Cattle grazing management affects soil microbial diversity and community network complexity in the Northern Great Plains.

Soil microbial communities play a vital role in the biogeochemical cycling and ecological functioning of grassland, but may be affected by common land uses such as cattle grazing. Changes in microbial diversity and network complexity can affect key ecosystem functions such as nutrient cycling. However, it is not well known how microbial diversity and network complexity respond to grazing in the Northern Great Plains. Consequently, it is important to understand whether variation in grazing management alters the diversity and complexity of grassland microbial communities. We compared the effect of intensive adaptive multi-paddock (AMP) grazing and conventional grazing practices on soil microbial communities using 16S/ITS amplicon sequencing. Samples were collected from grasslands in 13 AMP ranches and 13 neighboring, conventional ranches located across the Canadian prairies. We found that AMP grazing increased fungal diversity and evenness, and led to more complex microbial associations. Acidobacteria, Actinobacteria, Gemmatimonadetes, and Bacteroidetes were keystone taxa associated with AMP grazing, while Actinobacteria, Acidobacteria, Proteobacteria, and Armatimonadetes were keystone taxa under conventional grazing. Besides overall grazing treatment effects, specific grazing metrics like cattle stocking rate and rest-to-grazing ratio affected microbial richness and diversity. Bacterial and fungal richness increased with elevated stocking rate, and fungal richness and diversity increased directly with the rest-to-grazing ratio. These results suggest that AMP grazing may improve ecosystem by enhancing fungal diversity and increasing microbial network complexity and connectivity.

Timing and duration of access mat use impacts their mitigation of compaction effects from industrial traffic.

Grasslands are declining worldwide and are often impacted by industrial activities, including infrastructure development. Current best management practices for low-disturbance development on grasslands include the use of wooden access mats as temporary work platforms and roadways to mitigate soil compaction and rutting due to heavy traffic. We assessed the impacts of heavy traffic (TON), and the impacts of the same heavy equipment driven over top of access mats (AM), on soil physical, hydrological, and nutrient responses in sandy and loamy soils in the Dry Mixedgrass prairies over a 2-year period. We also assessed how the timing (early vs. late in the growing season) and duration (6 vs. 12 vs. 24 weeks) of AM and TON affected the same metrics. Compared to undisturbed soils, TON increased soil penetration resistance (15 cm depth) up to 93% in loamy and up to 101% in sandy soils, and decreased water infiltration rates from 53 to 71%, respectively. Notably, the negative impacts of TON on soil physical characteristics and hydrology were larger in sandy vs. loamy soils, and when moist soils were exposed to traffic early in the growing season. AMs were effective at mitigating soil compaction from industrial traffic when used on sandy soils. However, AM use increased the supply of total nitrogen and other plant macro- and micro-nutrients, particularly in soils subject to longer (12-24 wk) mat placement. Results indicate TON may have long-lasting effects on grassland (particularly sandy) soils, and that AM use represents an effective tool to mitigate traffic impacts. Further, early-season traffic should be avoided when soils are moist (whether with AM or not), and AMs should be placed on soils for limited durations (≤6 wk) to minimize potential nutrient losses.

Carbon and hydrogen isotopes of n-alkanes in soils reconstructed after mining disturbance.

Ecosystem reconstruction after mining disturbance is a challenge considering the multitude of factors that affect soil formation and revegetation. In the boreal forest of western Canada, peat material is often used as the organic amendment for land reclamation to upland forest. Carbon and water dynamics of peat-dominated ecosystems differ from natural upland forest soils. The objective of this work was to evaluate the evolution of soils reconstructed after mining disturbance using 13 C and 2 H analyses of n-alkane tracers. Ten soils from natural ecosystems were sampled (0-10 cm) and compared with 11 soils from novel ecosystems ranging in age from 0 to 30 yr, as well as a fresh peat sample. Soils supported different vegetation, including pine (Pinus spp.), aspen (Populus spp.), and white spruce [Picea glauca (Moench) Voss]. Despite overlaps for some individual n-alkanes, we found a dominance of n-C25 in reconstructed soils, also dominant in the peat material, and a dominance of n-C27 in natural soils, one of the dominant n-alkanes in natural forest vegetation. In addition, there was a significant difference in odd n-alkane δ2 H and δ13 C values between natural and reconstructed soils (p < .05). Differences in δ2 H values, more negative for reconstructed soils than for natural soils, were attributed to changes in soil moisture, from wetter peat-dominated soils to drier upland forests; among forest types, δ2 H values were most negative under pine vegetation. The δ13 C composition of odd n-alkanes, in particular n-C27 , was significantly related to tree age (p < .05). Overall, both 2 H and 13 C isotopic signatures of odd n-alkanes exhibited differences between natural and reconstructed soils. However, within the reconstructed soils, neither isotopic signature showed a clear evolution with age since reclamation.

Extraction and Analysis of Microbial Phospholipid Fatty Acids in Soils.

Phospholipid fatty acids (PLFAs) are key components of microbial cell membranes. The analysis of PLFAs extracted from soils can provide information about the overall structure of terrestrial microbial communities. PLFA profiling has been extensively used in a range of ecosystems as a biological index of overall soil quality, and as a quantitative indicator of soil response to land management and other environmental stressors. The standard method presented here outlines four key steps: 1. lipid extraction from soil samples with a single-phase chloroform mixture, 2. fractionation using solid phase extraction columns to isolate phospholipids from other extracted lipids, 3. methanolysis of phospholipids to produce fatty acid methyl esters (FAMEs), and 4. FAME analysis by capillary gas chromatography using a flame ionization detector (GC-FID). Two standards are used, including 1,2-dinonadecanoyl-sn-glycero-3-phosphocholine (PC(19:0/19:0)) to assess the overall recovery of the extraction method, and methyl decanoate (MeC10:0) as an internal standard (ISTD) for the GC analysis.

Understory Plant Community Composition Is Associated with Fine-Scale Above- and Below-Ground Resource Heterogeneity in Mature Lodgepole Pine (Pinus contorta) Forests.

Understory plant communities play critical ecological roles in forest ecosystems. Both above- and below-ground ecosystem properties and processes influence these communities but relatively little is known about such effects at fine (i.e., one to several meters within-stand) scales, particularly for forests in which the canopy is dominated by a single species. An improved understanding of these effects is critical for understanding how understory biodiversity is regulated in such forests and for anticipating impacts of changing disturbance regimes. Our primary objective was to examine the patterns of fine-scale variation in understory plant communities and their relationships to above- and below-ground resource and environmental heterogeneity within mature lodgepole pine forests. We assessed composition and diversity of understory vegetation in relation to heterogeneity of both the above-ground (canopy tree density, canopy and tall shrub basal area and cover, downed wood biomass, litter cover) and below-ground (soil nutrient availability, decomposition, forest floor thickness, pH, and phospholipid fatty acids (PLFAs) and multiple carbon-source substrate-induced respiration (MSIR) of the forest floor microbial community) environment. There was notable variation in fine-scale plant community composition; cluster and indicator species analyses of the 24 most commonly occurring understory species distinguished four assemblages, one for which a pioneer forb species had the highest cover levels, and three others that were characterized by different bryophyte species having the highest cover. Constrained ordination (distance-based redundancy analysis) showed that two above-ground (mean tree diameter, litter cover) and eight below-ground (forest floor pH, plant available boron, microbial community composition and function as indicated by MSIR and PLFAs) properties were associated with variation in understory plant community composition. These results provide novel insights into the important ecological associations between understory plant community composition and heterogeneity in ecosystem properties and processes within forests dominated by a single canopy species.

Tracking stable isotope enrichment in tree seedlings with solid-state NMR spectroscopy.

Enriching plant tissues with (13)C and (15)N isotopes has provided long-lasting, non-reactive tracers to quantify rates of terrestrial elemental fluxes (e.g., soil organic matter decomposition). However, the molecular location and level of isotope enrichment may differ among plant tissues. This factor is central to the integrity and interpretation of tracer data, but is seldom considered in experiments. We propose a rapid, non-destructive method to quantify molecular isotope allocation using solid-state (13)C and (15)N nuclear magnetic resonance spectroscopy. With this method, we tracked and quantified the fate of multiple pulses of (13)CO(2)(g) and K (15)NO(3)(l) in boreal tree seedling roots and leaves as a function of time. Results show that initial preferential (13)C carbohydrate enrichment in the leaves was followed by redistribution to more complex compounds after seven days. While (13)C allocation within the roots was uniform across molecules, (15)N results indicate an initial enrichment of amine molecules after two hours.

Biodegradation of naphthenic acids by rhizosphere microorganisms.

Naphthenic acids are components of most petroleums, including those found in the Athabasca Oil Sands of northeastern Alberta. Some naphthenic acids that are solubilized during bitumen extraction from oil sands are acutely toxic to a variety of organisms. Four-month enrichment cultures obtained from the rhizospheres of five plant species native to Alberta, and established with the addition of bitumen (0.5%) as the sole carbon source, revealed a high potential for aerobic degradation of a Merichem commercial preparation of naphthenic acids. Changes in the concentration and composition of the naphthenic acids mixtures during incubation were followed using high-performance liquid chromatography and gas chromatography-electron impact mass spectrometry. Concentrations did not significantly change in the sterile control, but they decreased by up to 90% after 10 days of incubation in the viable cultures. Lower molecular mass naphthenic acids were preferentially degraded, while the proportion of high molecular mass acids increased during incubation. By day 17, the most abundant ions were derived from cellular membranes, corresponding to an increase in microbial numbers in the cultures as naphthenic acids were metabolized. This study is the first to demonstrate the biodegradation potential of microorganisms from rhizosphere soils to biodegrade naphthenic acids.

Comparative analysis of cellulose preparation techniques for use with 13C, 14C, AND 18O isotopic measurements.

A number of operationally defined methods exist for pretreating plant tissues in order to measure C, N, and O isotopes. Because these isotope measurements are used to infer information about environmental conditions that existed at the time of tissue growth, it is important that these pretreatments remove compounds that may have exchanged isotopes or have been synthesized after the original formation of these tissues. In stable isotope studies, many pretreatment methods focus on isolating "cellulose" from the bulk tissue sample because cellulose does not exchange C and O isotopes after original synthesis. We investigated the efficacy of three commonly applied pretreatment methods, the Brendel method and two variants of the Brendel method, the Jayme-Wise method and successive acid/base/acid washes, for use on three tissue types (wood, leaves, roots). We then compared the effect of each method on C and O isotope composition (13C, 14C, 18O), C and N content, and chemical composition of the residue produced (using 13C nuclear magnetic resonance (NMR)). Our results raised concerns over use of the Brendel method as published, as it both added C and N to the sample and left a residue that contains remnant lipids and waxes. Furthermore, this method resulted in 18O values that are enriched relative to the other methods. Modifying the Brendel method by adding a NaOH step (wash) solved many of these problems. We also found that processed residues vary by tissue type. For wood and root tissues, the 13C NMR spectra and the 18O and 13C data showed only small differences between residues for the Jayme-Wise and modified Brendel methods. However, for leaf tissue, 13C NMR data showed that Jayme-Wise pretreatments produced residues that are more chemically similar to cellulose than the other methods. The acid/base/acid washing method generated 13C NMR spectra with incomplete removal of lignin for all tissues tested and both isotopic, and 13C NMR results confirmed that this method should not be used if purified cellulose is desired.