Soil freeze-thaw cycle experiments: Trends, methodological weaknesses and suggested improvements

Although freeze-thaw cycles can alter soil physical properties and microbial activity, their overall impact on soil functioning remains unclear. This review addresses the effects of freeze-thaw cycles on soil physical properties, microorganisms, carbon and nutrient dynamics, trace gas losses and higher organisms associated with soil. I discuss how the controlled manipulation of freeze-thaw cycles has varied widely among studies and propose that, despite their value in demonstrating the mechanisms of freeze-thaw action in soils, many studies of soil freeze-thaw cycles have used cycle amplitudes, freezing rates and minimum temperatures that are not relevant to temperature changes across much of the soil profile in situ. The lack of coordination between the timing of soil collection and the season for which freeze-thaw cycles are being simulated is also discussed. Suggested improvements to future studies of soil freeze-thaw cycles include the maintenance of realistic temperature fluctuations across the soil profile, soil collection in the appropriate season and the inclusion of relevant surface factors such as plant litter in the fall or excess water in the spring. The implications of climate change for soil freeze-thaw cycles are addressed, along with the need to directly assess how changes in soil freeze-thaw cycle dynamics alter primary production.     

Microbial activity and bacterial composition of H2-treated soils with net CO2 fixation

The effects of H₂ gas treatment of an agricultural soil cultivated previously with a mixture of clover (Trifolium pratense) and alfalfa (Medicago sativa) on CO₂ dynamics and microbial activity and composition were analyzed. The H₂ emission rate of 250 nmol H₂ g⁻¹ soil h⁻¹ was similar to the upper limit of estimated H₂ amounts emitted from N₂ fixing nodules into the surrounding soil ([Dong, Z., Layzell, D.B., 2001. H₂ oxidation, O₂ uptake and CO₂ fixation in hydrogen treated soil. Plant and Soil 229, 1-12.]). After 1 week of H₂ supply to soil samples simultaneously with H₂ uptake net CO₂ production declined continuously and this finally led to a net CO₂ fixation rate in the H₂-treated soil of 8 nmol CO₂ g⁻¹ soil h⁻¹. The time course of H₂ uptake and CO₂ fixation in the soils corresponded with an increase in microbial activity and biomass of the H₂-treated soil determined by microcalorimetric measurements, fluorescence in situ hybridization analysis (FISH) and DNA staining (DAPI). Shifts in the bacterial community structure caused by the supply of H₂ were recorded. While the H₂ treatment stimulated β-and γ-subclasses of Proteobacteria, it had no significant effect on α-Proteobacteria. In addition, FISH-detectable bacteria of the Cytophaga-Flavobacterium-Bacteroides phylum increased in numbers.     

Long-term impacts of infrequent biosolids applications on chemical and microbial properties of a semi-arid rangeland soil

A plot study was conducted to quantify long-term (>12 years) impacts of a single biosolids application, and short-term impacts (<2 years) of a repeated application, on semi-arid rangeland soil chemical and microbial parameters. In 2003 and 2004, plots which had received 0, 2.5, 5, 10, 21, or 30 Mg biosolids ha⁻¹ once in 1991 (long-term plots), or again in 2002 (short-term plots), were sampled and analyzed for soil chemical parameters, microbial biovolumes, C and N mineralization activities, Biolog EcoPlate substrate utilization potential, and plant productivity and tissue quality. Repeated applications temporarily exacerbated differences in soil chemical properties among treatments, but after 2 years, soil chemistry trends were similar between short-term and long-term plots. Soils which received a repeated application of 21 or 30 Mg biosolids ha⁻¹ had greater bacterial biovolumes and C and N mineralization activities. In long-term plots, mineralization activities were stimulated only at the highest rate. Biosolids-amended soil communities also utilized Biolog substrates more quickly compared to communities from control plots. Plant biomass increased, whereas plant diversity and plant C/N ratio decreased with increasing application rate for both short- and long-term plots. Infrequent biosolids application had positive ecosystem effects in terms of site management objectives, with relatively low extractable metal levels in soil and greater plant biomass and tissue quality despite reduced species richness.     

Application of para-nitrophenol (pNP) enzyme assays in degraded tropical soils

Enzyme activities have been used as indicators of soil quality and changes in biogeochemical function due to management or perturbations. The objective of this study was to answer a number of methodological questions regarding sampling schemes, sample handling recommendations, and assay procedures to facilitate the use of enzyme assays in the tropical highlands of East Africa. We used para-nitrophenol (pNP) based substrates for five enzymes: β-glucosidase, cellobiohydrolase, chitinase, acid phosphatase, and alkaline phosphatase. In the first experiment, we examined sampling procedures and compared the results of determining enzyme activities on a plot using composite or discrete samples. Composite samples usually had higher activities than the means of individual cores (P<0.05), but relative ranking of sites was the same if analyses were based on composite or discrete samples. In the second experiment, we examined the effects of storage time and conditions on enzyme activity. Enzyme activity degraded rapidly in frozen samples, but was better maintained in samples stored at 4 °C. Phosphatase and cellobiohydrolase activity declined after 14 days of storage, while the activity of the other enzymes remained close to the values of fresh samples for 28 or more days. In the third experiment, we examined the effect of the addition of an antiseptic, toluene, to prevent bacterial growth during the assay. We found no consistent toluene effect (P>0.4), probably because the assays were of short duration and microbial growth was minimized. Finally, we looked at the incubation time necessary to produce reliable results. Phosphatases, with relatively high activities could reliably be determined in 2 h incubations, but the other enzymes had much lower activities and required longer incubation times for reliable determination. For the enzymes we looked at, 4 h was a good standard time for determining the activity of even the lowest activity enzymes. The results of this study provide practical guidelines for applying these enzyme assays in the degraded tropical soils.     

Reciprocal transfer effects on denitrifying community composition and activity at forest and meadow sites in the Cascade Mountains of Oregon

In order to examine the effects of disturbance, vegetation type, and microclimate on denitrification and denitrifier community composition, experimental plots were established at the H. J. Andrews Experimental Forest in the Cascade Mountains of Oregon. Soil cores were reciprocally transplanted between meadow and forest and samples were collected after 1 and 2 years. Denitrifying enzyme activity (DEA) was measured using the acetylene block assay and terminal restriction length polymorphism profiles were generated with nosZ primers that target the gene coding for nitrous oxide reductase. Nitrate concentrations, C mineralization, and water content were measured to gain additional insights into soil properties controlling DEA. Meadow soils were significantly higher in DEA than forest soils, and the highest DEA levels were observed in cores transferred from the meadow into the forest. Nitrate concentrations were also different between forest and meadow soils, but did not correlate to DEA. DEA was higher in open versus closed cores, suggesting an association between denitrification and the rhizosphere. Denitrifier communities of undisturbed forest and meadow soils shifted through a 4-year period but remained distinct from each other. Similarly, denitrifier communities clustered by vegetation type of origin regardless of manipulation, suggesting that the overall denitrifier communities are well buffered against environmental changes.     

Increased N availability in grassland soils modifies their microbial communities and decreases the abundance of arbuscular mycorrhizal fungi

Two complementary studies were performed to examine (1) the effect of 18 years of nitrogen (N) fertilization, and (2) the effects of N fertilization during one growing season on soil microbial community composition and soil resource availability in a grassland ecosystem. N was added at three different rates: 0, 5.44, and 27.2 g N m⁻² y⁻¹. In both studies, Schizachyrium scoparium was the dominant plant species before N treatments were applied. Soil microbial communities from each experiment were characterized using fatty acid methyl ester (FAME) analysis. Discriminant analysis of the FAMEs separated the three N fertilizer treatments in both experiments, indicating shifts in the composition of the microbial communities. In general, plots that received N fertilizer at low or high application rates for 18 years showed increased proportions of bacterial FAMEs and decreased fungal FAMEs. In particular, control plots contained a significantly higher proportion of fungal FAMEs C18:1(cis9) and C18:2(cis9,12) and of the arbuscular mycorrhizal fungal (AMF) FAME, C16:1(cis11), than both of the N addition treatment plots. A significant negative effect of N fertilization on the AMF FAME, C16:1(cis11), was measured in the short-term experiment. Our results indicate that high rates of anthropogenic N deposition can lead to significant changes in the composition of soil microbial communities over short periods and can even disrupt the relationship between AMF and plants.     

Functional diversity of microbial communities in the mixed boreal plain forest of central Canada

This study was designed to examine whether or not specific tree species (Picea glauca, Picea mariana, Pinus banksiana, Populus tremuloides), their post-fire stand age, or their position in a successional pathway had any significant effect on the functional diversity of associated soil microbial communities in a typical mixed boreal forest ecosystem (Duck Mountain Provincial Forest, Manitoba, Canada). Multivariate analyses designed to identify significant biotic and/or abiotic variables associated with patterns of organic substrate utilization (assessed using the BIOLOG™ System) revealed the overall similarity in substrate utilization by the soil microbial communities. The five clusters identified differed mainly by their substrate-utilization value rather than by specific substrate utilization. Variability in community functional diversity was not strongly associated to tree species or post-fire stand age; however, redundancy analysis indicated a stronger association between substrate utilization and successional pathway and soil pH. For example, microbial communities associated with the relatively high pH soils of the P. tremuloides-P. glauca successional pathway, exhibited a greater degree of substrate utilization than those associated with the P. banksiana-P. mariana successional pathway and more acidic soils. Differences in functional diversity specific to tree species were not observed and this may have reflected the mixed nature of the forest stands and of their heterogeneous forest floor. In a densely treed, mixed boreal forest ecosystem, great overlap in tree and understory species occur making it difficult to assign a definitive microbial community to any particular tree species. The presence of P. tremuloides in all stand types and post fire stand ages has probably contributed to the large amount of overlap in utilization profiles among soil samples.     

Microbial activity affected by lime in a long-term no-till soil

Under conventional farming practices, lime is usually applied on the soil surface and then incorporated into the soil to correct soil acidity. In no-till (NT) systems, where lime is surface applied or only incorporated into the soil to very shallow depth, lime will likely not move to where it is required within reasonable time. Consequently, lime may have to be incorporated into the soil by mechanical means. The objective of this laboratory study was to characterize the effect of lime, incorporated to different depths, on chemical and biological soil properties in a long-term NT soil. Soil samples taken from the 0–5, 0–10, and 0–20 cm depths were analyzed in incubation studies for soil pH, nitrate, CO₂ respiration, and microbial biomass-C (MBC). Lime (CaCO₃) was applied at rates equivalent to 0, 4.4, 8.8, and 17.6 Mg ha⁻¹. Application of lime to both 0–10 and 0–20 cm depths increased soil pH from about 4.9 by 1, 1.7, and 2.8 units for the low, medium, and high liming rates, respectively. Soil nitrate increased over time and in proportion to liming rate, suggesting that conditions were favorable for N-mineralization and nitrification. Greater respiration rates and greater MBC found in lime-treated than in non-limed soils were attributed to higher soil pH. Faster turnover rates and increased mineralization of organic matter were found in lime-treated than in non-limed soils. These studies show that below-surface lime placement is effective for correcting soil acidity under NT and that microbial activity and nitrification can be enhanced.     

Effects of potato–grain rotations on soil erosion, carbon dynamics and properties of rangeland sandy soils

The potential for wind erosion in South Central Colorado is greatest in the spring, especially after harvesting of crops such as potato (Solanum tuberosum L.) that leave small amounts of crop residue in the surface after harvest. Therefore it is important to implement best management practices that reduce potential wind erosion and that we understand how cropping systems are impacting soil erosion, carbon dynamics, and properties of rangeland sandy soils. We evaluate the effects of cropping systems on soil physical and chemical properties of rangeland sandy soils. The cropping system included a small grain–potato rotation. An uncultivated rangeland site and three fields that two decades ago were converted from rangeland into cultivated center-pivot-irrigation-sprinkler fields were also sampled. Plant and soil samples were collected in the rangeland area and the three adjacent cultivated sites. The soils at these sites were classified as a Gunbarrel loamy sand (Mixed, frigid Typic Psammaquent). We found that for the rangeland site, soil where brush species were growing exhibited C sequestration and increases in soil organic matter (SOM) while the bare soil areas of the rangeland are losing significant amounts of fine particles, nutrients and soil organic carbon (SOM-C) mainly due to wind erosion. When we compared the cultivated sites to the uncultivated rangeland, we found that the SOM-C and soil organic matter nitrogen (SOM-N) increased with increases in crop residue returned into the soils. Our results showed that even with potato crops, which are high intensity cultivated cropping systems, we can maintain the SOM-C with a rotation of two small grain crops (all residue incorporated) and one potato crop, or potentially increase the average SOM-C with a rotation of four small grain crops (all residue incorporated) and one potato crop. Erosion losses of fine silt and clay particles were reduced with the inclusion of small grains. Small grains have the potential to contribute to the conservation of SOM and/or sequester SOM-C and SOM-N for these rangeland systems that have very low C content and that are also losing C from their bare soils areas (40%). Cultivation of these rangelands using rotations with at least two small grain crops can reduce erosion and maintain SOM-C and increasing the number of small grain crops grown successfully in rotation above two will potentially contribute to C and N sequestration as SOM and to the sequestration of macro- and micro-nutrients.