of environmental DNA extracts. BMC Microbiology 5:28.

These authors designed new primers for PCR and related molecular biology investigations of soil fungi, especially mycorrhizae. These are very diverse organisms, and, because of the commonalities between fungal and plant DNA, studies of fungal samples closely entwined with plant tissue as in root-associated mycorrhizae can be very complex. The new primers were designed to produce a range of PCR products suitable for techniques such as qPCR, Length-Heterogeneity PCR (LH-PCR) and T-RFLP analyses.

The primers as a suite were designed around a nested approach, with new outer primers amplifying a long DNA sequence of approximately 1000bp, and later primer pairs amplifying regions within that long sequence. Most inner primer pairs generate products of approximately 500bp length.

These authors also used a different DNA extraction method, based on xanthogenate and Tween (X/T) that involves little to no tissue grinding, compared to the standard method based on CTAB. The X/T method preferentially extracts fungal DNA from cells on the outside of particles, for example fungal cells not penetrating plant roots. This reduces the amount of plant DNA and associated plant-derived compounds in resulting extracts. Combining the two techniques may allow for some interesting studies of fungal micro-ecology.

This paper’s novel primers should be more specific and more useful to my own qPCR studies of Arctic soil microbes, including soil fungi. The methods section detailing some of the decisions made and considerations involved in primer design will also be useful.

Philippot et al. 2002

2010-02-24T17:15:00.001-05:00

Philippot L, Piutti S, Martin-Laurent F, Hallet S, Germon JC. 2002. Molecular analysis of the nitrate-reducing community from unplanted and maize-planted soils. Applied and Environmental Microbiology 68: 6121-6128.

These authors applied molecular techniques including PCR, RFLP, and sequencing to the study of soil bacteria relevant to crops. Dissimilatory nitrate reduction, the process that converts NO3- to NO2-, is widespread in prokaryotes, with the activity described in alpha, beta, and gamma Proteobacteria, gram-positive bacteria, and some archaea. There are two described enzymes that catalyze the reaction and provide energy to the organism; these authors focused on the membrane-bound protein, specifically one subunit that includes a distinctive set of components. Their approach was to design primers for a well-conserved region of the gene narG that amplify a 650bp region, and then subject the PCR product to cloning, RFLP analysis, and sequencing.

Community structure and diversity was compared between pots planted with maize versus unplanted controls. Maize (Zea mays) is a plant that facilitates gas diffusion in its roots under oxygen-stress soil conditions; this creates an aerobic region in the rhizosphere distinct from anaerobic conditions further from roots. While diversity, as measured by standard indices, did not differ between planted and unplanted soils, the structure of the communities did change, with numerous RFLP phylotypes found in only one or the other treatment. This suggests a role of rhizosphere conditions, likely involving both oxygen and root exudates, in selecting for particular groups of microorganisms.

Nitrate reduction occurs primarily or possibly only under aerobic conditions. The microbial cell gains energy from dissimilatory reduction of nitrate, and if it occurs in the rhizosphere, the plant may gain a readily-accessible form of nitrogen in the form of nitrite. Denitrification, the process that shuttles nitrogen atoms from nitrite to gaseous forms such as N2O or N2, can occur under a range of oxygen conditions, including aerobic, thus denitrifiers in the rhizosphere may compete with plant roots for nitrite. The fate of nitrite produced by dissimilatory nitrate reduction can also be to ammonium, though this appears to be rare in soil and more common in vertebrate guts and digested sludge, two environments typically lacking in oxygen.

This paper provides some molecular tools for my own studies of nitrogen dynamics in soils, especially the sequences of the degenerate primers. In addition, it provides some clarification of parts of the remarkably complex soil-nitrogen cycle.

Siciliano et al. 2007

2010-02-24T09:55:00.000-05:00

Siciliano SD, Ma W, Powell S. 2007. Evaluation of quantitative polymerase chain reaction to assess nosZ gene prevalence in mixed microbial communities. Canadian Journal of Microbiology 53: 636-642.

These authors examined the usefulness of qPCR in studying populations of soil bacteria, especially denitrifiers using the gene nosZ that codes for nitrous oxide reductase. This enzyme catalyzes the final reaction in the process of denitrification, converting N2O to N2. Normally, it is expressed only in severely anaerobic conditions, as it allows the use of N2O as the terminal electron acceptor during metabolism.

There are a number of factors that control the efficiency of PCR in quantitative PCR applications. The efficiency is a major component of the calculations that allow qPCR to estimate gene copy numbers in samples and thus to be used to examine population dynamics of non-culturable microorganisms from environmental samples. Of particular importance is consistency of efficiency between the amplification of the standard DNA template and the amplification of all templates in the unknown samples. Variation between the standard and the unknowns can lead to severe under- or over-estimation of target populations, while variation in efficiency between different templates within the unknown samples can lead to misestimations of relative proportions of organisms.

These authors evaluated the efficiency of qPCR in a range of experimental templates, and in a range of combinations simulating mixed populations. Little variance in efficiency was found, and this variance was not associated with genetic distance from a reference organism. The experimental design did not allow a direct examination of the influence of the geographical differences in the sources of the test sequences (Arctic, temperate-grassland, Antarctic), but this lack of association with the reference organism does indicate low or no variation among PCR efficiencies associated with some other variable.

The influence of varying PCR efficiencies among templates within a sample becomes less severe as the number of different templates rises. In a typical soil sample with perhaps 1000 different templates, no one template can utterly dominate amplification by outcompeting for primers, thus the resulting mix of amplicons at the end of 40 rounds of PCR will most likely be representative of the population mixture in the environment.

This paper is of obvious high utility to my own work, not least because the individual machine used to perform qPCR is the same individual machine that I will be using. For this and other reasons, this paper was suggested to me, repeatedly. Future reference to this paper, when I am developing my methods and when I am writing up the next paper or two, seems likely.

Dandie et al. 2007

2010-02-22T15:10:00.001-05:00

Dandie CE, Miller MN, Burton DL, Zebarth BJ, Trevors JT, Goyer C. 2007. Nitric oxide reductase-targeted real-time PCR quantification of denitrifier populations in soil. Applied and Environmental Microbiology 73: 4250-4258.

These authors examined the responses of two major components of the denitrifying bacteria fraction of soil bacteria to the addition of labile carbon (glucose) under denitrifying conditions. Denitrification is presented as a four-step process, with enzymes responsible for shuttling nitrate to N2 via nitrite, nitric oxide, and nitrous oxide. In this study, one of the enzymes responsible for the reduction of NO to N2O, cNOR, was examined using primers optimized for two different groups of denitrifying bacteria. This gene is found only in denitrifiers, unlike another enzyme, qNOR, found in many microorganisms and associated with detoxification, rather than utilization, of dangerous nitric oxide.

Primers for qPCR are presented in a table. Specific primers for the two variants of cNOR were developed in this study for use with SYBR green-based qPCR. 16s rRNA sequences were also studied, to examine the total population of soil bacteria; for these qPCR reactions, the TaqMan primers-plus-probe system was used, based on oligonucleotides published by Suzuki et al. 2000.

Two experiments were carried out. In the first, a preliminary experiment to establish the utility of qPCR in this area was based on inoculating soils with cultures of bacteria of known cell density, followed by qPCR evaluation of those soils. Under most conditions qPCR performed well, though at low cell densities of some genera of bacteria the signal was not distinguishable from the background noise also associated with sterilized soil. The second experiment forms the main body of work of this paper, and is an examination of the population dynamics of soil bacteria, divided into the hierarchical categories “denitrifiers” and “all bacteria”, under denitrifying conditions and with varying levels of added labile carbon in the form of glucose solutions in distilled water. In the second experiment, soil nitrate was maintained at a high level, to ensure sufficient raw material for detectable denitrification activity. As N2O accumulation was one of the measures of activity, nitrous oxide reductase activity that would reduce N2O to N2 was inhibited by maintaining an atmosphere of 10% acetylene in culture jars. Soils were maintained at 70% WFPS to encourage denitrification.

Total microbial biomass was also measured, using the CHCl3 fumigation-extraction technique. While cNOR sequences are almost certainly restricted to one copy per genome, 16s rRNA sequences may range in copy number up to 15 per genome, thus estimates of bacterial populations by qPCR of 16s may have a large error associated with it. Fumigation-extraction captures all carbon associated with cells, thus contributions by archaea and fungi will not be found by molecular methods such as 16s qPCR that are specific to bacteria. However, in this study, estimates of total bacterial population by the two methods were well correlated, with r2 = 0.69.

Denitrification occurred in this study. Soils treated with additional glucose showed greater depletion of nitrate, as expected when denitrifiers increase their activity in response to a food supply and conditions already favour denitrification. These authors provide two possible mechanisms, non-mutually-exclusive, that could lead to increased denitrification activity under added glucose. First, the population of denitrifiers could expand, through both additional cell replication and activation of dormant cells. This would increase the proportion of the bacterial population composed of denitrifiers. Second, the total population of soil organisms could increase, leading to increased respiration, a decrease in oxygenation, and establishment of anaerobic conditions more favourable for denitrification. This would not necessarily change the proportion of the population composed of denitrifiers. In this study, denitrifiers increased their proportion of the population as measured by comparative qPCR from less than 1% to about 2.4% of cell numbers.

This change in population components is central to the approach using qPCR advocated in this paper. As these authors state:

“Although absolute numbers may not be achievable, gross differences and changes in population size are still detectable. The differences observed between the two denitrifier populations studied are then real differences in the responses of these populations to the conditions tested.”

This general approach of examining relative changes in populations is applicable to a very wide array of studies of environmental microbiology, including my own planned studies in which the environmental factor under examination is biogeographical (i.e. latitude) and the functional diversity response is in terms of greenhouse gas cycling."

This paper is of great value to my studies. The qPCR methods are directly applicable, for example the primers presented here will be useful if I decide to examine multiple components of the denitrification pathway. The approach, as described above, is also useful. And the reference list is composed almost entirely of papers I am surprised I have not yet found in my literature searches.

Liptzin 2006

2010-02-19T16:45:00.001-05:00

Liptzin D. 2006. A banded vegetation pattern in a High Arctic community on Axel Heiberg Island, Nunavut, Canada. Arctic, Antarctic, and Alpine Research 38: 216-223.

This author attempted to explain the observation of banded vegetation on a slope that lacked the usual factors that generate such patterns. In temperate and tropical locations, banded vegetation, also known as “tiger stripes”, forms on shallow slopes in dry areas with a consistent direction of water flow. Plants at a position on the slope increase water retention and facilitate further colonization by plants. Similarly, some locations experience consistent wind direction carrying sea spray that kills trees at some positions. In cold environments, patterned ground from cryoturbation on shallow slopes can also lead to banded vegetation. However, the study site in this paper lacks all of these features, including cryoturbation despite the presence of permafrost within 50cm at most locations.

Some aspect of soil properties is the obvious explanatory hypothesis, which this author explores after describing the transects measuring plant diversity and the soil pits used to examine soil properties. In general, features that would normally be expected to influence plant diversity and abundance such as soil moisture or exchangeable cation levels, had no significant impact in the various statistical tests employed in this study. However, soil type did have some effect, as a few species of plants were found only on sandy soil, and nitrogen levels were negatively correlated with species richness.

The discussion section of this paper is an excellent example of a chain of logical reasoning working through a series of potential explanations. While this paper is interesting, it’s only relevant to my own studies in a narrow area around potential starting points in looking for explanations for whatever patterns I may find in my biogeography studies in 2010. However, this paper seems remarkably suitable as an introduction to the basics of modern soil science research, and may be relevant to my not-quite-mothballed interest in an undergraduate course about the current state of the scientific literature.

Klotz and Stein 2008

2010-02-18T14:00:00.002-05:00

Klotz MG, Stein LY. 2008. Nitrifier genomics and evolution of the nitrogen cycle. FEMS Microbiology Letters 278: 146-156.

These authors review the role of nitrifying microorganisms in the current nitrogen cycle, and their evolution and the emergence of biological nitrogen cycles in early Earth history. The current global nitrogen cycle has changed considerably in the past decades, due to the large increase in nitrogen in the cycle due to human activities. The early-Earth nitrogen cycle was probably mostly driven by abiotic processes. After the development of an oxygen-rich atmosphere, nitrogen cycling was almost entirely biotic, with most key processes driven largely or entirely by bacteria and archaea. In the last few decades, the anthropogenic abiotic processes of fertilizer production and fossil-fuel combustion combined with increased cultivation of N2-fixing crops, has transformed the global nitrogen cycle.

As presented in this paper, there are two lobes to the global nitrogen cycle. N2 gas in the atmosphere is fixed to NH3, by nitrogenase in bacteria and archaea, by the Haber-Bosch industrial process, and (in small quantities) by hydrothermal vents. The process of nitrification converts this ammonia to nitrite/nitrate. Nitrite/nitrate are returned to the atmosphere as N2 through denitrification, with production of N2O under weakly anaerobic conditions. The other lobe of the cycle is a “short circuit” that avoids the atmospheric N2 pool and cycles nitrite/nitrate back to ammonia through the processes of ammonification and through production and decomposition of organic matter containing nitrogen. There is another, minor short circuit, as “anammox”, anaerobic ammonia oxidation, returns ammonia to N2 directly.

This “mini review” focuses on the nitrification portion of the cycle. The first step, oxidation of ammonia to hydroxylamine (NH2OH) is carried out by ammonia oxidizing bacteria, abbreviated AOB. There are many acronyms in this paper, reflecting the many acronyms in the existing literature regarding global biogeochemical cycles. NOB are nitrite oxidizing bacteria, and they take the oxidized products of AOB, especially nitrite through to nitrate. Anammox bacteria, on the other hand, may run the same net process of NH3 to NO3- directly, without collaboration with other cells.

The discussion of the plausible evolutionary scenarios in this paper is interesting but not particularly relevant to my current research. This discussion focuses on the relative timing of major events, such as the emergence of nitrification, complete and incomplete denitrification, an oxygenated atmosphere, and nitrogen fixation. These factors interact with each other, creating conditions favourable or not to the evolution of each other and of possible detail shifts within.

The description of the role of hydroxylamine produced by early nitrifiers in stimulating evolution of metabolic pathways responsible for its detoxification initially reads as speculation, but a long and detailed description of the ways in which the various components of those metabolic detox enzymes and pathways function provides plenty of support for the arguments. One aspect of this discussion is that some enzymes are currently misclassified, and that very similar enzymes in different organisms have different names reflecting different ultimate functions rather than the usual (and preferred) enzyme naming scheme that reflects proximate function.

In the discussion concerned with anthropogenic climate change and nitrogen dynamics, especially in soils and ocean waters, interactions with methane are briefly considered. This is based on the observation that methanotrophs are often also ammonia-oxidizers, operating under a budget of consumption of both molecules that shifts as ammonia from fertilizer is added to the system.
Of greatest relevance to my current work is the section describing the gene ncyA. This encodes an enzyme (nitrosocyanin) involved in the pathway from ammonia to nitrite, and has only been found in AOB to date, as opposed to NOB, anammox, or heterotrophs; it seems to be involved in the chemistry of obligate chemolithotrophy as expressed by AOB. It seems likely the enzyme binds and reduces NO, a highly toxic intermediate in ammonia oxidation. The regulatory region adjacent to the gene is also suggestive of roles in this metabolic pathway, and regulation is linked to concentration of various nitrogen-with-oxygen compounds.

This review is very useful to my current research. This paper and the major references in it will be key to constructing a diagram of the complex nitrogen transformations occurring in soils, which will allow targeted hypothesis generation and testing regarding the communities and processes in the soils I am studying.

Li et al. 2009

2010-02-18T10:20:00.001-05:00

Li X-R, Du B, Fu H-X, Wang R-F, Shi J-H, Wang Y, Jetten MSM, Quan ZX. 2009. The bacterial diversity in an anaerobic ammonium-oxidizing (anammox) reactor community. Systematic and Applied Microbiology 32: 278-289.

These authors studied the bacterial community that developed inside a bioreactor running on sewage sludge under anaerobic conditions. Like Lim et al. (2008), the main focus of this study was in the applications of ammonia-oxidizing bacteria (AOB) to water treatment facilities. The expected chemistry of anaerobic ammonia oxidation catalyzed by microorganisms (“anammox”) includes the use of nitrite as the electron acceptor in a near-one-to-one ratio with the consumption of ammonia or ammonium. The energy derived from this process is used by the cell to fix CO2, thus making these organisms autotrophs. This alters the underlying stoichiometry slightly, as some nitrite is diverted to CO2 fixation rather than ammonia oxidation.

The study of anaerobic AOB is still quite new, with the five described genera of such organisms all named with “Candidatus” prefixes, indicating recent species descriptions. All are in one group (taxonomic level unknown), the Brocadiales, within the phylum Planctomycetes. Aerobic AOB are in other groups, and include some species within the genus Nitrosomonas in the Beta-Proteobacteria that are capable of limited ammonia-oxidizing activity under anaerobic conditions, and can apparently survive long periods without oxygen.

These authors did not develop novel primers for PCR or qPCR in this study. Instead, they used published primer sets; I gather they did not use the TaqMan double-dye system for qPCR, as no mention of probes is made. The target genomic sequences were portions of the 16s rRNA gene, using E. coli as a standard. Oddly, the overall procedure included normal PCR, followed by cloning and insertion into plasmids, followed by qPCR of plasmid DNA containing the 16s sequences. It is unclear to me exactly why this was done, though later in the paper there are a few sequence-based phylogenetic trees that might have been based on sequences derived from this cloning procedure. In any case, the qPCR did provide informative results regarding the composition of bacterial groups within the reactor.

Of the sequences identified, the great majority were unlike cultivated organisms, highlighting the utility of these techniques in studying environmental samples. AnAOB produced approximately 16% of sequences, with aerobic AOB less than 1%. Non-AOB in three phyla constituted the majority of sequences, including 38% Chlorobi, 21% Chloroflexi, and 7% Bacteriodetes. These are filamentous heterotrophic bacteria, and appear to be closely associated with the granules that formed in the reactor solution after a few months. These authors suggest further research on the ecophysiology of these groups to answer questions regarding energy and material cycles within these systems.

In addition to 16s sequences, the hzo locus was also studied. This is a gene that produces an enzyme that catalyzes the oxidation of hydrazine (rocket fuel; N2H4) to N2 gas. No mention is made of the possibilities for N2O production or consumption in this process. The gene is restricted to AnAOB only, or at least that is the inference based on the observation that hydrazine is a unique intermediate molecule of the anammox process.

This study provides a useful example of the combination of qPCR and molecular-phylogenetic approaches in studying a microbiological system. Applied together, the two approaches allow the extraction of useful information regarding taxonomic diversity, both richness and evenness, among functional groups of organisms.

Lim et al. 2008

2010-02-17T14:20:00.000-05:00

Lim J, Do H, Shin SG, Hwang S. 2008. Primer and probe sets for group-specific quantification of the genera Nitrosomonas and Nitrosospira using real-time PCR. Biotechnology and Bioengineering 99: 1374-1383.

These authors developed precise primer and probe sets for TaqMan-based quantitative PCR to examine ammonia-oxidizing bacteria (AOB) associated with wastewater treatment facilities. These molecular tools have very low rates of false-positive and false-negative errors associated with them, and will be useful primarily to work on improving the nitrogen-removal capacity of wastewater treatment. However, AOB are nearly ubiquitous, such that these molecular tools will also be useful to a wide variety of less-directly-applied studies.

These authors purchased eight strains of nitrifying AOB commonly found in water treatment plants, eight non-nitrifying bacteria also commonly found in such plants, and collected, identified, and purified seven strains of nitrifying bacteria directly from a pair of water treatment plants in operation. Using published sequences of the 16s rRNA genes of these organisms, sets of primers and probes were constructed.

The development and evaluation of these primer/probe sets followed two basic procedures: first, “in silico” evaluation of potential primer and probe binding sites, and calculation of potential mismatches in various combinations. For example, a set developed for one organism may also be highly likely to amplify a related organism, reducing specificity of the assay. Second, sets were optimized for PCR conditions and trialed with the often-variable real sequences derived from culture collections or field samples. Iterating between these two processes allowed a series of final best-fit sequences to emerge, that have high specificity and low failure rates.

As a template for developing very good molecular tools, this paper provides some excellent advice regarding qPCR primer development. I will be using a different system that does not include probes, simplifying some steps of this process, but the basic pattern of invention, computer evaluation, wet-lab evaluation, further computer evaluation, and refinement will still be useful.

Himmelheber et al. 2009

2010-02-17T09:05:00.001-05:00

Himmelheber DW, Thomas SH, Löffler FE, Taillefert M, Hughes JB. 2009. Microbial colonization of an in situ sediment cap and correlation to stratified redox zones. Environmental Science & Technology 43: 66-74.

These authors previously studied the changes in geochemistry associated with the common practice of adding a sediment cap to cover contaminated sediments at the sediment-water interface. Such caps are commonly clean sand, with the underlying idea being the layer of sand provides a transport barrier to various contaminants moving through the system by diffusion. Sediment geochemistry, like soils, includes layers of redox conditions generated by both biotic and abiotic factors. These zones of chemical conditions migrate upwards when a sediment cap is added; not surprising considering the effect the cap has on diffusion of oxygen and other chemicals important for redox considerations.

This study shows that the microbial populations also migrate upwards when a cap is added. The primary concern here seems to be the effect this population shift may have on the transport and decontamination of such pollutants as are often found in the river-bottom sediments of the eastern USA. The primary effect is likely positive: populations of bacteria and archaea in sediments will metabolize, mineralize, and generally detoxify most compounds moving up from the sediments to the cap. A few classes of contaminants, however, may not be decontaminated and it is possible their transport and release into the water column may be accelerated by these microbes.

There are two key parts of the methods of this paper that interest me. First, the microbial populations were analyzed by a range of techniques including real-time quantitative PCR (qPCR). The procedure of primer design, evaluation, and data interpretation looks very similar to what I will be attempting with my own samples. Second, diversity estimates for the various strata within the sediments, derived from qPCR data, includes the use of the statistical technique Canonical Correspondence Analysis. This allows direct testing of hypotheses regarding the relationship between environmental parameters, in this case depth below surface, and estimates of biodiversity such as the Shannon-Weiner index.

Kellman and Kavanaugh 2008

2010-02-12T17:15:00.000-05:00

Kellman L, Kavanaugh K. 2008. Nitrous oxide dynamics in managed northern forest soil profiles: is production offset by consumption? Biogeochemistry 90: 115-128.

These authors measured surface flux and subsurface profiles of N2O at a number of paired sites in the managed forest of Nova Scotia. Half of the sites were clear-cut harvested three years before the study, the other half more than 50 years previously. Climate factors such as air temperatures and solar radiation were consistent across the study area. Fluxes and profiles were measured periodically through a 9-month snow-free period in 2005, from early March to late November.

Surface fluxes were measured by pulling samples into evacuated containers from chambers mounted on permanent collars. Similarly, profiles were measured by sampling from permanent probes buried in the walls of soil pits. Actual measurement of gas concentrations were in the laboratory using a gas chromatograph system.
The soil probes consist of 50cm PVC tubes, covered with a “water resistant porous membrane” (could they be using Gore-tex?) and buried in the walls of pits at depths of 0, 5, 20, and 35cm, with 0 at the mineral soil-organic layer interface. This provides a 50cm-long sampling space at four depths, replicated across 40 sites.

The relationship between profile N2O concentrations and surface flux was almost always non-significant. These authors attribute this lack of correlation to consumption of N2O in the soil profile. In contrast, the studies that have linked CO2 profiles to surface flux have relied on the (probably true) assumption that CO2 is not consumed in the soil, and moves through diffusion in a manner that can be predicted from soil physics. N2O profiles that include regions of consumption are complicated by the biological and chemical factors that control production and consumption, as well as movement. All of this leads to a disconnection between soil N2O cycling and surface-atmosphere exchange.

This paper should almost certainly be included in the introduction, methods, and/or discussion section(s) of my pits & probes manuscript. This is one of the few studies I have found that examined N2O in soil profiles; most others appear to focus on CO2 or in some cases the biogeochemistry of CH4.

Lamb et al. 2006

2010-02-11T17:15:00.001-05:00

Lamb EG, Cahill JF, Dale MRT. 2006. A nonlinear regression approach to test for size-dependence of competitive ability. Ecology 87: 1452-1457.

These authors describe a statistical analytical technique, “nonlinear regression”, that potentially provides more information than linear regression techniques including ANCOVA. The basic linear regression formula includes the intercept and the slope as parameters; nonlinear regression adds an exponent parameter. In this paper, these parameters are referred to as k1 (intercept), k2 (slope), and k3 (exponent), in the formula

y = k1 + k2x^k3

where y is the response variable and x is the explanatory variable.

These authors tested their technique on three example datasets from previous studies. The first example is most thoroughly examined, and involves a plant competition experiment. One key feature of all of the example analysis is the dataset must be paired, such that each data point on the x axis corresponds to a partner data point on the y axis. In the plant competition example, individual plants that did not experience competition are paired with individuals that did, because each pair of plants was grown in a communal pot that was treated at the pot level with manipulations such as fertilizer application. The continuous dataset is plant size, measured as the absolute gain in mass over the course of the growing season.

The exponent parameter k3 can take on any value between negative and positive infinity, to describe curves that may be accelerating, saturating, or straight. At values of 1 or -1, k3 is not informative as a parameter, and should be discarded from the model. This analysis is based on a model-building and model-testing technique, where models with various values for the three parameters are tested against null models and each other in an iterative fashion to find the model that best fits the data.

This approach is likely to be useful in the analysis of some of the data collected at Alexandra Fjord in 2009.

Michelsen et al. 1999

2010-02-04T23:25:00.000-05:00

Michelsen, A., Graglia, E., Schmidt, I.K., Jonasson, S., Sleep, D., and Quarmby, C. 1999. Differential responses of grass and a dwarf shrub to long-term changes in soil microbial biomass C, N and P following factorial addition of NPK fertilizer, fungicide and labile carbon to a heath. New Phytologist 143(3): 523-538.

These authors measured the responses of two tundra plants, one graminoid and one shrub, to additions of fertilizer, labile carbon, and a fungicide over four years in a heath in subarctic Sweden. The underlying hypothesis is that differences in growth strategy between these two groups of plants would be reflected in their responses to changes in microbial biomass, also measured by these authors.

Molecules inaccessible to plants by virtue of being incorporated into microbial cells in soils dominate their respective pools. For example, in this study, microbial N concentrations were up to 50 times higher than free, inorganic N such as ammonium. Similar ratios were found for C and P. Rapid incorporation of these nutrients into microbes therefore constitutes competition between microbes and plants. Consistent with the variety of ecological and growth strategies that plants have evolved interacting with each other and with the abiotic features, plant strategies impact their interactions with microbes and how they react to such competition.

As predicted, the graminoid Festuca ovina responded rapidly to changes in nutrient levels. Added N from fertilizers led to increased ground cover by this grass, while added labile C, in the form of sugar, led to decreased ground cover, presumably due to a rapid increase in microbial biomass and associated increased microbial uptake of soil N and P. In contrast, the shrub Vaccinium uliginosum did not appreciably react to changes in nutrient levels, consistent with a long-view growth strategy and successful escape from severe competition with soil microbes. This may have been mediated by the mycorrhizal associations the shrub has but the graminoid lacks.

This paper, interestingly, does not cite any of the work by Wardle or Nilsson, the authors who severely criticized the earlier work by this research group (Michelsen et al. 1995).

Wardle and Nilsson 1997

2010-02-03T17:47:00.001-05:00

Wardle DA, Nilsson M-C. 1997. Microbe-plant competition, allelopathy and arctic plants. Oecologia 109: 291-293.

These authors critique Michelsen et al. (1995), a study that came to several important conclusions regarding the interactions between Arctic plants and the soil microbial communities. This is a very negative review of that paper, in which these authors question almost all of the conclusions of Michelsen et al. (1995).

These authors make two main criticisms. First, they question the measures of soil microbial activity used by the earlier paper. Second, they question the conclusions regarding the allelopathy of Empetrum hermaphroditum. Soil microbial activity was measured by Michelsen et al. (1995) in two ways: soil respiration, and soil ergosterol content. Neither approach is necessarily informative about one of Michelsen et al.’s (1995) main claims, that soil microbial biomass was increased by the addition of plant leaf extracts. There are a number of studies, many of them with Nilsson as a co-author, in which a lack of association between microbial biomass and soil respiration was demonstrated. Furthermore, ergosterol is presented by Michelsen et al. (1995) as an indicator of fungal biomass, but previous work by Newell and colleagues (e.g. Newell and Fallon, 1991; Newell 1992) showed that ergosterol is not a reliable indicator of biomass nor is it useful as a proxy measure of soil fungal activity; the ratio of ergosterol to fungal biomass is highly variable.

From the reference list in this short paper, it appears that Wardle and Nilsson had, by early 1997, completed a considerable body of work regarding the allelopathic and other ecological interactions of E. hermaphroditum in sub-arctic environments. The conclusion by Michelsen et al. (1995) that the chemicals released by this plant have a greater impact on microbial communities than potential surface-plant competitors is not supported by this work by Wardle, Nilsson, and their colleagues.

The conclusion of Michelsen et al. (1995) that currently has the most direct bearing on my own work is that key plant traits often possessed by prostrate shrubs in tundra ecosystems such as a high root: shoot ratio and storage of nutrients such as nitrogen in the roots allow those plants to escape from or outcompete soil microorganisms. This conclusion was not addressed by these authors, but given the devastation inflicted upon the other conclusions, my confidence in the utility of Michelsen et al. (1995) in addressing issues of interactions between Cassiope tetragonal and soil microbial communities has been shaken.

Michelsen et al. 1995

2010-02-03T17:42:00.001-05:00

Michelsen A, Schmidt IK, Jonasson S, Dighton J, Jones HE, Callaghan TV. Inhibition of growth, and effects on nutrient uptake of arctic graminoids by leaf extracts – allelopathy or resource competition between plants and microbes? Oecologia 103: 407-418.

These authors conducted an experiment to examine the potential allelopathic effects of plant leaf extracts among a few species of arctic tundra plants. Three arctic/subarctic plants suspected of releasing phytotoxic compounds upon their competitors including Cassiope tetragonal were harvested and their leaves and branches ground up to make leaf extracts. Three species of arctic graminoids (Carex bigelowii, Festuca vivipara, Luzula arcuata) were then treated with these extracts while growing in either sterilized or non-sterilized soil in greenhouses in southern Sweden. All of the graminoids, the soil they grew in, and two of the leaf extract-providing plants came from a montaine subarctic hillside in northern Sweden; the third leaf extract came from Betula pubescens ssp. tortuosa, a birch, were collected from individuals growing near the tree line at 450m altitude near Abisko Scientific Research Station, above the Arctic Circle in a subarctic ecosystem.

The experimental design was factorial, with two soil types (sterilized vs. non-sterilized) and four leaf-extract treatments including a control of distilled water. In addition to growth of the graminoids, measurements were made of the chemicals in the leaf extracts and soils, nutrient uptake by excised roots, soil ergosterol content, and soil respiration. Excised roots take up nutrients in a manner directly correlated to the nutrient-limitation status of the plant; more phosphorus-starved plants, for example, have roots that more rapidly take up phosphorus when offered. Soil ergosterol content is a measure of fungal biomass, while soil respiration was taken as a measure of total microbial activity.

Sterilized soil had higher extractable nutrients, probably as a result of the breakdown of microbial cells during autoclaving. Nitrate levels were negligible, both in soils and in leaf extracts; nitrate was a component of the dilute nutrient water used to maintain the plants while growing. Some mycorrhizae were found, but they covered less than 1% of the roots in non-sterilized soil, and had no impact on other measured parameters.

The highest growth of all three test graminoids was recorded in sterilized soil with no extract added (i.e. distilled water added instead of leaf extract solution). Plants growing under these conditions experienced no inhibition from the materials of other plants, and did not compete with soil microbes for nutrients, at least during the early stages of the experiment before microbes recolonized the sterilized soils. Recolonization was much faster by prokaryotes than by fungi, as measured by the contrast in soil respiration rates and soil ergosterol contents. Recolonization also varied between leaf extract treatments, with a negative correlation between microbial activity and plant growth; strongly growing plants were able to outcompete colonizing microbes, while poorly growing plants were further inhibited by colonizing and rapidly growing microbes.

All three leaf extracts significantly reduced growth of all three graminoid species. However, it is not clear that allelopathy alone was responsible for this effect. The results of this study indicate that competition between plants and microbes also played a major role. In particular, the components of the leaf extracts, especially labile carbon and (in the case of the Betula extract) phosphorus, appeared to stimulate the microbial community, increasing competitive pressure on the plants. Added nitrogen, for example, appears not to have much benefited the plants, as their roots were N-limited when grown in non-sterilized soil even though the leaf extracts included high concentrations of inorganic nitrogen.

The susceptibility of plants to this combined effect of allelopathy and microbial competition probably varies by species. Plant traits of particular importance are probably the root: shoot ratio, in which plants with more robust roots are less harmed, and the storage of nutrients such as nitrogen in plant roots, in which plants with a growth strategy favouring nutrient storage rather than immediate use are less harmed. Such traits appear to be widespread among the dominant plants of the Arctic tundra, including Cassiope tetragonal and probably other prostrate shrubs. Many of these plants may form associations with mycorrhizal fungi, which provide some protection against microbial competition.

This paper is directly relevant to the discussion section of my current-high-priority Pits & Probes manuscript. The patterns of soil respiration and microbial GHG activity under some of the lowland communities are consistent with successful competition against soil microbes by Cassiope tetragonal and possibly Salix arctica plant roots.

This paper was critiqued quite harshly by Wardle and Nilsson (1997).

Moffat et al. 1990

2010-02-03T17:40:00.000-05:00

Moffat AJ, Johnston M, Wright JS. 1990. An improved probe for sampling soil atmospheres. Plant and Soil 121: 145-147.

These authors describe a new design of soil gas probe they produced to examine soil O2 levels, especially at landfills and other potentially polluted sites. The design is fairly complex, with moving parts. Essentially, the probe is driven into the soil to the desired depth, and then lifted slightly to create a void below the tip. The top of the probe is then rotated and tapped to open the tip, and a gas sample can be drawn from a tube on the side. These authors tested this probe by measuring O2 concentrations in a column of sand that had been flushed with pure N2; anoxic conditions were recorded in the deeper parts of this column.

Farías et al. 2009

2010-02-03T17:38:00.000-05:00

Farías L, Fernández C, Faúndez J, Cornejo M, Alcaman ME. 2009. Chemolithoautotrophic production mediating the cycling of the greenhouse gases N2O and CH4 in an upwelling ecoystem. Biogeosciences 6: 3053-3069.

“Chemolithoautotrophy is the non-photosynthetic biological conversion of C1 molecules (usually CO2 or CH4) into organic matter."

These authors studied the chemolithoautrophic community in the south-eastern Pacific ocean, at a long-term research position on the small continental shelf of Chile. At this position, the water is about 90m deep, and is part of one of the largest and most productive upwelling regions of the world, where productivity is extremely high. For about 70% of the year, surface winds drive upwelling that brings nutrients such as NO3- up to the photic zone, allowing massive planktonic productivity. Some of these plankton are non-photosynthetic prokaryotic autotrophs, using inorganic molecules such as NH4+, NO2-, and HS- as electron donors to drive the energy-intensive process of fixing inorganic carbon, chiefly CO2, as organic matter. These organisms require oxygen as an electron acceptor, thus they are all aerobic organisms and do not thrive in anaerobic environments. Assimilation of CO2 in the dark is the diagnostic signal of the presence of these organisms. Some chemolithoautrophs use CH4 in addition to or instead of CO2, but always under aerobic conditions; anaerobic methanotrophs are outside the scope of this study, and have not been identified in this system.

Nitrous oxide accumulates in these waters, indicative of greater production than consumption. These authors state that N2O reduction can only occur through a single identified pathway, that of total denitrification that takes NO3- or NO2- all the way to N2, and only under extremely limited O2 conditions; they cite Elkins et al. (1978) and Farías et al. (2009). Variation in space and time in dissolved N2O patterns do suggest some consumption is occurring, but in general production outweighs consumption.

Kostina et al. 1994

2010-02-03T17:37:00.000-05:00

Kostina NV, Stepanov AL, Umarov MM. 1994. Study of the complex of nitrous oxide-reducing microorganisms in the soil. Eurasian Soil Science 26: 81-87.

These authors extracted microorganisms from a range of soils apparently collected from various places in Russia and produced cultures of organisms capable of reducing nitrous oxide. The only nitrogen source in culture vials was N2O, and conditions were rendered anaerobic by flushing with argon gas.

Most cultures gradually lost their ability to reduce N2O, especially mixed-species cultures. A few pure strains were isolated that did not show this loss, and maintained high levels of activity in storage.

Pseudomonas spp. and Bacillus spp. contributed the vast majority of N2O-reducing activity in all soils, with other groups including Aeromonas spp., Micrococcus spp., Flavobacterium spp., Erwinia spp., and an organism identified as “similar to Corynebacterium” also showing some activity. Neither actinomycetes nor eukaryotes were found in any of the cultures capable of N2O-reduction, indicating this is a physiological process not possessed by these organisms.

This is an English translation of a paper that was probably originally in Russian: Pochvovedeniye 1993 25: 72-76.

Yu and Patrick 2004

2010-01-28T18:20:00.001-05:00

Yu K, Patrick WH Jr. 2004. Redox window with minimum global warming potential contribution from rice soils. Soil Science Society of America Journal 68: 2086-2091.

These authors followed up a previous study (Yu and Patrick 2003) that discovered a critical range of soil redox potential (Eh) across a range of pH for rice-agriculture soils, by examining soil Eh in more detail. The critical range is based on minimizing total global warming potential of all 3 major greenhouse gases, in terms of CO2-equivalents; methane and nitrous oxide have much higher radiative forcing than does CO2, when looked at on a 100-year horizon.

The critical range, for all 8 soils studied, is between 180 and -150 mV, what might be considered “moderately reducing” for soils. CO2 production is modest in this range, though it is lower at more reducing conditions. CH4 production is nearly absent in this range, but is very large at redox conditions below -150 mV. N2O production is modest in this range as well, with much higher N2O production under oxidizing conditions (Eh > 180) due to strong nitrification activity.

This paper serves to support earlier ideas about the net effects of redox conditions on the production and consumption dynamics of these gases. The story with CO2 and CH4 is fairly simple, while N2O dynamics are more complicated because there are more pathways for both production and consumption of this gas.

Ettwig et al. 2009

2010-01-28T15:20:00.000-05:00

Ettwig KF, van Alen T, van de Pas-Schoonen KT, Jetten MSM, Strous M. 2009. Enrichment and molecular detection of denitrifying methanotrophic bacteria of the NC10 phylum. Applied and Environmental Microbiology 75: 3656-3662.

These authors describe a series of experiments and procedures designed to investigate an enigmatic organism known as NC10, a bacteria in its own eponymous phylum that currently represents the only demonstrated case of biological reduction of nitrate coupled to oxidation of methane under anaerobic conditions. While anaerobic methane consumption has been observed in some archaea, it has not been found coupled to denitrification.

A laboratory culture eventually dominated by NC10 organisms of group a (a distinction within the phylum) was established, based on sediment collected from a eutrophic ditch draining agricultural land on the floodplain of the Rhine river in the Netherlands. This culture was grown and maintained under conditions in which the only carbon source was the sparge gas of CH4-CO2, and nitrogen was supplied with the mineral inputs as nitrate and nitrite, along with a wide range of other inorganic compounds and trace elements.

The major finding of this study was a wealth of knowledge of the basic characteristics of the NC10 organism, and confirmation that it does indeed oxidize methane under anaerobic conditions coupled to denitrification. This process is energetically favourable, and the theoretical stoichiometry matches the observed changes in chemical composition in these experiments, with the nitrite reduction to methane consumption ratio of 8:3.5, versus 8:3 based on mass balance calculations.

One of the surprising aspects of this organism is that its methane-oxidizing activity is completely inhibited by black butyl rubber, as is found in black rubber stoppers for serum vials and other glassware. Grey or red butyl rubber stoppers do not show such inhibition, and repeated boiling of black butyl rubber stoppers in HCl did not remove the inhibitory effects. Strictly anoxic conditions are not required for all aspects of working with this organism; brief exposure to atmospheric oxygen during liquid transfer, for example, did not inhibit methanotrophic activity.

Another strange feature of NC10 concerns its 16s rDNA sequences. General 16s primers do not amplify NC10 DNA. These authors developed new primers for the 16s region based on the DNA in their culture, which they were able to confirm as NC10 based on FISH observations. The new primers allowed them to work more easily with the NC10 DNA, which is not surprising, but the sequences of NC10 16s found did not differ in critical ways from the target regions of the general 16s primers. So, it is unknown why the general 16s primers do not work on NC10 DNA.
This paper was recommended to me and I would not likely have discovered it without this recommendation. I have results from the 2009 work at Alexandra Fjord that suggest simultaneous consumption of CH4 (oxidation) and N2O (reduction), and I did not know if these two processes might be linked in a single organism or within a system such as a symbiosis or food-chain as 2 halves of a redox couple.

My results may be more suggestive of an alternate situation. Rather than anaerobic oxidation of methane (weirdness) coupled to nitrate reduction, I may be looking for cases of reduction of nitrous oxide under aerobic conditions (weirdness) coupled to methane oxidation.

Firestone et al. 1980

2010-01-28T11:45:00.000-05:00

Firestone MK, Firestone RB, Tiedje JM. 1980. Nitrous oxide from soil denitrification: Factors controlling its biological production. Science 208: 749-751.

These authors measured the faction of N2O in nitrogen gas outputs from soil slurries under a range of conditions of substrate and oxygen availability. Slurries were employed to avoid problems associated with diffusion of materials through a soil matrix, and the process of denitrification was studied using isotopic tracers, especially in the form of 13N in nitrate and other inputs.

The controls on the production of N2O from denitrification are the concentration of nitrite (NO2-) and the availability of oxygen (O2), with time-since-anoxic another important factor. Increasing nitrite increases N2O production and increasing NO3- does as well, but less strongly, suggesting the role of NO3- is indirect, and it is the NO2- produced from NO3- that matters. Aerobic conditions inhibit denitrification, rendering the entire pathway moot. The establishment of anaerobic conditions turns on denitrification, but in a stepwise process apparently related to protein synthesis. In a series of experiments, these authors found that in the initial period of anaerobiosis, N2 is the major output. Later, N2O production increases without an increase in its consumption, and N2O is the major output. Finally, N2O consumption catches up with production, and N2 is once again the major output. Adding O2 increases the proportion of total denitrification output that is N2O, but eventually O2 does inhibit denitrification completely.

Patrick and DeLaune 1972

2010-01-28T10:50:00.001-05:00

Patrick WH Jr., DeLaunce RD. 1972. Characterization of the oxidized and reduced zones in flooded soil. Soil Science Society of America Proceedings 36: 573-576.

These authors measured the thickness of the oxidized layer in flooded soils. For Eh measurements, a platinum electrode was pushed down through the soil at a rate of 2mm/hour, sufficiently slowly for the electrode tip to reach near-equilibrium conditions as it descended. Concentrations of reduced and oxidized forms of Manganese, Iron, Sulfur, and Nitrogen were also measured.

Within a few days of submergence, soils showed a clear redox-potential profile as measured by the platinum electrode, with the oxidized layer above the reduced layer, and a transition from above 200 mV to below 200 mV across a relatively narrow intermediate layer. The profiles as measured by the chemical species distribution were similar, though Mn showed a narrower and S a deeper oxidized layer, probably relating to the redox conditions needed to reduce the oxidized compounds present in the soil; sulfate, for example, requires lower Eh values to be reduced than do ferric oxides.

Manganese, Iron, and Sulfur did not diffuse appreciably in these experiments, but Nitrogen compounds did. These authors propose the following process is occurring in these flooded soils:

“…ammonium diffusion from the reduced layer to the oxidized layer -> ammonium oxidation to nitrate (nitrification) -> nitrate diffusion from the oxidized layer to the reduced layer -> denitrification…”

To explain the observation that nitrate was absent from the reduced layer, and never very abundant in the oxidized layer, and that ammonium was rapidly depleted in the oxidized layer. In this system, nitrification and denitrification are occurring simultaneously at different positions and redox potentials.

Yates et al. 2007

2010-01-27T16:05:00.000-05:00

Yates TT, Si BC, Farrell RE, Pennock DJ. 2007. Time, location, and scale dependence of soil nitrous oxide emissions, soil water, and temperature using wavelets, cross-wavelets, and wavelet coherency analysis. Journal of Geophysical Research 112, D09104.

These authors analyzed a dataset of soil parameters and N2O emission using three subtly-different wavelet-based statistical techniques. There were two main purposes to this study; first, to examine the predictive relationships (if any) between soil parameters such as water filled pore space (WFPS) or temperature and N2O emissions; second, to evaluate the utility of these 3 wavelet techniques in analyzing this type of data.

N2O emission data is characterized by high variance in space and time, and frequent extreme values. These characteristics make many sophisticated geospatial statistical techniques not suitable, and the high spatial and temporal autocorrelation of many soil parameters eliminates many other techniques. These authors describe these limitations and some of the techniques that have been employed, and settle on 3 varieties of wavelet analysis.

Wavelet techniques are related to Fourier-transforms, and they appear to be highly complex and sophisticated methods to transform data for analysis, rather than being analytical methods per se. A large fraction of this paper is concerned with detailed description of the parameters of the transformation, and the interpretation of the results. One of the key advantages of these techniques is they usually allow examination of data across a broad range of spatial scales, thus permitting identification of the spatial scale at which important soil processes occur. Beyond that, I did not understand much of this paper.

Besides the interpretation of the differences between the 3 wavelet techniques, which was quite frankly beyond my understanding, the main result of this study was that the soil parameters that can predict N2O emissions in this landscape vary through the season. Early, around snowmelt and soil thawing, soil temperature is predictive of emissions. Later in the season, temperature loses its usefulness, and individual landscape features may present WFPS as predictive, but not in a global sense. By mid-summer, the soil parameters measured in this study no longer bore any relationship to N2O emissions. This loss of predictive value shows how complex this system is, and shows how some modeling efforts need to change in order to improve estimates of landscape-scale N2O processes.

Besides demonstrating my ignorance of advanced geospatial statistical techniques, this paper is primarily useful to me for its clear introduction describing the basic controls on and processes of N2O production in soils. My previous understanding centred on the role of water in restricting O2 availability in soils leading to changes at both the community and cell-physiology levels and consequently N2O production patterns in space and time appears to be essentially correct, and is reinforced by the early introduction section of this paper and the references therein.

Conrad 1999

2010-01-26T17:50:00.000-05:00

Conrad R. 1999. Contribution of hydrogen to methane production and control of hydrogen concentrations in methanogenic soils and sediments. FEMS Microbiology Ecology 28: 193-202.

This author reviews the chemistry behind methane production by Archaea in anaerobic environments, focusing on the contribution of H2 rather than acetate to methanogenesis. The thermodynamics and kinetics of H2-driven CH4 production are distinct from those of acetate-driven, and the stoichiometry of the situation indicates that H2 should contribute 33% of the CH4 from a given ecosystem.

Methanogenesis in anaerobic sediments and soils is the end of a short chain of microbial interactions. First, organic matter is broken down by fermenting bacteria. The products of fermentation includes H2, and the other components such as alcohols and fatty acids, are further decomposed by syntrophic bacteria, also supplying some acetate to the environment. Finally, methanogens consume either H2 and CO2 or acetate (CH3CO2-) to produce methane.

There are many studies that show this expected pattern of methanogenesis, but many other that show either over- or under-representation of H2. Where H2 contributes less CH4 than expected, the most likely explanations involve sulfate reducers, microbes capable of outcompeting H2-consuming methanogens by more efficient use of H2 and faster population growth, based on the thermodynamics of the two guilds respective metabolisms. Such situations are common in marine and acidic freshwater sediments.

Where H2 contributes more CH4 than expected, including an Antarctic soil where H2 is the basis of 100% of CH4 production, the explanation is not as well established. The explanations that have been proposed, by this and other authors, include additional sinks of acetate such as scavenging by other organisms, additional sources of H2 including geological sources, or measurements of the system taken when it was far from equilibrium. The models of H2 and CH4 dynamics are mostly based on equilibrium conditions.

That addition of sulfate inhibits methanogenesis is well established. Competition explains this observation in sediments and soils where the biological community has had time to reach something like equilibrium, with methanogens outcompeted by sulfate reducers. However, addition of sulfate to sediment immediately and completely inhibits H2-driven CH4 production, which cannot be explained by ecosystem dynamics. A model involving a threshold H2 concentration, in which H2 levels lower than some critical level determined by the thermodynamics of the situation shut down that pathway, does explain these observations.

Pennock 2004

2010-01-26T14:10:00.001-05:00

Pennock DJ. 2004. Designing field studies in soil science. Canadian Journal of Soil Science 84: 1-10.

This author reviews the major issues surrounding field-based (as opposed to strictly laboratory-based) research, focusing on issues specific or of greatest importance to soil science. Soil science’s history could perhaps be described as a fusion of physical geography and geology with agronomy, and many published studies in the soil science journals show these roots. Following the lead of previous authors, who have included ecologists, statisticians, and philosophers and historians of science, this author divides field research into 2 major categories, broadly manipulative studies and mensurative studies. Manipulative studies are, under some definitions including one tentatively employed in this paper, the only type of study that qualify for the name “experiment”, and involve complete control over experimental conditions by the researcher. Treatments in an experiment are directly related to replication, and can be applied with great precision. Mensurative studies are those that at least partly use features of the environment beyond the control of the researcher to test hypotheses or discover new information. The key feature of a mensurative study is that the features of interest are clearly defined but not controlled (i.e. not randomized) by the person conducting the study.

Replication, and avoiding pseudoreplication, is of great importance in all types of studies. However, the replication built into a manipulative experiment in the form of repeated application of treatments is distinct from the replication of a mensurative study using repeated features of the environment. That these are different types of replication is stated in this paper, but I found no more detail or explanation than that. Pseudoreplication in this paper is discussed little in the context of independence of samples; rather the discussed risk is of attempting to draw inferences beyond the inference space of the study. This is a problem in both major types of study, and can be avoided by carefully determining and describing the inference space, and expanding that space by greater replication; too-small sample sizes are quite simply labeled as unpublishable in this paper, a sentiment I can agree with.

Determining the required sample size is a major issue for all types of studies. In this author’s presentation, this is an early step in the design of the study, after the biological and statistical questions have been established but before data collection begins. There is some discussion here as well of statistical power (the chance of avoiding a Type II error, that is of failing to reject a false null hypothesis) and recommendations of flexibility regarding especially alpha values (the chance of making a Type I error, that is of rejecting a null hypothesis that is not false). For a number of reasons, some of which are practical and logistical, alpha values larger than the ubiquitous 0.05 are encouraged, because in many cases the consequences of the 2 types of error are not even, and one may wish to concentrate on reducing the probability of a Type II error.

This paper describes 10 commonly-encountered study designs in soil science and related disciplines, and then discusses study-design concerns common to all such as replication and the need to clearly define study units, samples, populations, and other important aspects. Finally, this author presents the conclusions from all of these examples and considerations in the form of a short list of key recommendations. Quoting directly:

1. A clear definition of the research question is the initial (and most critical) step. This definition dictates the type of research design that is appropriate and the specific design issues associated with different research types.
2. The appropriateness of a given research design can be judged only after a thorough review of what is known about the research question. Exploratory pattern studies can be very informative at an early stage of research, but yield little new information for well-established research topics. Equally, the imposition of a set of treatments if little is known of the processes controlling responses is unlikely to produce comprehensive interpretations.
3. There is never a good reason for haphazard sampling – the rationale for selecting sampling points in pedological, soil geomorphic, or inventory studies should be clearly stated.
4. A clear definition of the population and the elements that comprise the population under study is very important.
5. The definition of the population dictates the extent of the study and the physical or temporal space that the results pertain to, which is critical to avoid pseudoreplication.
6. The sample support, spacing, and extent of the study must be consistent with what is known of the processes controlling the phenomena being studied.
7. The construction of hypotheses for formal testing should be based on sound physical or biological reasoning, and sufficient samples should be taken to allow reliable testing of the alternative hypotheses.
8. The exclusion of phenomena because they cannot be replicated is inherently limiting to the expansion of our knowledge of soils. Innovative approaches must continue to be developed and applied so that we can expand the scale at which field studies can be undertaken.

Bremer et al. 2009

2010-01-25T17:15:00.000-05:00

Bremer C, Braker G, Matthies D, Beierkuhnlein C, Conrad R. 2009. Plant presence and species combination, but not diversity, influence denitrifier activity and the composition of nirk-type denitrifier communities in grassland soil. FEMS Microbiology and Ecology 70: 377-387.

These authors ran a manipulative, common-garden experiment to examine interactions between surface-plant community and soil denitrifier community diversities. The major finding of this paper, as described in the title, is that denitrifier community diversity is influenced by the species of plants in the system, but not how many species are there.

I saw 2 major problems with this paper that calls their major finding into question at least in my mind. First, the single greatest effect on denitrifier community composition found here was the very distinct community in the control, no-plant plots. These authors never acknowledge that zero plants is a point on their spectrum of species diversity; they state their range of plant community species richness was 2 to 8, but it was actually 0 to 8, with a strong effect of the 0 community. They do not analyze their data in this way that I can see, so I do not know if this 0-community effect does or does not reverse their conclusion that plant species diversity does not influence denitrifier diversity.

Second, the single most distinct with-plants community that was included in analysis is probably an outlier and should be excluded, because the plant community was 2 species of grasses (rather than mixed grass / forb), which also had the highest productivity in one of the study years. Furthermore that year was a high-temperature drought year across much of Europe including the study site. This probably-outlier result is acknowledged by these authors as having some of these problems, but their discussion of the probable role of greater niche-space in the soil under the more-diverse-but-same-species-richness plots does not suggest they have considered the confounding effects.

Despite the suspect results, this paper provides a useful overview, especially in the introduction and large parts of the discussion sections of denitrifier communities in soils and their probable interactions with plant communities.

Bedard-Haughn et al. 2006

2010-01-22T11:40:00.000-05:00

Bedard-Haughn A, Matson AL, Pennock DJ. 2006. Land use effects on gross nitrogen mineralization, nitrification, and N2O emissions in ephemeral wetlands. Soil Biology and Biochemistry 38: 3398-3406.

These authors used a combination of stable-isotope and measurements of chemical pools and emissions from soils techniques to examine the role of various microbial-mediated processes in contributing to N2O production in an agricultural landscape. N2O emissions are the result of a complicated suite of metabolic activity in soils, with local oxygen concentrations, driven by soil moisture, and concentrations of reactants in these chemical pathways both contributing to net processes.

In the Canadian prairies, gross N2O production is positively correlated with soil moisture, with the highest emissions associated with lower-slope and wetland soils. This is consistent with the major contributing process in N2O emissions being denitrification, the process that reduces NO3- under anaerobic conditions. However, nitrification, the production of NO3- from NH4+, has also been observed to contribute to N2O emissions, especially from drier and aerobic soils.
Simple measurements of NO3- and NH4+ concentrations in soils will not capture information about the processes cycling N between these and other pools of soil matter. Used in conjunction with measurements of those processes, such as the 15N technique used here, does provide information about the factors controlling those processes. In this case, little variation through time or space in either pool combined with patterned variation in N2O emission and 15N movements allowed these authors to infer that both nitrification and denitrification are not limited by the substrate pools, despite the quite different other aspects of these processes.

This paper provides a very detailed description of the 15N procedures used, as well as a clear discussion of the various N-cycling processes in soils.

Zuur et al. 2010

2010-01-19T17:30:00.001-05:00

Zuur AF, Ieno EN, Elphick CS. 2010. A protocol for data exploration to avoid common statistical problems. Methods in Ecology & Evolution doi: 10.1111/j.2041-210X.2009.00001.x

These authors present a step-by-step guide and recommendations for data exploration, a procedure in analysis of statistical data that should be carried out before primary statistical techniques such as regression. The point of data exploration is to look for errors in measurement, calculation or data-entry, to remove outliers, and to ensure no critical assumptions are being violated. Data exploration is not an instantaneous process, and may take up to 50% of the time spent on data analysis.

Their Figure 1 shows the steps in data exploration. Not all steps need be conducted for every dataset, for example, PCA is not sensitive to normal distribution, so the construction of histograms to evaluate normality is not necessary. On the other hand, almost all statistical techniques are very sensitive to violations of the assumption of independence.

(To avoid potential copyright issues, I have not pasted Fig. 1 from the paper here)

Figure 1 from Zuur et al. (2010). The procedures in italics are described in detail in this paper.
This paper was assigned reading for a course I am taking, Plant Sciences 813, Statistical Methods in the Life Sciences. I think the advice and instructions here will be useful.

Michalyna 1971

2010-01-15T13:30:00.001-05:00

Michalyna W. 1971. Distribution of various forms of aluminum, iron and manganese in the orthic gray wooded, gleyed orthic gray wooded and related gleysolic soils in Manitoba. Canadian Journal of Soil Science 51: 23-36.

This author examined soil Al, Fe, and Mn in some poorly drained soils of the Gleysolic order in western Manitoba, looking for indicators for soil classification that would cover some of the deficiencies of the previous criteria. The distribution of these metals, including the ratio of oxalate-extractable to dithionite-extractable iron representing amorphous and total Fe(III)-oxide forms, respectively, was a useful criterion for classification.

Iron of all types was concentrated in the upper horizons of these soils, with the highest levels in the BA and B horizons. The ratio of amorphous to total Fe(III) was also highest in these horizons, and declined with depth. This suggests amorphous Fe(III) is most abundant in relatively oxidizing conditions in these wet soils. Water content is not reported, except to note that some soils are “imperfectly drained”, others are “poorly drained”.

The ratio of amorphous to total Fe(III) found in these soils ranged from about 0.1 to about 1.2, with most measurements between 0.4 and 0.8. My own measurements, converted to the same ratio, range between 0.14 and 0.83.

Howarth 1979

2010-01-15T10:00:00.000-05:00

Howarth RW. 1979. Pyrite: Its rapid formation in a salt marsh and its importance in ecosystem metabolism. Science 203: 49-51.

This author investigated sulfur and iron dynamics in a salt marsh in the United States. Previous work by other authors had suggested pyrite (FeS2), one end-product in sulfur reduction, forms slowly over years or decades in marine sediments. This paper includes an experiment involving buried Teflon bags in which pyrite formation was detected after 48 hours. From this and other measurements, an estimate of total marshland bacterial sulfur-driven respiration was formed that is of a similar magnitude in CO2 release as is total net productivity of the marshland.

Iron metabolism in this system involves formation of amorphous iron compounds under oxidizing conditions, with a predominance of crystalline forms only under more reducing conditions.

Soulides and Allison 1961

2010-01-14T16:40:00.000-05:00

Soulides DA, Allison FE. 1961. Effect of drying and freezing soils on carbon dioxide production, available mineral nutrients, aggregation, and bacterial population. Soil Science 91: 291-298.

These authors conducted a series of experiments to investigate previously reported claims of a burst of CO2 production following drying or freezing of soils, with an associated change in soil bacterial populations. Drying soils killed large fractions of bacterial populations; freezing as well, to a lesser extent. Combined drying and freezing killed many bacteria, but did not sterilize soils. CO2 production was raised 20 to 40% over controls following drying and rewetting under different schemes, which these authors attribute to rapid breakdown of organic material by large numbers of bacterial cells in an early growth phase; CO2 production declines as populations stabilize.

I was hoping this paper would provide clues to the soil water levels tolerable by bacteria, but these authors do not discuss critical levels of moisture or temperature, beyond noting that severe drying is detrimental, and temperatures below 2ºC prevent most bacterial growth.

Clément et al. 2005

2010-01-13T10:05:00.001-05:00

Clément J-C, Shrestha J, Ehrenfeld JG, Jaffe PR. 2005. Ammonium oxidation coupled to dissimilatory reduction of iron under anaerobic conditions in wetland soils. Soil Biology and Biochemistry 37: 2323-2328.

These authors observed an unexpected chemical reaction involving the accumulation of both nitrite (NO2-) and ferrous iron (Fe(II)) under anaerobic conditions. They investigated this phenomenon further, and propose a chemical reaction in wet soils in which ammonium is oxidized under reducing conditions by transferring electrons to Fe(III), generating NO2- and Fe(II).

Nitrite does not usually accumulate in soils. These authors suggest that under normal conditions, it is consumed at least as fast as it is produced, but their experimental conditions included inhibition of denitrification, allowing nitrite to build up to detectable levels. Other oxidizers besides Fe(III), such as Mn(IV), were not detected in soil samples and were not included in the experiment.
The proposed chemical pathway is thermodynamically feasible at pH 7, though it appears to rely on goethite as the ferric iron source; from my understanding of dissimilatory iron reduction (e.g. Lovley 1991), I would expect strongly crystalline forms of iron oxide such as goethite to be highly resistant to such destructive forces, and the iron source in the systems (natural and experimental) described here to be amorphous ferric oxides instead. But the underlying chemistry appears plausible to me.

One unexpected aspect of this short communication was the authors’ use of a Dionex ion chromatography system, apparently very similar to the device I will be using to analyze root exudates. Additionally, this paper discusses the “ferrous wheel”, a memorable name for chemical cycling of iron between valencies, as an established hypothesis; I need to track down the origins of this term and learn its importance regarding my own attempts to relate measured Fe(III) contents to redox conditions.

Lovley 1991

2010-01-12T16:10:00.001-05:00

Lovley DR. 1991. Dissimilatory Fe(III) and Mn(IV) reduction. Microbiological Reviews 55: 259-287.

This author comprehensively reviews microbe-mediated reduction of iron and manganese in soils and sediments, in a long and detailed paper. Dissimilatory reduction is distinct from assimilatory reduction, in which metal ions are reduced when they are incorporated into cellular macromolecules such as enzymes and cofactors. Dissimilatory reduction is a process that ends with the accumulation of reduced metal outside the cell, and is responsible for the majority of iron and manganese reduction in sediments. While it has been observed in aerobic environments, such reduction occurs mainly in anaerobic conditions. Fe(III) in particular is most often reduced when it is the final electron acceptor in the anaerobic oxidation of organic molecules.

Microbes capable of dissimilatory metal reduction can be categorized in a number of ways. This author presents 5 categories, though I think there is considerable overlap between them, as in cases where one cell is able to reduce Fe(III) and metabolize a range of carbon sources. In the first category, reducing fermenting bacteria, the amount of Fe(III) reduced during metabolism is far less than the stoichiometry of the redox couple would suggest. This implies that Fe(III) is a minor electron acceptor during fermentation reactions, rather than the electron acceptor of choice or necessity for these organisms. Sulfur-oxidizing species in contrast do appear to reduce quantities of iron in line with stoichiometric predictions, but do not seem to gain significant energy from these reactions which occur under aerobic conditions on elemental sulfur. Hydrogen-oxidizing reducers appear to be abundant in anaerobic sediments, and seem to have high affinity for hydrogen gas; where Fe(III) is being reduced, hydrogen concentrations are low, and when hydrogen is added, Fe(III) reduction increases. These organisms need other material, such as simple organic molecules, to grow, as this reaction provides energy but little else. Organic-acid oxidizing reducers and aromatic-oxidizers are probably in many cases the same cells. As these molecules are sometimes the result of fermentation metabolisms, one possible food chain in anaerobic environments is fermentation, with some Fe(III) reduction, followed by greater levels of Fe(III) reduction linked to the decomposition of smaller organic molecules such as acetate. Such an ecological pairing is probably widespread, given the known abundance and diversity of fermenting species and the probable abundance of more aggressively iron-reducing species, many of which are probably Archaea rather than Bacteria.

There are three competing models for how Fe(III) is reduced in natural environments. The first is the enzymatic model, in which microbial cells employ membrane-bound or intracellular enzymes to transfer electrons to Fe(III) ions during the process of anaerobic oxidation of organic matter. The second is termed the redox model, and posits the majority of Fe(III) reduction is driven by equilibrium thermodynamics, with the relative levels of Fe(III) and Fe(II) in sediments controlled by abiotic factors such as temperature and pH. The third model is termed the direct-reduction model, in which some organic molecules react directly with Fe(III), without the intervention of cells or enzymes.

At first glance, the observation that Fe(III) reduction rates fall when microbes are removed from sediments supports the enzymatic model, but in fact all three models rely on microbial metabolisms. In the redox model, competing electron acceptors such as oxygen and nitrate are consumed by microbes, lowering their concentrations to levels too low to influence the transitions between Fe(III) and Fe(II). And the direct model relies on microbes releasing key organic molecules known to reduce Fe(III) in vitro. However, several other lines of evidence, such as the lack of spontaneous shifts in iron valency ratios in stored sediments, the large changes in pH associated with widely-used extraction methods, and the general rarity of rapidly-acting direct-reducing molecules all lead to the conclusion that while the other mechanisms may contribute some iron reducing activity in some situations, the overwhelming majority of Fe(III) reduction to Fe(II) occurring is driven by microbes and their enzymes.

This author spends nearly as much time discussing Mn(IV) reduction as Fe(III) reduction, but I have little interest in Mn chemistry at this time. However, in the discussion of the competing models of metal reduction, mention is made that Fe(II) in solution may reduce Mn(IV), removing accumulated Fe(II) from iron reduction and abiotically returning the iron to Fe(III) while generating Mn(II). This abiotic back-reaction closes the loop on cycling Fe(III).

Iron reduction is postulated as one of the first, if not the first, globally important metabolic pathway, with early Archaea using Fe(III) as their final electron acceptor in a generally reducing environment lacking free oxygen and nitrate. If Fe(III), a non-soluble, precipitating substance is the primary oxidizing agent, the redox environment of the Earth 2 billion years ago was upside-down compared to today: the surface and atmosphere was reducing, while buried and water-saturated sediments were oxidizing.

Freshwater swamps differ from aquatic and marine sediments in a number of ways, including a generally high organic content and plenty of sulfates. Under these conditions, both iron and manganese may cycle rapidly between reduced and oxidized forms, further reinforcing the idea that iron may cycle in a closed or nearly-closed loop between Fe(III) and Fe(II) in wet soils. Soils that periodically dry and become oxic, such as rice paddies, may also employ molecular oxygen in this cycling, with the formation of amorphous Fe(III) oxides during the dry season, and reduction of this iron during the wet season. These reactions would restrict methane production, at least during the early part of the wet season, because Fe(III) reduction diverts electrons away from methane production.

The physical structure of iron oxides in the environment has a large effect on rates of Fe(III) reduction and populations of microbes. More strongly crystalline forms, such as goethite and hematite, are not readily reduced, while amorphous forms are consumed rapidly. Presumably it is amorphous forms that accumulate when Fe(II) is oxidized to Fe(III) and precipitates from solution as an oxide, leading to a labile pool of iron distinct from the highly resistant pool of crystalline mineral iron.

When it is not back-oxidized rapidly, Fe(II) and Mn(II) accumulate in solution, and are subject to water movements, potentially removing them from the site of production. Dissolved, reduced metal can be problematic, as these ions are readily oxidized by atmospheric oxygen if water flows contact the atmosphere, and will precipitate as rusty powder in drinking water and irrigation infrastructure. This also suggests that Fe(II) will not accumulate in natural systems over long periods, and that measuring the amount of Fe(II) in a sample is not a good indicator of iron-reducing microbial populations outside of microcosm experiments.

Iron oxides tend to provide strong adsorption surfaces for many other substances, including phosphate and heavy metals. The release to solution and movement of these substances can be a major concern when iron oxides are reduced.

Banded iron formations in some sediments and rocks, in which magnetite is deposited, appear to be the result of dissimilatory iron reduction. Magnetite is a mixed-valence iron oxide that behaves magnetically; small, characteristically-shaped crystals of it are produced intracellularly by magnetotactic bacteria, but much larger amounts are produced by a range of iron-reducing bacteria. The associated organic matter, masses and crystal structures of banded iron formations strongly suggest ancient dissimilatory iron reduction coupled to the decomposition of organic matter.

There are a range of factors controlling iron and manganese reduction in natural environments. Metal reduction is decreased in the presence of alternate electron acceptors, especially oxygen and nitrate. Oxygen is a thermodynamically favourable electron acceptor compared to either Fe(III) or Mn(IV), and nitrate appears to inhibit Fe(III) reduction by lowering electron availability below necessary levels. From a biological perspective, aerobic bacteria can usually outcompete iron reducers, many of whom are obligate anaerobes and are killed or inhibited by the presence of oxygen.

As previously described, the form of metal available in the environment also has a major effect on metal reduction rates, with more strongly crystalline forms most resistant to chemical alteration by microorganisms. “Poorly-crystalline” forms, presumably including amorphous metal oxides, are the major source of oxidized metals for reducing cells. Thus, extraction and measurement procedures that involve only the least crystalline forms of Fe(III) oxides are a good measure of the iron available for reduction by microbes. The widely-used oxalate extraction method normally does not extract significant quantities of Fe(III) oxides bound in strongly crystalline forms, except when “catalytic quantities” of Fe(II) are also available, as when weakly crystalline mixed-valence minerals such as magnetite are present. In such systems, oxalate extraction will gather a larger fraction of the total dithionate-citrate extractable iron, even though much of the iron so extracted will be in a form not actually available to soil microbes.

This paper is an enormous review of iron metabolisms in soils and sediments. It did not actually answer my current question, about the cycling of Fe(III) in soils and how comparisons of amorphous (oxalate-extractable) and total (dithionate-extractable) can be used to support inferences about long-term redox status in soils. However, I do think I now have a better grasp of general soil iron and manganese conditions and chemical transformations.

Liang and Balser 2008

2010-01-09T15:20:00.001-05:00

Liang C, Balser TC. 2008. Preferential sequestration of microbial carbon in subsoils of a glacial-landscape toposequence, Dane County, WI, USA. Geoderma 148: 113-119.

These authors examined the microbial communities at a range of depths in soils near the University of Wisconsin, Madison campus. Soil organic carbon includes markers of microbial groups such as amino sugars, molecules that are absent from plants and specific to some groups of soil microorganisms. As markers, these molecules have several advantages; besides their utility in identifying organisms, they are stable in soils, persist after cell depth, and can apparently be extracted and examined using fairly simple laboratory techniques plus access to a gas chromatograph.

This study represents a general survey of soil carbon in this system, an examination of the pools and fates of different carbon molecule classes as well as the contributions of broad groups such as bacteria and fungi to soil physiology in different soil horizons. There were three main conclusions:
1. Upper soil horizons are relatively enriched both in total SOC and amino sugars. The source of this material almost certainly is some combination of surface plant litter and root exudates, not surprisingly supporting a large community of microorganisms in the near-surface soil.
2. Amino sugars accumulate in subsoils, despite the redox environment (presumably somewhat negative) associated with the water table.
3. Amino sugars, while useful, are not sufficient on their own to elucidate mechanisms of SOC turnover and sequestration by soil microbes. Variability between sites and between horizons suggests a major role of both history and site-specific factors in structuring communities at a level distinguishable by ratios of various amino sugars.

This paper is one of a handful I have that explicitly examine microbial communities and variation by depth. However, as this paper describes what I think is a first-look at a soil microbiological system, it lacks some detail and strong conclusions. Amino sugars may be useful in my own research, though I think our lab has more familiarity with other techniques useful for examining interactions between soil bacteria and fungi.

Beyond my current research, this paper is clearly written, not too long, and presents a set of well-described investigations built on a solid foundation of general theory. This suggests it may be useful as a teaching tool, perhaps as a paper a 2nd-year undergraduate would have the skills and knowledge to understand.

Wagner et al. 2009

2010-01-09T11:50:00.001-05:00

Wagner D, Kobabe S, Liebner S. 2009. Bacterial community structure and carbon turnover in permafrost-affected soils of the Lena Delta, northeastern Siberia. Canadian Journal of Microbiology 55: 73-83.

These authors examined the microbial communities at two depth bands (near-surface and near-permafrost) in low-centred tundra polygons at the vast permafrost wetland of the delta of the Lena River. The delta covers more than 60 000 km^2, and much of it appears to be a reserve or national park of Russia. The CAVM (Walker et al. 2002) describes most of the delta as vegetation type W2, sedge, moss, dwarf-shrub wetland, and satellite images from Google maps shows very extensive lake and pond coverage of the landscape. In short, it’s pretty wet, and generally cold.

The general finding of this paper is that while near-surface communities include a wide diversity of aerobic and facultatively-anaerobic bacteria, the deeper, colder, anaerobic portions of the soil contain almost no aerobes, and are instead dominated by “fermenting” species capable of decomposing recalcitrant organic carbon molecules under negative-redox conditions. There is a sharp temperature gradient, which combined with the poorer quality of carbon, the lack of oxygen and negative redox conditions, and the general water saturation at depth creates conditions near the permafrost suitable only for the slow microbial metabolisms. None of this is particularly surprising, but the observation of decreased biodiversity with water saturation does suggest the worrying possibility that increased water in this system, driven by melting permafrost and climate change (particularly upstream in the long and North-flowing Lena) could drive these microbial communities to lose some “physiological skills” such as the ability to oxidize methane, a metabolic pathway possessed only by some aerobic prokaryotes.

This paper is quite important to my own work, I think. Besides emphasizing the role of water content in structuring soil chemical and especially biological conditions, the description of the methods used to measure microbial biodiversity should be useful. However, while the BIOLOG plates seem interesting, the results of this technique are not at all well explained in this paper. I do not know what is indicated by the relationship shown in Figure 3, for example, of changes in colour development associated with carbon turnover of various categories of organic substrates. Several of the figures are simple plots of principal component analysis (PCA), literally just PC1 vs. PC2 with some outlines drawn around some clusters. I’m sure there is more of interest in this paper besides the coarse outline of biodiversity differences in communities, but without a more thorough explanation of the nearly-raw data I cannot see it.

Lovley and Phillips 1986

2010-01-08T17:37:00.001-05:00

Lovley DR, Phillips EJP. 1986. Organic matter mineralization with reduction of ferric iron in anaerobic sediments. Applied and Environmental Microbiology 51: 683-689.

These authors examined the reduction of iron, both supplemented and already present, in sediments collected from the bottom of the Potomac river across a salinity gradient from freshwater to brackish estuarine. A previous hypothesis in the literature suggested that observed decreases in methane production in the presence of trivalent iron, Fe(III) were caused by Fe(III) being toxic to methanogenic prokaryotes. This hypothesis was disproved in this study, and the results of this study suggest instead that methanogens are outcompeted by iron-reducing species because of the greater thermodynamic benefits of Fe(III) reduction compared to methane production. Van Bodegom et al. (2004) reported instead that rather than competition, Fe(III) directly inhibits methanogenesis, though I think this is due to metabolic switching within individual cells, not interactions between distinct methanogenic and iron-reducing populations.

The other major finding of this paper is that the form of the Fe(III) in the environment has a major effect on Fe(III) reduction. Amorphous ferric oxyhydroxides are reduced much more readily than are crystalline forms. These authors do not speculate on the mechanism underlying this difference, though I suspect surface area exerts a major controlling influence.

These authors did not measure Fe(III) forms in native sediments, rather they added amorphous Fe(III) and measured Fe(II) after incubation. Thus, while my own studies of the ratios of amorphous to crystalline Fe(III) are not assisted by these techniques, these authors do provide a clear and apparently fairly simple protocol for the measurement of Fe(II), involving extraction by HCl and reaction with a molecule that turns purple when complexed with Fe(II) allowing measurement of Fe(II) amounts from the absorbance spectrum of the resulting solution. I need to learn more about Fe(III) biogeochemistry before deciding to pursue such an analysis; I suspect one fate of Fe(III) is to cycle through an organism and be returned to the oxidized state, rather than shuttling directly from Fe(III) to Fe(II).

Siciliano et al. 2009

2010-01-06T14:45:00.001-05:00

Siciliano SD, Ma WK, Ferguson S, Farrell RE. 2009. Nitrifier dominance of Arctic soil nitrous oxide emissions arises to due fungal competition with denitrifiers for nitrate. Soil Biology and Biochemistry 41: 1104-1110.

These authors examined the nitrous oxide emissions, microbial communities, and some components of nitrogen cycling in soils from three landforms at Truelove Lowland, on Devon Island. Previous results (Ma et al. 2007) had indicated that Arctic nitrous oxide emissions are not sensitive to soil moisture, at least in the range of 50% to saturated water filled pore space. This study includes a series of incubations of soil samples at a range of temperatures similar to ambient conditions, and treatments to disrupt fungi or particular types of prokaryotes.

Large differences in community composition were found between the three landforms, with the highest biomass and fungi:bacteria ratio in the wet sedge meadow and lowest in the raised beach crest (the lower foreslope was intermediate by these measures). Competition between fungi and denitrifiers for soil nitrate pools was inferred as the mechanism allowing dominance of emitted N2O by nitrifiers; fungi and denitrifiers are busy scavenging every available electron acceptor starting with nitrate and running all the way down to N2 gas, so almost any N2O that escapes was generated by nitrifiers in conditions not favoured by either of the other major groups.

This paper serves to demonstrate the very complex nature of soil biology, especially regarding the multiple and interacting pathways that may produce or consume materials of interest such as N2O. The references in this paper should be useful for digging into this complexity.

Elberling 2007

2010-01-05T13:45:00.001-05:00

Elberling B. 2007. Annual soil CO2 effluxes in the High Arctic: the role of snow thickness and vegetation type. Soil Biology and Biochemistry 39: 646-654.

This author studied the total annual efflux of CO2 at three vegetation communities in Endalen valley on Svalbard. The three communities are each dominated by one characteristic species of plant, and are named accordingly: Dryas, Cassiope, and Salix, and from the description of the sites and their environmental parameters, there appears to be high agreement between these communities and those found at Alexandra Fjord, Ellesmere Island.

The depth and duration of snow cover was a major factor controlling (directly and indirectly) soil conditions and thus respiration. Snow depth varied with vegetation type, though the causal relationship is probably snow to plants, via soil temperature (more snow = higher winter temperatures) and soil moisture content (snow accumulates at and melts into depressions and certain slope positions). Higher temperatures and wetter conditions correlated with higher soil respiration, both in winter and summer. All sites experienced a brief period of water saturation in the upper 5cm of the soil during spring thaw, though sites varied in when thaw happened, with Dryas first and Salix last, corresponding with winter snow cover depth.

Soil conditions among the sites seem to have been broadly similar; not surprising considering the close proximity of sites and the consistent soil type across the valley, though soil under Cassiope tetragonal patches was more acidic. This acidity seems related to a reduced concentration of base cations (especially Ca2+ and K+) under Cassiope plants.

Summer water content did not correlate with annual CO2 flux, which this author attributes to the generally well-drained soils, a lack of large precipitation events, and long periods without rain leading to typically dry soils everywhere, though soil respiration at the Dryas site may have been water-limited, as this was the driest site.

Winter temperatures in the soil averaged warmer than -10ºC at all sites, warm enough for microbial activity. A burst of CO2 during spring thaw was not predicted from soil parameters, but was attributed to increasing microbial activity associated with warming temperatures and the release of high-quality organic material from winter-killed microbial cells. Winter CO2 efflux averages were 0.11 to 0.28 µmol / m^2 / s, not far from values we found (for example) at the Cassiope site at Alexandra Fjord.

This paper contains much that is valuable to my current research, including both the data and patterns found and the discussion with other relevant references.

Angel and Conrad 2009

2009-11-26T09:45:00.000-05:00

Angel RA, Conrad R. 2009. In situ measurement of methane fluxes and analysis of transcribed particulate methane monooxygenase in desert soils. Environmental Microbiology 11: 2598-2610.

These authors examined the methanotroph communities of sub-tropical desert soils in Israel, using both field and lab measurements of methane fluxes and molecular investigation of sampled microbes. Negative surface flux, indicating consumption of atmospheric methane by soil, was found only at an undisturbed site in this study; the 4 other sites were varying degrees of agricultural and did not show clear patterns of consumption of methane at low concentrations. Addition of water, simulating a typical local rainfall event, eliminated methanotroph activity for about 12 hours, then this activity rebounded to well above background for up to 48 hours. This process of apparent short-term community dynamics was not investigated or discussed in much detail by these authors, though I found it one of the most interesting observations.

Methanotrophs were identified in soil samples by the usual suite of molecular biology methods. One set of primers used here is described as also targeting certain clades of amoA, a gene prominent in ammonia oxidation. These primers were successful at amplifying sequences even from soils in which methanotrophic activity had not been detected, suggesting that many or all of the sequences amplified by these primers were not actually methanotroph sequences, but rather sequences from apparently ubiquitous ammonia oxidizing bacteria.

The target gene for methanotrophs encodes a membrane-bound protein involved in transporting methane into the cytoplasm. From the way some primers also targeted amoA, I think perhaps there is a shared ancestry among the pathways for scavenging environmental ammonia and for scavenging environmental methane, though these authors do not delve into that discussion.

This paper was apparently instrumental in structuring the thoughts of my co-author (Dr. Siciliano) regarding how we should structure the manuscripts we are preparing based on the 2009 Alexandra Fjord field season. Up to this paper, we had been considering including both molecular analysis (based mainly on qPCR) and soil-properties (nutrients, root exudates, moisture, trace gases, etc.) in our nascent “Pits & Probes” manuscript. However, this paper demonstrates the considerable volume of work required to achieve a useful molecular dataset, suggesting that we would be better off saving these DNA data for a subsequent study, where they can be described and analyzed at the appropriate level of detail.

Wrage et al. 2004

2009-11-12T09:45:00.000-05:00

Wrage N, Lauf J, del Prado A, Pinto M, Pietrzak S, Yamulki S, Oenema O, Gebauer G. 2004. Distinguishing sources of N2O in European grasslands by stable isotope analysis. Rapid Communications in Mass Spectrometry 18: 1201-1207.

These authors used the signature ratios of stable isotopes of oxygen and nitrogen in a range of soil chemicals and microbial metabolic pathways to identify the source of N2O produced in grasslands monitored as part of a long-term greenhouse gas international experiment. There are 3 known pathways to N2O production: nitrification, in which N2O is produced as a by-product of ammonium oxidation to nitrate, nitrifier denitrification, in which nitrifying organisms reduce nitrite to dinitrogen gas via N2O, especially under anaerobic conditions, and denitrification, in which nitrate is reduced to dinitrogen gas via N2O by denitrifying organisms.

Previous studies had often used acetylene to inhibit N2O production, but this has been found to be unreliable. In contrast, the stable isotope approach used here was able to detect both N2O production and consumption even when reservoirs of N2O were very low. Nitrification was the most important N-transforming process found in these systems, with most N2O produced probably from reduction pathways.

Tack et al. 2006

2009-11-12T09:40:00.000-05:00

Tack FMG, Van Ranst E, Lievens C, Vandeberghe RE. 2006. Soil solution Cd, Cu and Zn concentrations as affected by short-time drying or wetting: the role of hydrous oxides of Fe and Mn. Geoderma 137: 83-89.

These authors examined the effects of short-term changes in redox conditions on the interaction between trace heavy metals and oxides of iron and manganese in agricultural soils of Flanders. The expectation was that two weeks of water-saturated and hence reducing conditions would allow these iron and manganese oxides to dissolve, and then re-precipitate when soils were returned to oxidizing conditions at field capacity or dry conditions. This process of iron and manganese chemistry would dominate redox conditions in soil solution, and dominate the chemistry of trace metals, controlling the solubilities of the trace metals.

Contrary to expectations, short-term wetting and drying cycles did not push the chemistry of trace metals around to a significant degree. The authors state that they cannot distinguish between the competing explanatory hypotheses of very slow transformations of the expected type or that these processes occurred at the expected rates but reverted during the later stages of the incubations of soils at particular moisture levels.

In the conclusion, these authors describe the importance of periods of drying, where soil moisture levels fall far below field capacity. Unlike cycles of saturation and field capacity, dried soil has much higher oxygen levels, and the moisture shortage has important effects on soil microorganisms, and the local redox conditions in microhabitats. Occasional very dry periods have a disproportionate effect on heavy-metal chemistry in soils.

For my purposes, this paper was most useful for the clear description of the methods used to measure iron oxides in soils, from a distinctly geochemistry perspective. These authors clearly state that measurement of soil levels of “amorphous” and “crystalline” Fe(III)-oxides are based on operational definitions, as the measurements are based on not-entirely-specific dissolutions of particular fractions of soil iron with particular solutions. These dissolution methods are widely used, and it was useful to see the limitations of these methods clearly described.

Nemergut et al. 2007

2009-11-12T09:35:00.000-05:00

Nemergut DR, Anderson SP, Cleveland CC, Martin AP, Miller AE, Seimon A, Schmidt SK. 2007. Microbial community succession in an unvegetated, recently deglaciated soil. Microbial Ecology 53: 110-122.

These authors describe the partly-predictable patterns of succession among the soil microbes of a glacial foreland in Peru. Primary succession on new terrain, as found in front of a receding glacier, has been studied to some extent, especially regarding the vegetation. Studies of the microbial communities have been rarer, but the few that have been conducted have suggested that these communities also show predictable patterns of community assembly and turnover associated with soil age. Basic ecological theory has led to the nitrogen paradigm of primary succession in soils: nitrogen is absent from new mineral substrate, thus nitrogen fixing organisms have a competitive advantage and are therefore abundant. The presence of nitrogen fixers is tightly linked to the accumulation of soil nitrogen; hence these organisms may facilitate later successional stages.

The study site in this paper is in Peru, at a glacier that seems to be receding quickly. These authors sampled from 3 transects arranged parallel to the front of the glacier, located adjacent to the glacier on soil less than a year old, 100m away on soil about 4 years old, and 500m away on soil about 20 years old. This area receives very high inputs of pollen, leading to the hypothesis that heterotrophic, nitrogen-fixing organisms may be present, using the pollen as a carbon source but drawing nitrogen from the atmosphere because the C:N ratio of pollen is higher than that of microbial biomass. Surface soil samples were collected, kept at 0C, and analyzed in Colorado. Much of the analyses were detailed phylogenetic examination, including the P-test of Martin (2002); note that he is one of the authors of this paper. OTU and a range of sequence-data fine-tuning techniques were also employed.

Over the study area, autotrophic nitrogen fixers were abundant. The bacteria found were extremely diverse at the highest taxonomic levels, and many sequences identified were not closely related to existing sequences in public databases. Diversity increased rapidly from the youngest soils to the 4-year-old, then plateaued.

One very interesting group of bacteria found are the Comamodaceae; sequences reported in other studies were in many cases derived from glacial or ice-sheet ice. The Comamodaceae found in the youngest soils here may have persisted as viable populations in the glacier; differences between the two closely-examined youngest communities suggest physical and genetic isolation for hundreds to thousands of years, allowing speciation events to accumulate differences.

Other patterns among the sequences identified suggest that the earliest colonizers of new terrain may be cosmopolitan – some of the Comamodaceae sequences, for example, are similar to those derived from a glacier in Nunavut. Later colonizers may be more endemic, and displace the earliest colonizers as soils age. The trophic status of the first colonizers is not clear; these authors did not have a test for definite autotrophs or heterotrophs, as Comamodaceae are known to include both modes. At the macrobiological level, the earliest arrivals on new terrain are typically heterotrophs, insects that feed on deposited organic matter such as wind-blown pollen.

This paper is very useful to me, describing as it does a complete set of analytical procedures for my planned biogeographic / phylogenetic studies, as well as providing data in the form of publicly-accessible sequences and analyzed information on patterns of soil microbial community assembly.

van Bodegom et al. 2004

2009-11-09T15:45:00.000-05:00

van Bodegom PM, Scholten JCM, Stams AJM. 2004. Direct inhibition of methanogenesis by ferric iron. FEMS Microbiology Ecology 49: 261-268.

These authors present the novel finding that inhibition of methanogenesis in anaerobic sediments by Fe(III) is a direct effect, and not the result of competition for resources between different species of microbes. They used three pure strains of methanogenic archaea under controlled-atmosphere conditions, and found a clear signal of methanogenesis inhibition related to the amount of Fe(III) added to liquid culture.

This paper raises some issues I need to consider in the methanogenic, anaerobic soils at some of the systems of Alexandra Fjord, especially regarding redox conditions and available electron acceptors. Additionally, this paper provides references for widely-accepted and apparently highly effective methods for measuring Fe(II) and Fe(III) in soil solutions.

Lindsay 1991

2009-11-09T14:40:00.000-05:00

Lindsay WL. 1991. Iron oxide solubilization by organic matter and its effect on iron availability. Plant and Soil 130: 27-34.

This author reviews the chemistry of bioavailable iron in soil solutions. The major controls on the availability of iron in soil solution, which is normally very low, are pH, redox status, and the presence of organic matter and microsites where organic matter is decomposed by microorganisms under oxygen-limited conditions.

The usual concentration of iron in solution in soils is extremely low, and its exact value suggests contributions from multiple solid iron species, including amorphous and a range of crystalline forms of Fe(III) oxides. Organic matter produces transient small organic acids as it is decomposed, which complex with iron and help to bring it into solution. However, these organic acids are in the middle of the soil organic matter decomposition pathway, and do not persist for long in soils. Bringing more iron into solution relies on a combination of local reducing conditions around respiring roots and among microbe-and-SOM microsites, and local pH. Plants and other organisms secrete compounds that are effective at solubilizing iron and making it available to plants. The other major source of iron for organisms is local fluctuations of reducing and oxidizing conditions, which bring iron into solution and precipitate it as metastable, mixed-valence ferrosic hydroxide, which provides iron to solution at much higher concentrations than most other solid forms of iron.

This paper provides useful background information about the chemistry of iron in soils, but it is not clear to me which forms of iron should be targeted and measured in soils if one wishes to learn something about local aerobic and redox conditions.

Elberling et al. 2004

2009-11-06T16:40:00.001-05:00

Elberling B, Jakobsen BH, Berg P, Sondergaard J, Sigsgaard C. 2004. Influence of vegetation, temperature, and water content on soil carbon distribution and mineralization in four High Arctic soils. Arctic, Antarctic, and Alpine Research, 36: 528-538.

These authors examined the carbon pools and carbon dioxide effluxes from four ecosystems at Zackenberg, in north-east Greenland. Their four ecosystems are very similar to the Alexandra Fjord systems of Dryas (CAVM P1), Cassiope (P2), Salix (G3), and Wet Sedge Meadow (W1).

There are two major sources of soil CO2: respiration by plant roots, and microbial respiration. Which of these two processes dominates CO2 production is a matter of some debate, with studies in the 1990s and 2000s indicating either when measuring similar arctic ecosystems. This study does not settle that debate, with estimated ratios of the two processes ranging from 9:1 to 1:9. What is clear is that plants and microbes compete for resources in arctic soils, with considerable variation in both time and space, even among vegetation communities in one valley with a consistent above-ground climate.

These soils include a buried A-horizon, with birch leaves present in pockets of organic-rich former topsoil indicative of surface conditions during the previous climatic mild period approximately 5000 years ago. These pockets of “Ab” create something of a wildcard situation for CO2 evolution, being responsible for a considerable fraction of the measured CO2 efflux at all ecosystems to varying extent.

To measure subsurface concentrations of CO2, these authors extended a probe to a range of depths and connected it to their gas analyzer also used for measuring surface fluxes, much as we have done at Alexandra Fjord. However, their gas analyzer only measures CO2, and they did not allow their probes to equilibrate to subsurface conditions for very long; a few minutes seems to be the usual protocol in this case.

A range of soil parameters were measured, and in general variation between vegetation types in these parameters exceeded variation within. The Cassiope system was the most variable, but also had the most variable Ab layers, which probably accounted for most of the variation. These patterns of variation at all ecosystems, however, suggest that the effects of climate change will not be uniform across the High Arctic, with increased temperatures leading to perhaps increases or decreases in the decomposition of buried organic matter and CO2 effluxes.

Like our own results from Alexandra Fjord, the Salix ecosystem at Zackenberg showed the highest below-ground CO2 concentrations. At Zackenberg, CO2 concentrations in Salix were mostly related to microbial decomposition of organic matter, with reduced soil water content leading to more oxygenation and higher temperatures, both increasing the rate of decomposition. In contrast, the water-saturated Eriophorum system had very high carbon stores and the highest CO2 efflux, but a decrease in water levels here leads to a shift from methane production to CO2 production, rather than a simple increase in one rate.

Overall, this paper provides some results and considerations of high relevance to my own work, especially given the high overlap in the range of ecosystems under consideration and the range of methods employed in their analysis. Their systems are not identical to those I studied; for example, the Dryas system at Zackenberg appears to be considerably drier and with less vegetation cover compared to the system with the same name at Alexandra Fjord.

Jones and Henry 2003

2009-11-02T17:30:00.000-05:00

Jones GA, Henry GHR. 2003. Primary plant succession on recently deglaciated terrain in the Canadian High Arctic. Journal of Biogeography 30: 277-296.

These authors examined five glacial foregrounds on Ellesmere Island, one intensively and the other 4 “extensively”, to determine patterns of succession among plant communities on sterile ground. The ecological literature recognizes several different modes of succession, including a categorization by Henry and Svoboda (1987) based on the relative strengths of biotic and abiotic factors. This model of succession recognizes 3 modes; directional succession with species replacement, directional succession without species replacement, and non-directional succession without replacement. They are arranged in increasing importance of abiotic factors, referred to here as “environmental resistance”, which operates in opposition to “biological driving forces”.

In temperate regions, where much of the relevant ecological theory has been developed, biotic factors are mainly competition. In the High Arctic, a successional pattern consistent with directional-with-replacement was found, demonstrating that this can occur even in environments with obviously severe abiotic factors. However, these authors argue that the biotic factor driving this succession was probably not competition, because total plant cover remains below 10% by area even at the fourth stage recognized here, and species richness is always very low. The polar oasis landscape with 80-100% plant cover was never reached within the approximately 50-year old glacial forelands examined by these authors, though it is likely that competition is important in that “stage 5” level of High Arctic succession.

Other biotic variables suggested playing a role in successional dynamics in the High Arctic included facilitation and life-history characteristics. These factors are not independent; later successional species such as Salix arctica appear not to be able to establish until soil fertility has been improved by mats of very-early-colonizing mosses, and are long-lived, slow-growing species that contribute little to the early seed bank and seed rain. Thus, multiple plant and environmental characteristics appear to interact when structuring early communities.

I read this paper to try to gain some understanding of ecological succession and the role of time-since-deglaciation among the ecosystems of Alexandra Fjord. Rather than being distinct successional stages in sequence as I had previously supposed, it appears the various lowland ecosystems are all of a similar age, and have different vegetation communities as a result of other factors besides simply relative proximity to the Twin Glacier. Dryas integrefolia and Cassiope tetragona were important parts of this study, and both appear in stage 4, after primary-colonizing mosses, and early-colonizing forbs such as Papaver radicatum and early-colonizing deciduous shrubs like Saxifraga spp. Both Dryas and Cassiope form associations with mycorrhyzal fungi, a requirement that may slow their colonization of novel habitats; earlier-spreading plants do not form these associations, and instead may be limited by seed dispersal.

This was helpful in organizing the structure of the manuscript I am currently working on, which will describe some of the soil biotic communities both in the Alexandra Fjord lowlands and in the adjacent polar desert. It is not a simple story of succession from one ecosystem to the next, but succession does play a role.

Hashimoto and Suzuki 2002

2009-10-15T17:35:00.001-04:00

Hashimoto S, Suzuki M. 2002. Vertical distributions of carbon dioxide diffusion coefficients and production rates in forest soils. Soil Science Society of America Journal 66: 1151-1158.

These authors inverted the usual soil-gas measurement approach of inserting probes into soil: they inserted soil into their apparatus. “Undisturbed” soil samples approximately 20cm in diameter and 40cm long were collected from a research forest in Japan, dried for up to 17 days, and then placed into the laboratory apparatus. This consists of a sample cylinder big enough to take the soil samples, with chambers above and below that allow control of boundary conditions. Sample ports through the side of the cylinder allow measurement of CO2 concentrations at various positions in the sample. A set of pumps, valves, and tubes allow control of soil water conditions, including manipulation of water potential.

The point of this device is to measure CO2 diffusion in large soil samples, and control for water and temperature effects. Theories of soil gas diffusion can be tested using this apparatus.

Kammann et al. 2001

2009-10-15T16:55:00.000-04:00

Kammann C, Grünhage L, Jäger H-J. 2001. A new sampling technique to monitor concentrations of CH4, N2O and CO2 in air at well-defined depths in soils with varied water potential. European Journal of Soil Science 52: 297-303.

These authors invented a buried probe based on silicone tubing for long-term monitoring of soil gas concentrations. In essence, this is simply a length of silicone tubing coiled into a flat “snail” shape and secured with wire mesh, with silicone stoppers at both ends. One end is penetrated by a stainless steel tube that extends to the surface, and is topped with a stopcock that allows sampling by syringe. These authors tested their design in a few laboratory and field studies, and demonstrated the gases CH4, N2O and CO2 do indeed diffuse into the hollow interior of the probe, and reach 95% equilibration in as little as 7 hours. They state the advantages of their design over other soil gas probes, most notably the ability to use this design in very wet and saturated soils, which is where CH4 processes may be of greatest interest.

The downside of this probe design from my point of view is the need to dig a pit for these probes, and the consequent long wait before the disturbance has dissipated and accurate sampling can begin. However, for long-term monitoring of soil gas processes, particularly in wetlands and soils prone to heavy rainfall events, these probes appear to be very useful.

Nemergut et al. 2005

2009-10-09T17:20:00.000-04:00

Nemergut DR, Costello EK, Meyer AF, Pescador MY, Weintraub MN, Schmidt SK. 2005. Structure and function of alpine and arctic soil microbial communities. Research in Microbiology 156: 775-784.

These authors review the current state of knowledge of microbial communities in cold- and snow-affected soils. Their primary study site is a ridge in Colorado with a range of habitats from sub-alpine forest to glaciated mountain-tops; all areas receive significant snow cover for much of the year. They describe only three studies of microbial communities in the Arctic, stating these are the only such studies to their knowledge at the time of preparation of this paper.

The referenced work in this review clearly demonstrates that microbial communities are active when snow covered, contrary to the previous assumption that low temperatures would effectively prohibit microbial metabolisms during winter. Indeed, microbial biomass is actually highest in winter in the alpine tundra systems studied and lowest in spring after an apparent population crash. A wide diversity of microbes has been found, from Bacteria, Archaea, and Eucaryea, including deeply divergent lineages with no known associations with described groups. The physiologies and ecological functions of many of these microbes are completely unknown.

This paper provides a useful overview of the state of the field of cold-soils microecology, with many interesting references and some surprising synthesized findings. This research group in Colorado appears to be one of the few groups in the world studying cold soil microbial communities and their links to climate change.

Lipson et al. 2009

2009-10-08T17:30:00.001-04:00

Lipson DA, Monson RK, Schmidt SK, Weintraub MN. 2009. The trade-off between growth rate and yield in microbial communities and the consequences for under-snow soil respiration in a high elevation coniferous forest. Biogeochemistry 95: 23-35.

These authors conducted a multiply-combined approach study that examined soil microbial communities in the sub-alpine forest of Colorado. They investigated growth and respiration of microbes including both bacteria and fungi, how those processes varied between summer (snow free) and winter (snow covered), and linked these processes to measures of community composition, and built a mathematical model of soil microbial metabolism and temperature. The overall purpose of this study was to thoroughly examine soil microbial processes relating to CO2 emissions and carbon cycling.

The major finding of this study was that there are effectively two distinct microbial communities in this ecosystem. In summer, there is a community of slow-growing, high biomass-yield microbes with a low specific respiration; in other words, the summer microbes grow slowly but efficiently, capturing much of the available carbon as biomass and releasing relatively little CO2 per unit biomass. In winter, the community is composed of fast-growing, low yield microbes that release much more CO2 per unit biomass.

There are effectively two ecological strategies at work, during different seasons. The winter strategy is one of competition. Available nutrients are consumed rapidly, releasing large amounts of CO2 but producing little growth. In summer, the strategy is more cooperative, with slower, less scramble-like growth that more fully uses available nutrients in growing new cells.

In general, the bacteria in the system seem more capable of the high-competition strategy, as these authors found little contribution of fungi to total ecosystem respiration in winter, by using a set of bacterial and fungal inhibitors. The winter community has a much higher response to temperature (Q10) than the summer community. A winter community at intermediate temperatures produces much more CO2 than does a summer community.

In analyzing the composition of the communities, these authors employed the P-test method of Martin (2002), as I intend to as well. I found this paper through a Web of Science search for papers citing Martin (2002); this was one of 153 papers found. The first author of this paper, D.A. Lipson, appears to have a substantial history of publications examining soil microbial communities.

Bohannan and Hughes 2003

2009-10-06T17:25:00.001-04:00

Bohannan BJM, Hughes J. 2003. New approaches to analyzing microbial biodiversity data. Current Opinion in Microbiology. 6: 282-287.

These authors review the use of three broad approaches to studying microbial biodiversity in environments. The three are 1) parametric, 2) nonparametric, and 3) community phylogenetics. Each has advantages and disadvantages, and these authors suggest a combined approach may be most beneficial. Both 1) and 2) are based on Operational Taxonomic Units, to avoid the many problems of bacterial species identification, while 3) is based on molecular phylogenies, typically 16s rDNA.

Parametric approaches make simplifying assumptions and are based on some model of species richness in microbial communities; often this model is log-normal, in which some taxa are rare, some are abundant, and most are intermediate. These approaches extrapolate from patterns in a sample to the total environment. The obvious downside to parametric approaches is the vulnerability of the model to incorrect and difficult to test assumptions.

Nonparametric approaches avoid assuming any model, and instead are typically built on an approach analogous to mark-release-recapture. Sequences encountered more than once in a sample are recaptures, and the frequency of these doubletons is assumed to be related to how many unique sequences are present: more doubletons means fewer total sequences. As a downside, these approaches provide only a lower limit to actual richness, thus generally underestimating total diversity.

Community phylogenetics approaches avoid the OTU concept and thereby preserve useful data in the form of genetic information about sequences and sequence relationships. The downside of community phylogenetics approaches is they sample a clone library derived from the environment, not the environment directly, and can therefore not extrapolate from the sample to the environment.
This paper provides several useful examples of each approach, and supports the utility of Martin’s (2002) combined approach, which is what I would like to apply to my data to be collected in 2010. Figure 3 in this paper, for example, provides a useful overview of what Martin (2002) did, and how to make inferences about observed patterns.

Fang and Moncrieff 1998

2009-10-06T17:20:00.002-04:00

Fang C, Moncrieff JB. 1998. Simple and fast technique to measure CO2 profiles in soil. Soil Biology and Biochemistry 30: 2107-2112.

These authors present a method they developed to measure sub-surface CO2 concentrations in soil. Their test environment was a slash pine (Pinus elliotti) plantation in Florida, with sandy soil subject to a broadly fluctuating water table that occasionally comes to the surface.

Their method, essentially, involves burying a perforated aluminum probe connected to the surface with flexible tubing, waiting several weeks for the disturbance to settle, and then measuring recirculated gas samples by injecting them with a syringe into an IRGA. Recirculation breaks the trade-off between errors associated with the syringe sucking air out of surrounding soils at depths different from the intended sample, and large buffer volumes inside probes become dead spaces with internal gas concentration gradients.

This is an interesting approach to soil gas probes, but is fundamentally unsuitable for my own studies because of the necessity of waiting weeks before sampling due to the disturbance of burying the probes.

Cockell and Stokes 2004

2009-10-06T17:15:00.000-04:00

Cockell CS, Stokes MD. 2004. Widespread colonization by polar hypoliths. Nature 431: 414.

These authors present a short report about the colonization of the undersides of rocks in the polar deserts by cyanobacteria and unicellular green algae. Where rocks have a protected but accessible underside, these organisms colonize, forming a pale green band a few centimetres across, between the part of the rock too exposed and dry, and too dark for photosynthesis. In polygon terrain, where the ground is sorted by frost heave (“periglacial processes”) and even large rocks are periodically jostled, areas of finer texture form in the centers of polygons, with larger rocks around the edges. The edge rocks were 100% colonized, while central areas were 5% colonized.

Fierer and Jackson 2006

2009-10-06T17:10:00.000-04:00

Fierer N, Jackson RB. 2006. The diversity and biogeography of soil bacterial communities. Proceedings of the National Academy of Sciences of the U.S.A. 103: 626-631.

These authors measured biodiversity of soil bacterial communities from 98 samples across North and South America. Rather than the expected variables that structure macroorganism biodiversity such as climate and latitude, the single most important factor for bacterial community diversity was soil pH. Soils with higher pH had greater biodiversity, regardless of geographic separation between sites or geographic position. Because soils with pH greater than 8.5 are rare, it is impossible to distinguish between a unimodal distribution, and a plateau at near-neutral pH.

Most surprising to me was the lack of correlation between bacterial diversity, as either species richness or Shannon-index diversity (richness and evenness) and plant diversity. While these authors did not assess plant diversity, they did express their own surprise that sites in the Peruvian Amazon, with acidic soils, had very low bacterial diversity despite having one of the highest measured plant biodiversities on Earth, and expanded their soil collecting sites to include other tropical and sub-tropical forests with very high plant diversity. Sites with near-neutral pH provided the highest estimates of bacterial diversity, and were primarily dry-grassland, dry-forest, and humid temperate forest sites, with low-to-intermediate plant diversity.

The method used to estimate diversity of bacterial communities was terminal-restriction fragment length polymorphism analysis, or T-RFLP. This technique relies on PCR to amplify the sequence of interest, in this case 16S rDNA, followed by digestion with restriction endonucleases and generation of “fingerprints” for each community. This is a fairly low-resolution method, with less than 100 bands per community, and no ability to distinguish between species with similar restriction sites. However, it does permit high throughput and costs much less per sample than does cloning and sequencing.

This paper is an excellent example of a recent study supporting the hypothesis that microbial global biodiversity is controlled by factors quite distinct from the factors controlling macroorganism (plant and animal) biodiversity. In this case, soil pH is the master control variable, rather than climate or climate factors.

Martin 2002

2009-09-29T13:45:00.000-04:00

Martin AP. 2002. Phylogenetic approaches for describing and comparing the diversity of microbial communities. Applied and Environmental Microbiology 68: 3673-3682.

This author presents a synthesis of a set of statistical techniques for detailed analysis of biodiversity in the context of microbial communities. One new test, the P-test (for phylogenetics) is combined with the FST test to generate inferences about the quantified levels of difference in community composition when examining multiple microbial communities.

A review of existing methods for quantifying diversity is provided first, rapidly pointing out the not-unlikely circumstances under which inter-community differences would be either under- or over-estimated in the absence of explicit phylogenetic inference. Other types of phylogenetic inference in this context are examined, but one main problem with techniques such as the Shannon-Wiener index is its dependency on accurate information about frequency of taxa. The P-test, novel to this paper as far as I can tell, avoids this pitfall, and instead is based on an examination of the covariance between a phylogeny and the distribution of taxa in communities.Figure 3 from Martin (2002). The basis of the P test is the covariance between which community a sequence was found in, and the positions of sequences on the phylogenetic tree.

The P test is combined with the FST test to examine the partitioning of sequence variation between communities. A P test on its own is not particularly informative, because it says little about how variation is partitioned between communities vs. the total pool.The 2x2 grid of comparison of P test and FST test results, from Figure 4 of Martin (2002). Each possible outcome of significance for the two tests allows inference about the evolutionary and ecological history of a particular situation of microbial communities.

The raw data for the P test is sequence data, typically 16S rDNA. This author advocates whole-gene sequences for comparison, to provide the maximum data and maximum compatibility between different studies, but acknowledges the trade-off between sequence length and number of sequences that can be produced. These are also the raw data for FST, but how those raw data are treated before going into each test varies.

Under the P test, the sequence data are used to construct a phylogeny, incorporating all sequence data from all communities. This phylogeny is set to equal total branch lengths from the root to the tips (the tips being the currently-measured sequences), and a null model of branching through time (lineage-per-time) is built. Then the community occurrence of each sequence is mapped onto the phylogeny, and the covariance calculated.

The FST test takes in Theta values as its meat of calculation. Theta is the total genetic variation in a sample, and in FST the grand total theta for all communities combined is compared to the average within-community theta for all communities under consideration.

This combined approach is intended to be complimentary to existing methods of examining microbial diversity, such as methods for estimating species richness, and methods for examining microbial phylogenies. I think the author’s own words at the beginning of the discussion section provide a good summary:

“In this study I used standard quantitative methods of analysis borrowed from population genetics and systematics for describing and comparing microbial communities. Information gained from analysis of DNA sequences provided the basis for statistical analysis of communities in ways that advance inferences about the processes that may govern the compositions and functions of microbial communities. Furthermore, the analytical approaches advocated here make it possible to accomplish broad comparisons of ecological communities. For instance, a comparison of lineage-per-time plots across a diverse set of ecosystems might reveal differences in the phylogenetic compositions of ecological communities that would be invisible with standard ecological statistics that ignore the magnitude of genetic differences among sampled sequences.”

I think I would like to use this approach in the analysis of microbial communities I will conduct based on soil samples from the polar desert. This method seems at this point like a useful way to quantify diversity across the gradient of latitude I will be covering.

Nannipieri et al. 2003

2009-09-28T16:20:00.000-04:00

Nannipieri P, Ascher J, Ceccherini MT, Landi L, Pietramellara G, Renella G. 2003. Microbial diversity and soil functions. European Journal of Soil Science 54: 655-670.

These authors present a review of the current state of knowledge about microbial diversity and ecosystem function such as organic matter decomposition in soils. They devote sections of the paper to the structure of soil as a habitat for microbes and other soil-dwelling organisms, methods of measuring microbial diversity, measuring soil functions, the current understanding of these methods and prominent results, and how these measures fit together in various contexts.

Unlike above-ground systems, soils appear to have no link between function and microbial diversity, or the direction and magnitude of the relationship varies considerably with which function is studied. General ecology theory and results suggest there should be a hump-shaped relationship between biodiversity (species richness and evenness) and productivity, such that productivity increases with diversity to some point, before declining. This relationship has not typically been found in soil systems, though there are relatively few studies of this relationship specifically in soils.
Measuring function in soils is complicated by the structure of soil. It is largely composed of non-living matter, some of which such as clay surfaces are capable of catalyzing reactions normally associated with living cells. In addition, these surfaces can adsorb large organic molecules such as enzymes and nucleic acids and protect them from degradation while still allowing some catalytic activity. Thus, even after all cells have been killed in a soil sample, enzymatic activity may be detectable. Distinguishing between biotic and abiotic chemical reactions in natural soil systems is therefore extremely difficult.

Measuring biodiversity in soil is not much easier than measuring function. Plate-count methods have been widely criticized because they will measure only culturable organisms, variously estimated to compose a small fraction of actual biodiversity. Countering this criticism, some researchers have suggested that the biomass, rather than species richness, of unculturable microbes is a minority, rendering plate counts of culturable species much more relevant to ecological studies. However, much attention has been focused recently on molecular methods, further divided into DNA-based techniques and fatty-acid based techniques.

DNA-based techniques deployed to study microbial diversity in soils often include a PCR step. However, DNA extraction methods for soil must balance several trade-offs, for example gram positive bacteria have very tough cell wall structures that require harsh treatment to break down and access their DNA. This same harsh treatment can shred DNA from less-tough cells to under 1kb fragments, which will often form chimeras during PCR, especially when using universal primers for such popular markers as 16s rDNA. Similarly, high-efficiency methods of DNA extraction and isolation are also efficient at extracting humic acids, which interfere with PCR. Despite these concerns, a large number of studies based on PCR of soil-derived DNA templates have been published, providing a large database of sequences for phylogenetic comparison.

Fatty-acid based techniques avoid the PCR-based concerns of DNA methods, but are less specific in their results: fatty acid composition is generally not species-specific the way DNA sequence data can be. However, techniques such as PFLA provide useful estimates of soil microbial biomass.

There is an ecological puzzle in the observed high biodiversity of near-surface soils. Two competing, though probably not mutually-exclusive hypotheses centre on a lack of competition among soil microbes. Under the first hypothesis, microbial microhabitats tend to be isolated from each other, preventing contact and competition. Community mixing occurs when water droplets bridge the gaps between soil aggregates, as during rainfall when soil pore spaces are filled. Countering this hypothesis is the observation that much of the near-surface soil environment is not especially prone to pore-drying, for example the plant root-soil interfaces, yet contains high species richness. The second hypothesis suggests that high specialization for organic substrates (i.e. microbe food) prevents competition among cells in close physical proximity. There are higher quantity and diversities of organic molecules in surface soils compared to greater depths, but flow channels such as cracks, fissures, and worm burrows also have high levels of organic molecules, and high microbial biomass, but do not show higher diversity than the surrounding bulk soil. The puzzle remains unsolved.

Much of the discussion of various measurements in this paper is of direct relevance to my own work. The various methods for assessing soil function, for example, are almost all measures of enzyme activity, which is precisely what my gas-flux measurements are as well. I intend to measure biodiversity, by molecular means, and the references and discussion here are valuable. Overall, this review paper does a good job of providing an overview of some issues I will also be exploring.

Freeman et al. 2009

2009-09-23T18:10:00.000-04:00

Freeman KR, Pescador MY, Reed SC, Costello EK, Robeson MS, Schmidt SK. 2009. Soil CO2 flux and photoautotrophic community composition in high-elevation, ‘barren’ soil. Environmental Microbiology 11: 674-686.

These authors measured photosynthetic carbon fixation and microbial community composition in sub-nival barren soils in the Colorado Front Range of the United States, at 40ºN latitude and approximately 3600m altitude. Like polar desert soils, these sub-nival soils lack conspicuous macrophytic vegetation (vascular plants and bryophytes) and are snow-covered for most of the year. Previous examinations of these systems had suggested the majority of carbon input to these soils was derived from wind-blown dust, but this study demonstrated a much larger input of carbon from in-situ photosynthesis.

Net carbon fixation was estimated by subtracting in-light measurements from in-dark measurements of CO2 flux. All measurements were made using an IRGA system with a 1.18L transparent chamber; dark measurements were made by covering the chamber with a dark, opaque cloth. After measurement of CO2 flux, one site was carefully dug up and transported to the laboratory for molecular-phylogenetic analysis.

The soil was divided into 2 depths: 0-2cm and 2-4cm, then DNA was extracted and PCR using universal bacterial primers for the 16s region was carried out, followed by sequencing. This generated more than 1000 sequences, in 4 bacterial divisions containing known photoautotrophic microorganisms, plus some sequences from eukaryotic green algae.

The most intriguing group of bacteria found were the Chloroflexi, an understudied group found in both depth layers. The taxa composition found in the deeper layer was highly different from the community found in the surface, light-receiving zone, and the authors suggest, based on a few studies done of Chloroflexi in hot-springs environments, that this group may use longer-wavelength light which penetrates deeper in soils. These authors do not make it, but this suggests to me the microphotoautotrophs in this system may be partitioning their environment in both space (depth) and spectrum (red).

This paper includes a large number of references and introductory descriptions for techniques and findings I will need to incorporate into the planning stages (at least) of my future studies in the polar desert. In particular, the molecular approach to the phylogenetics and biodiversity of the soil photoautotrophs seems both powerful and relatively uncomplicated. There are many procedures to carry out, to be sure, but the justification for each is clear, and the sequence of operations appears to be linear.

Uchida et al. 2002

2009-09-23T17:10:00.001-04:00

Uchida M, Muraoka H, Nakatsubo T, Bekku Y, Ueno T, Kanda H, Koizumi H. 2002. Net photosynthesis, respiration, and production of the moss Sanionia uncinata on a glacier foreland in the High Arctic, Ny-Ålesund, Svalbard. Arctic, Antarctic, and Alpine Research 34: 287-292.

These authors constructed a model of moss physiology that uses meteorological data to estimate productivity, based on data collected during one field season at Svalbard. In 2000, these authors measured the response of a common High Arctic moss species to water content, temperature, and light, then determined the relationship between those variables and available meteorological data, then applied previous-years meteorological data to their model and estimated previous-years productivity. These estimates suggest a great deal of variation in year-to-year productivity, driven largely by differences in water availability. Water content of fresh moss tissue was the single most important controlling variable in moss photosynthesis rates. The response to temperature was nearly flat between 7 and 23ºC, with near-freezing photosynthetic rates still a large fraction of maximum under saturating light conditions. Saturating light conditions were estimated at near 800µmol/m^2/s, which is not uncommon on sunny days in this environment.

The glacial foreground in question is at 79º North, but is not polar desert as it receives approximately 360mm of precipitation per year. The moss species studied is dominant in the local ecosystem, but appears to represent an intermediate successional stage, with high-productivity vascular plants replacing bryophytes in older sites in the area (i.e. further from the toe of the glacier).

Floyd et al. 2002

2009-09-22T15:45:00.000-04:00

Floyd R, Abebe E, Papert A, Blaxter M. 2002. Molecular barcodes for soil nematode identification. Molecular Ecology 11: 839-50.

These authors present a detailed description of and theory behind the MOTU concept. This analysis technique uses molecular sequence data to identify taxonomic units, hence the name Molecular Operational Taxonomic Unit. This paper uses the MOTU concept to examine and draw inferences about a collection of nematodes from a Scottish farm, finding high levels of species richness, and demonstrating a set of methods for rapid, inexpensive phylogenetics of a taxonomically-difficult group of animals.

Büdel et al. 2009

2009-09-22T12:00:00.000-04:00

Büdel B, Darienko T, Deutschewitz K, Dojani S, Friedl T, Mohr KI, Salisch M, Reisser W, Weber B. 2009. Southern African biological soil crusts are ubiquitous and highly diverse in drylands, being restricted by rainfall frequency. Microbial Ecology 57: 229-247.

These authors examined biological soil crusts (BSCs) along a 2000km transect running roughly north-south through Namibia and South Africa. A number of hypotheses relating to BSC composition, frequency, and succession were proposed and tested, with most hypotheses partly confirmed. In general, BSCs are an important and abundant component of these dryland ecosystems, and show patterns of biodiversity associated with biomass, as measured by chlorophyll-a concentrations and species counts.

The major finding of this study, as implied in the title, is that BSC distribution and composition is primarily controlled by patterns of rainfall, but not total rainfall. Species richness and successional stage of BSCs was highest in the winter rain zone, which has a shorter dry season though less total annual rainfall than the summer rain zone. This implies that most BSC organisms are limited by drought tolerance rather than annual water input.

This study is interesting to me for a number of reasons. First, it includes in the references a number of reviews of BSCs and methods to study them, such as protocols for measuring chlorophyll-a concentrations per square metre, and molecular methods for species richness estimation. Second, because BSCs are expected to be the major photosynthetic organisms in the polar desert, I need to know what patterns of their distribution and diversity I should expect. This paper’s Hypothesis 4, that biomass (and productivity) of BSCs increases with species richness, which was essentially confirmed, is of particular interest in this context, as it provides another layer of background expected pattern in addition to my general expectation of a species-richness gradient associated with latitude, particularly as one crosses Lancaster Sound north of Baffin Island. This paper provides some ideas for ways to measure species richness in BSCs, which (third) contribute strongly to the overall biodiversity of dryland regions and therefore will be interesting in their own right in studies of Arctic Biogeography.

Pilegaard et al. 2006

2009-09-21T18:10:00.000-04:00

Pilegaard K, Skiba U, Ambus P, Beier C, Bruggemann N, Butterbach-Bahl, Dick J, Dorsey J, Duyzer J, Gallagher M, Gasche R, Horvath L, Kitzler B, Leip A, Pihlatie MK, Rosenkranz P, Seufert G, Vesala T, Westrate H, Zechmeister-Boltenstern S. 2006. Factors controlling regional differences in forest soil emission of nitrogen oxides (NO and N2O). Biogeosciences 3: 651-661.

These (abundant) authors present an analysis of a large combined dataset covering NO and N2O emissions from a range of forest systems in Europe. The measurements contributing to this large dataset were continuous measures (at least daily, usually hourly or better) and run at least one year. This provides a high-quality dataset that includes variation induced by seasonality.

One of the most interesting findings in this study is a scale-dependent relationship between soil parameters and N2O emissions. Within-forests, soil temperature and moisture were highly predictive of N2O flux, but not at scales encompassing multiple forests in comparison. At larger spatial scales, stand age and C/N ratio were much better predictors.

Chen et al. 2008

2009-09-21T17:25:00.000-04:00

Chen Y, Dumont MG, Neufeld JD, Bodrossy L, Stralis-Pavese N, McNamara NP, Ostle N, Briones MJI, Murrell JC. 2008. Revealing the uncultivated majority: combining DNA stable-isotope probing, multiple displacement amplification and metagenomic analyses of uncultivated Methylocystis in acidic peatlands. Environmental Microbiology 10: 2609-2622.

These authors used a multiple-methods approach to isolate and identify DNA from a group of methanotrophic prokaryotes that have previously resisted attempts at culture. These microbes were previously estimated to be highly abundant in peatland soils, and were found in soils from a range of peatlands in Europe.

The three methods used to investigate these microbes were 1. a microarray built using sequences derived from a key enzyme in the methanotrophic process, 2. DNA-SIP, DNA Stable-Isotope Probing, used to examine DNA replicated with an injection of 13C-labelled CH4 (such that only methanotrophs would be able to use the carbon in their metabolisms), and 3. MDA, Multiple-Displacement Amplification to generate sufficient template DNA for fosmid-library construction and subsequent DGGE and cladistic analysis.

This triple-combined approach allowed the isolation, identification, and some basic phylogenetic analysis of a group of ecologically-important microbes previously unstudied in such a way. From my perspective, currently the most useful parts of this paper are the references (containing reviews of metagenomics and microarrays) and the methods section, as I may be attempting similar analyses of polar desert soils.

Barrett et al. 2006

2009-09-18T09:30:00.000-04:00

Barrett JE, Virginia RA, Wall DH, Cary SC, Adams BJ, Hacker AL, Aislabie JM. 2006. Co-variation in soil biodiversity and biogeochemistry in northern and southern Victoria Land, Antarctica. Antarctic Science 18: 535-548.

These authors examined soil biota at three sites across about 7 degrees of latitude in the drier part of Antarctica. The latitudinal gradient here covers a range of different ecosystems, from relatively wet, more northern and coastal systems to extremely arid and barren southern systems. Here, latitude is not studied for its effects on ecosystems; its effects on ecosystems are exploited to cover the widest available range of conditions. Within each of the three sites, one wet and one dry location were chosen a priori based on obvious surface features such as meltwater drainage channels and the presence of moss beds. At each location, a set of transects were laid out and samples were collected. To quote them directly:

"We investigated the structure (bacterial and metazoan diversity) and functioning (soil respiration) of soil communities and the influence of soil biogeochemical properties (organic matter, inorganic nutrients, physicochemical properties) on habitat suitability."

This is quite similar in many respects to my planned investigations in the Arctic polar desert. The list of molecular techniques used in this paper, for example, serves as a useful guide or checklist of the procedures I intend to use.

Soil invertebrates were investigated using both morphological and molecular techniques. There are only four metazoan phyla with any significant presence in Antarctica’s soils (Arthropoda, Nematoda, Rotifera, and Tardigrada), all of which are difficult to identify to species using standard morphological methods. While these authors were able to identify nematodes to species, rotifers, tardigrades, and mites were handled as MOTUs, molecular operational taxonomic units (Floyd et al. 2002). These MOTUs, based on ribosomal DNA sequences, were also used to generate a series of cladograms used to assess biodiversity at the research sites. Unfortunately, while the text descriptions of the methods and results are reasonably clear, the figures relating to the biodiversity and metazoan-molecular work are confusing and poorly described.

The results of the microbial analyses are broadly similar to the metazoan dataset. These authors were able to examine biodiversity at the level of microbes, and compare this diversity to both soil chemical characteristics such as water content and C:N ratio, and to the metazoan diversity. Perhaps surprisingly, they found no evidence to support the hypothesis of top-down control on bacterial populations by metazoan predators, as there was no correlation between DGGE and FAME-derived estimates of bacterial population size and species richness and cladogram and sugar-extract-derived estimates of nematode biodiversity. Bacterial diversity is described as not varying across sites, though community composition does. I take this to mean that while total species richness of bacteria (as measured by DGGE) was constant across sites, species turnover (Beta-diversity) was high. This is interesting to me, though not mentioned in the paper, because it seems to directly contradict the microbial-biogeography hypothesis of “everything is everywhere”. There is a mention in the description of this comparison of the importance of geography in structuring biodiversity, which reads to me like an opportunity to apply explicit geospatial techniques to their dataset.

Overall, while it is true that Antarctic soil ecosystems are extremely simple relative to other systems, there are a great many complex and variable interactions between even the few components of these systems, creating a great deal of complexity. This paper will be very useful to me in structuring some of my own investigations in the Arctic.

Ettema & Wardle 2002

2009-09-15T13:50:00.000-04:00

Ettema CH, Wardle DA. 2002. Spatial soil ecology. Trends in Ecology & Evolution 17: 177-183.

These authors review the growing use of explicit geospatial analysis techniques in soil biology. As this is a TREE article, there are several helpful boxes that explain fundamentals of geospatial analysis such as the terminology and key case studies. This is also a review article, so there are descriptions of various previous studies that include evidence useful in answering the questions set out in this paper. These questions are 1) What are the scales, patterns and causes of spatial variability in soil organism distributions? 2) What are the implications of spatial variability for the structure and function of soil communities? 3) How do spatial properties of the soil biota influence plant communities?

Regarding question 1, the scales and patterns of spatial variability in soil organisms range from 10s and 100s of metres down to millimetres. Studies of soil microbes including methanogenic Archaea have included soil corers of 1mm diameter (based on a hollow needle) and aggregations of organisms separated by distances of 2 to 4mm.

Soil communities and their influence on plant communities were found to be highly non-uniform, and show predictable though complex spatial patterns. However, while much was made of the role of individual plants (especially trees) to structure the soils around them and create spatial patterns of microbes and invertebrates on the same scale as the trees themselves are distributed, very little was made of the role that small-scale aggregations play in structuring larger patterns. This is surprising, given the highly biased view of soil processes in this paper and more generally in the soil science literature: soil is viewed as something that exists primarily to support plants, rather than a system of its own independent importance. That is the impression I have gotten, at least.

This paper is a very useful overview of geospatial analysis, and the reference list includes a number of similarly useful papers. In particular, further exploration of the statistics of semivariance patterns seems useful.

Garten et al. 2007

2009-09-14T17:15:00.001-04:00

Garten CT Jr., Kang S, Brice DJ, Schadt CW, Zhou J. 2007. Variability in soil properties at different spatial scales (1m-1km) in a deciduous forest ecosystem. Soil Biology & Biochemistry 39: 2621-2627.

These authors examined some of the fundamental assumptions of geospatial analysis as applied to soil properties, in a pair of transects that I strongly suspect have been used repeatedly for many studies in Tennessee (see, e.g. Zhou et al. 2008). One of the fundamental assumptions is of spatial autocorrelation, that is, samples in close proximity will be more similar to each other than samples separated by greater distances. In this study, this assumption was stated as the null hypothesis “there are no differences in variance at different spatial scales”; a rejection of this null hypothesis can be interpreted as support for the “common sense” (their wording) principle of spatial autocorrelation, at least among the spatial scales discussed here (i.e. metres to kilometres). This and other important assumptions of geospatial analysis come from a series of papers applying these principles to soils, which I should probably read soon.

The 11 soil variables examined in this paper were distinctly non-orthogonal in their relationships. Many of the variables were calculated directly from other variables, and the majority takes the form of either ratios (such as C-to-N) or fractions (such as silt content). The Principle Components Analysis (PCA) these authors conducted on their final, grand-total dataset indicated that the usual suspects of soil properties were important – soil Carbon, soil Nitrogen, and soil Texture are one way to summarize the first three PC variables.

I looked up and read this paper mainly because of the statistical tests used here and the discussion of them. They conducted 5 main statistical tests.
1. Bartlett’s test for equal variances at different distances.
2. Bartlett’s test is sensitive to non-normal data, so they also used the non-parametric Spearman’s Rank Correlation between coefficients of variation (100 x S.D./mean). The other reason a non-parametric test was used was that the functional relationship (linear vs. non-linear) between variance and sampling distance was unknown.
3. Mantel and Partial Mantel tests. These were the central analysis, I think, and provided most of the key results regarding the tests of the main hypotheses. Apparently, Burrough (1993) recommends semivariogram analysis, but the present data set was not amenable to such.
4. PCA, as mentioned above.
5. Power analysis. How many samples would they need to collect to be more certain of being close to the true mean value in their estimates?
Besides the PCA, which produced utterly unsurprising results, I think the statistical tests deployed here will serve as models for my own analysis of 2009 and putative 2010 datasets from the High Arctic. In particular, the Mantel tests and the Power analysis should be very useful in my own examinations.

Overall, the geospatial assumption of spatial autocorrelation was not very well supported by this study. Many soil properties appear to be highly variable at small spatial scales, such that samples collected within a few metres of each other are as variable as samples collected from up to a kilometre away, at least in a temperate forest ecosystem as studied here. This is particularly surprising in light of the consideration of the structure of such a forest, where individual trees presumably have strong impacts on soil properties within perhaps 5 to 10 metres of their trunks.

Zhou et al. 2008

2009-09-10T17:50:00.000-04:00

Zhou J, Kang S, Schadt CW, Garten CT Jr. 2008. Spatial scaling of functional gene diversity across various microbial taxa. Proceedings of the National Academy of Sciences of the USA 105: 7768-7773.

These authors used a microarray-based technique to estimate biodiversity of soil microbes across a pair of transects in a forest in Tennessee. Their analysis found rates of species turnover through space much lower than rates for macroorganisms such as “higher” plants and animals.

The species-area relationship, generalized to the Taxa-Area-Relationship (TAR), is S=cA^z, where S is the number of species, A is the area, c is the intercept in log-log space, and z is a measure of the rate of species turnover across space. Values of z for macroorganisms have been estimated close the theoretically derived value of 0.25, while previous estimates for microbes have been often much lower, but occasionally much higher. This study found a range of z-values, all a bit less than 0.1.

The key technique used in this study was the GeoChip, a microarray with nearly 25000 50-mer probes for more than 10000 genes in functional groups such as denitrification or heavy-metal resistance. As such, it represents an excellent tool for such investigations, because it reduces or avoids many of the microbe-diversity sampling artifacts such as undersampling that plague other methods.

A large fraction of the observed variation in sequences across the transects was unexplained. These authors speculate that a fraction of this unexplained variation may be driven by unexamined patterns and processes including biotic interactions (competition, trophic interaction), abiotic interactions (O2 concentrations, labile C pool), and microscale effects below 1m scales.

One interesting suggestion by these authors is to use metagenomic approaches to characterize key sequences of interest in a particular system, and then examine biodiversity using a microarray customized for these sequences. This is in line with what I was thinking in regards to using such techniques in polar desert soils – first, characterize what is there; second, look at biodiversity and patterns within diversity relating to groups of interest.

Broll et al. 1999

2009-09-04T12:00:00.000-04:00

Broll G, Tarnocai C, Mueller G. 1999. Interactions between vegetation, nutrients and moisture in soils in the Pangnirtung Pass area, Baffin island, Canada. Permafrost and Periglacial Processes 10: 265-277.

These authors examined soils from 6 pedons in Pangnirtung Pass, a north-south pass between mountains on Cumberland Peninsula. Three pedons were from moist soils, and three from dry soils. The moisture content drove a major difference in soil structure: dry soils are not cryoturbated, resulting in strong differences in nutrient content and mineralization rates.

The goal of the study was to compare in detail these differences between dry and moist soils. This seems very similar to my PhD goals surrounding examinations of Polar Desert soils. This study thus represents a possible template for some of my own investigations.

Bockheim 1979

2009-09-02T17:20:00.000-04:00

Bockheim JG. 1979. Properties and relative age of soils of southwestern Cumberland peninsula, Baffin island, N.W.T., Canada. Arctic and Alpine Research 11: 289-306.

This author sampled soils from more than 60 sites on the Cumberland peninsula of Baffin Island, mostly near the hamlet of Pangnirtung. This covered soils from two tundra vegetations (Dwarf shrub-sedge-moss-lichen on lowlands and coastal, stony sedge-moss-lichen in highlands and northern fjords) and the Polar Desert of Baffin island. The tundra soils ranged from mesic to subxeric, while the desert near Penny icecap was xeric. A similar gradient driven by latitude rather than altitude is referenced in Tedrow (1973).

Descriptions are made of the pH and various exchangeable and free minerals in the soils, along with how those components change with depth in each area. pH increases with depth, for example, especially in the Polar Desert. Phosphorus was found in surprisingly high levels in all soils. The active layer, or at least the layer above the permafrost, is much deeper than found on Ellesmere island, and appears to be deeper than 1m everywhere studied in this paper.

Niederberger et al. 2008

2009-09-02T12:00:00.001-04:00

Neiderberger TD, McDonald IR, Hacker AL, Soo RM, Barrett JE, Wall DH, Cary SC. 2008. Microbial community composition in soils of Northern Victoria Land, Antarctica. Environmental Microbiology 10: 1713-1724.

These authors present an analysis of a large collection of data regarding both microbial and metazoan biodiversity at relatively small scales in one part of Taylor Valley, Antarctica, one of the famous Dry Valleys. This contributes to both the Latitudinal Gradient Project, an international effort to characterize Antarctica, and to the biogeographical debate regarding the distribution and community assemblages of microbes and soil microfauna.

Biodiversity was higher than expected based on the physical characteristics of this extreme environment, and was much more variable at small (~200m) spatial scales. While the microbes identified by 16s sequences were not particularly surprising, the changes in community composition between study sites was high. This supports the hypothesis that extreme environments “select for” particular microbial physiologies, and that differences in soil physical features such as moisture and temperature are highly important, in distinct contrast to the “everything is everywhere” hypothesis of microbial biogeography.

NB October 1 2009: the “everything is everwhere” hypothesis (Beijerinck 1913) includes the second clause “the environment selects”, which implies my earlier impressions, above, are incorrect. This paper’s demonstration that extreme environments select for particular soil communities, and that local-scale variables such as moisture and temperature, rather than regional-scale variables such as climate factors, actually supports Beijerinck’s (1913) hypothesis, rather than countering it.

Rolston 1986

2009-04-20T19:54:00.001-04:00

Rolston DE. 1986. Gas diffusivity. Pp 1089-1102 in Methods of Soil Analysis Part 1: Physical and Mineralogical Methods 2nd Ed. ed. A Klute. American Society of Agronomy Inc. Soil Science Society of America Inc, Madison, WI.

This author presents a review of the principles and measurement methods of soil gas diffusion, including formulae and relevant calculations. At the heart of all considerations of soil gas diffusion is Fick’s law and the measurement of Dp, the soil gas diffusion constant for a particular gas. There are a wide range of laboratory methods for measuring gas diffusion, but all are based on measuring the passive movement of a target gas through a volume of soil, often by measuring the accumulation of the target gas in a chamber that initially lacks that gas.

Many variables will impact rates of gas diffusion; perhaps of greatest importance is the moisture content of the soil. Wet soils make take hours to measure, while dry soils only a few minutes. Temperature also has a strong effect, and this author urges the reporting of temperatures with all measures of soil diffusion. Additionally, a simple formula is presented that relates diffusion at one temperature to diffusion at a second temperature.

Further calculations surround the determination of Dp, and correcting for errors associated with the details of the measurement chamber. For example, a correction factor can be applied when the ratio of soil air volume to chamber volume is more than about 0.005; this corrects for the gas “stored” in the soil of the measurement apparatus.

This paper is the first of two adjacent chapters in this edited volume by this author, both dealing with soil gas movements. The calculations and formulae here will be very useful in attempts to calibrate the FTIR and the flux chambers.

Widén & Lindroth 2003

2009-03-17T19:53:00.001-04:00

Widén B, Lindroth A. 2003. A calibration system for soil carbon dioxide-efflux measurement chambers: description and application. Soil Science Society of America Journal, 67: 327-334.

These authors describe a system for the absolute calibration of CO2-flux from soils, for both open- and closed-chamber type systems. The basic design is a large box, topped with a layer of sand, into which CO2 is pumped at a known rate. The measurement chamber sits on top of the layer of sand, and the actual gas flux can be calculated and compared to that measured by the machine.

The system appears to be useful, and a significant improvement in the field. The authors caution that improvements need to be made to the system, cheifly in its size and consideration of the effects of water in the soil on gas flux variations.

Staal et al. 2001

2009-03-17T19:51:00.002-04:00

Staal M, te Lintel-Hekkert S, Harren F, Stal L. 2001. Nitrogenase activity in cyanobacteria measured by the acetylene reduction assay: a comparison between batch incubation and on-line monitoring. Environmental Microbiology 3(5): 343-351.

These authors present two methods for measuring nitrogenase activity in cyanobacteria, both based on continuous on-line measurement of ethylene produced by the reduction of acetylene by nitrogenase. One method relies on a gas chromatograph to detect ethylene, the other on a not-yet-commercially available laser system. Nitrogenase normally reduces N2 to NH3, but will also reduce other triple bonds such as that between the carbon atoms in acetylene, hence this measurement assay was developed in the late 1960s. The nitrogenase enzyme is inhibited by oxygen, but is very energy-intensive when reducing N2, thus cyanobacteria may fix Nitrogen in a manner dependent upon but separated from photosynthesis in either (at night) time or space (specialized cells).

Previous ethylene-based methods were based on incubations of cells in air-tight containers, for incubation periods sufficient to saturate nitrogenase with acetylene and accumulate sufficient ethylene for detection. Changes in O2 and CO2 concentrations during these hours-long incubations introduce conflating variables; O2 is depleted and CO2 accumulates in the dark, vice-versa in the light. CO2 concentration affects pH, while O2 inhibits nitrogenase and indirectly relates to available energy. In addition, long incubations will fail to detect any event occuring on a frequency shorter than the incubation time, such that processes occurring on time scales of seconds to minutes will not be registered. Finally, saturation of nitrogenase with acetylene eventually leads to nitrogen starvation and the synthesis of more nitrogenase.

In contrast, on-line methods involve the continuous flow of gas over the sample. This can be used to measure gas flux only when the system reaches a steady state (however, see the discussion of steady-state and non-steady-state modes in gas sampling chambers in Davidson et al., 2002). For nitrogenase-ethylene, this steady state may be reached as quickly as 1 minute under ideal, high-surface-area conditions. In addition, while O2 and CO2 concentrations are controlled during on-line measurement, ethylene cannot accumulate, thus only very low concentrations will be present.

Nitrogenase activity was higher under light-saturation conditions than in the dark, but became inhibitory with longer incubation times. Changes in nitrogenase activity with time and light level probably relate to both energy limitation in the dark and oxygen inhibition in the light. Changes relating to growth, internal rythms, or energy depletion only occurred after very long incubations, such as 24 hours. I am not certain how long-term energy depletion is distinct from short term energy limitation in the dark.

This paper suggests it may be possible for us to measure nitrogenase activity with acetylene and ethylene using the Gasmet FTIR system and its chambers.

Martin et al. 2004

2009-03-17T19:50:00.001-04:00

Martin JG, Bolstad PV, Norman JM. 2004. A carbon dioxide flux generator for testing infrared gas analyzer-based soil respiration systems. Soil Science Society of America Journal 68: 514-518.

These authors constructed a system for calibrating soil CO2 flux using a closed-chamber Infrared Gas Analyzer (IRGA), in this case a Li-Cor 6400. Two detectors were used: one to monitor the CO2 concentration inside the reservoir, and one as the test machine placed on top of the artificial soil on top of the reservoir. The basic construction was quite simple, compared with the systems of Butnor and Johnsen (2004) and Widen and Lindroth (2003). Essentially, this system is just a cylinder topped by a level layer of uniform glass beads. CO2 is added to the reservoir beneath, apparently by the simple method of exhaling into the input valve, and diffuses through the reservoir (mixed by a small fan) and through the glass bead layer.

Much of the refinement of this system concerns the placement of mixing fans to 1) ensure the reservoir is well-mixed but not pressurized and 2) disrupt the boundary layer on top of the glass beads. Boundary layer effects are blamed for some of the measurement error reported here.

The test system underestimated low fluxes, and overestimated high fluxes. These authors suggest that wind-speed differences inside vs. outside the closed chamber, and associated boundary layer differences, are the main drivers of these measurement errors. They also strongly caution that variation in the set point inside the reservoir has a large effect on measured flux rates. Under field conditions, with soil composed of smaller particles, these errors are expected to be less important.

Marion et al. 1997

2009-03-17T19:48:00.000-04:00

Marion GM, Henry GHR, Freckman DW, Johnstone J, Jones G, Jones MH, Levesque E, Molau U, Molgaard P, Parsons AN, Svoboda J, Virginia RA. 1997. Open-top designs for mainpulating field temperature in high-latitude ecosystems. Global Change Biology 3 (suppl. 1): 20-32.

These authors evaluated 4 different chamber designs under field conditions, examining many variables but focusing on temperature differences between the inside and outside of the chambers, and unintended ecological effects. The four designs were termed “ITEX corners”, “cone chambers”, “hexagon chambers”, and “plastic tent”. There were 6 field sites, 5 in the Arctic from Sweden to Canada, and 1 in Antarctica.

This paper represents one of the outcomes of a meeting that established the International Tundra Experiment (ITEX); at this meeting a list of requirements for long-term temperature manipulation devices was constructed, leading to these 4 designs and a requirement to measure ecological effects such as changes in snow accumulation or melting.

The results were fairly consistent across chamber designs. In general, open-top chambers cause fewer and less severe ecological side-effects than closed designs, but warm the surface of the soil by 1-2 degrees compared with up to 10 or 15 degrees for some closed designs. Side-effects of the open-top chambers included some shading and interception of PAR by the chamber materials, changes in moisture concentrations in the air immediately above the soil surface (though these may have been driven by changes in temperature), and the possibility of interference from animals, such as nutrient addition by birds perching on the chambers. However, CO2 concentrations were not affected by chambers.

One of these authors, GHR Henry, will be working with me this summer at Alexandra Fjord; this was also one of the study sites in this paper.

Mastepanov et al. 2008

2009-03-17T19:47:00.001-04:00

Mastepanov M, Sigsgaard C, Dlugokencky EJ, Houweling S, Strom L, Tamstorf MP, Christensen TR. 2008. Large tundra methane burst during onset of freezing. Nature 456: 628-631.

These authors describe a large emission of methane from a wet tundra site in the Greenland High Arctic, which occurred in late autumn and early winter as the ground froze. This burst of methane emission is of a similar magnitude to the total methane emission from this site during the growing season, and accounts for the previously observed “shoulder” of methane in autumn at high altitudes.

The site is Zackenberg Valley, in the north-east of Greenland at about 74ºN latitude. This site appears to be broadly similar to other High Arctic tundra meadows such as Truelove Lowlands (Devon Island) and Alexandra Fjord (Ellesmere Island) and large parts of northern Russia, with an active layer 20 to 100 cm thick. The growing season measurements here were similar to previous years, and similar to another study in Siberia.

These measurements were made using an automated methane-only laser-based system, that took readings of methane flux every hour, with a data-gathering time of 1 second. Late-season pulses of methane were not observed at lower-latitude sites, possibly because a deeper permafrost “floor” allows methane to diffuse down to deeper soil layers rather than being forced upwards. Spatial and temporal variablity of the freezing methane emissions were very high, suggesting the diffusion paths of methane squeezed out of the soil were dependent upon plant root systems and similar structures.

This paper is the reason my field season at Alexandra Fjord in 2009 may extend as late as August 25 (planned) or September 10 (worst-case scenario). The figures in this paper imply the difference between measured and modeled methane emissions became apparent approximately in late August.

Kammann et al. 2005

2009-03-17T19:45:00.000-04:00

Kammann C, Grünhage L, Grüters U, Janze S, Jäger H-J. 2005. Response of aboveground grassland biomass and soil moisture to moderate long-term CO2 enrichment. Basic and Applied Ecology 6: 351-365.

These authors present the results of the first 5 years of the GiFACE experiment in Germany (see Jäger et al. 2003). The major findings, as alluded to in the title, concern the response of aboveground biomass and soil moisture to moderate, year-round (but not 24-hour) CO2 enrichment in a temperate, mesic, semi-natural grassland ecosystem.

Compared to other similar studies, the GiFACE experiment found increased grass biomass under CO2 enrichment, no increase in forbs, and no changes in soil moisture. Other studies found less biomass increase, especially of grasses, generally increased forbs both by measures of diversity and by biomass, and generally increased soil moisture. Differences associated with GiFACE include the low CO2 step increase of 20% compared with much higher in other studies, such as doubling, the cutting frequency that is lower than most other studies, and the year-round CO2 enrichment compared with many studies enriching only during the active growing season. Additionally, the mix of species at Giessen may have been “right” for a strong biomass response, with an interaction from the low cutting frequency allowing these strongly responding grasses to increase above ground biomass to a large degree.

This paper suffers from an irritating flaw – a non-significant difference in annual biomass yield between enriched and control plots is described as “non-significantly higher”, an oxymoron. If it’s not significantly higher, it’s not higher.

Jäger et al. 2003

2009-03-17T19:44:00.001-04:00

Jäger HJ, Schmidt SW, Kammann C, Grünhage L, Müller C, Hanewald K. 2003. The University of Giessen free-air Carbon dioxide enrichment study: description of the experimental site and of a new enrichment system. Journal of Applied Botany – Angewandte Botanik 77: 117-127.

These authors describe the study site and technical details of the operation of a long-term experiment designed to study the impact of rising atmospheric Carbon dioxide concentrations, the GiFACE. In essence, this study system is unique and important, being the only such study currently ongoing in Europe, and is based on what appears to be the leading edge of relevant technology. At its heart, the system consists of a circular open-topped chamber into which CO2 is released under the control of a concentration monitor in the center. Release occurs at the upwind side of the ring, and consistently acheives the target enrichment of about 25% additional CO2 at 40cm above ground. Grassland canopy heights at this site and similar sites in Europe are almost never higher than 50cm.

Turbulence from the blowers disrupts microclimates in and near the rings during the ambient quiet at night, so the blowers are only run during daylight hours. Control plots without enrichment show the expected pattern of higher ambient CO2 concentrations at night, associated with nocturnal respiration and diurnal photosynthesis. These authors do not address the effects of this blower and enrichment schedule may have on a simulation of globally enriched atmospheric CO2.

This paper is the reference provided by Dr. Kammann to provide needed details for my application to the Canadian Food Inspection Agency to import soil samples from the GiFACE site to Canada.