Poa secunda local collections and commercial releases: A genotypic evaluation.

The genetics of native plants influence the success of ecological restoration, yet genetic variability of local seed collections and commercial seed releases remains unclear for most taxa. Poa secunda, a common native grass species in Intermountain West grasslands and a frequent component of restoration seed mixes, is one such species. Here, we evaluate the genetic variation of local Poa secunda collections in the context of wild populations and commercial seed releases. We evaluated AFLP markers for seven Poa secunda collections made over a 4000-hectare area and four commercial releases (High Plains, MT-1, Opportunity, and Sherman). We compare the genetic distance and distribution of genetic variation within and between local collections and commercial releases. The extent and patterns of genetic variation in our local collections indicate subtle site differences with most variation occurring within rather than between collections. Identical genetic matches were usually, but not always, found within 5 m2 collection sites. Our results suggest that the genetic variation in two Poa secunda releases (High Plains and MT-1) is similar to our local collections. Our results affirm that guidelines for Poa secunda seed collection should follow recommendations for selfing species, by collecting from many sites over large individual sites.

Restoring ecological properties of acidic soils contaminated with elemental sulfur.

Elemental sulfur (S0) accumulates in the environment from anthropogenic sources as a byproduct from oil and gas refining and from trap and skeet shooting targets. Bacteria can oxidize S0 to H2SO4, which acidifies soil. We explored whether combinations of soil amendments can be used to remediate acidic soils contaminated with S0 by restoring soil chemistry, plant growth, and bacterial communities in a greenhouse. Results were compared to a contamination gradient in a field that had been limed with CaMg(CO3)2 two years prior. Amendments in the greenhouse included CaCO3 by itself, and in combination with fertilizer, compost, biochar, and chitin. Amended soils were incubated for one week and half of all containers were planted with Poa nevadensis. We sequenced bacterial DNA from a subset of amended soils and along the field gradient. CaCO3 additions in the greenhouse initially raised the pH of contaminated soil to values found in uncontaminated soils. However, pH decreased over time, which was likely caused by the oxidation of S0 to H2SO4. This was also apparent in the field, where CaCO3 additions raised pH to 4 but not to the desired value of 5 or higher. Plants in the greenhouse failed to grow in the unamended contaminated soil, but CaCO3 alone reduced concentrations of toxic cations and resulted in more plant growth than in the uncontaminated soil. CaCO3 also partially restored the bacterial communities in the greenhouse and in the field by increasing richness and diversity to near values found in uncontaminated soil, suggesting that bacteria can be resilient to prolonged acidic conditions. Organic amendments did not provide a significant benefit to restoration. This study demonstrates that acid neutralization alone can restore abiotic and biotic components and productivity of soils contaminated with S0, but multiple CaCO3 applications may be required to avoid future acidification.

Invasive Plants Rapidly Reshape Soil Properties in a Grassland Ecosystem.

Plant invasions often reduce native plant diversity and increase net primary productivity. Invaded soils appear to differ from surrounding soils in ways that impede restoration of diverse native plant communities. We hypothesize that invader-mediated shifts in edaphic properties reproducibly alter soil microbial community structure and function. Here, we take a holistic approach, characterizing plant, prokaryotic, and fungal communities and soil physicochemical properties in field sites, invasion gradients, and experimental plots for three invasive plant species that cooccur in the Rocky Mountain West. Each invader had a unique impact on soil physicochemical properties. We found that invasions drove shifts in the abundances of specific microbial taxa, while overall belowground community structure and functional potential were fairly constant. Forb invaders were generally enriched in copiotrophic bacteria with higher 16S rRNA gene copy numbers and showed greater microbial carbohydrate and nitrogen metabolic potential. Older invasions had stronger effects on abiotic soil properties, indicative of multiyear successions. Overall, we show that plant invasions are idiosyncratic in their impact on soils and are directly responsible for driving reproducible shifts in the soil environment over multiyear time scales. IMPORTANCE In this study, we show how invasive plant species drive rapid shifts in the soil environment from surrounding native communities. Each of the three plant invaders had different but consistent effects on soils. Thus, there does not appear to be a one-size-fits-all strategy for how plant invaders alter grassland soil environments. This work represents a crucial step toward understanding how invaders might be able to prevent or impair native reestablishment by changing soil biotic and abiotic properties.

Extreme soil acidity from biodegradable trap and skeet targets increases severity of pollution at shooting ranges.

Lead pollution at shooting ranges overshadows the potential for contamination issues from trap and skeet targets. We studied the environmental influence of targets sold as biodegradable by determining the components of the targets and sampling soils at a former sporting clay range. Targets comprised approximately 53% CaCO3, 41% S(0), and 6% modifiers, and on a molar basis, there was 2.3 times more S(0) than CaCO3. We observed a positive correlation between target cover and SO4(2-) (ρ=0.82, P<0.001), which indicated the oxidation of S(0) to H2SO4. Sulfate was negatively correlated with pH (ρ=-0.93, P<0.001) because insufficient CaCO3 existed in the targets to neutralize all the acid produced from S(0) oxidation. Plant cover decreased with decreasing soil pH (ρ=0.62, P=0.006). For sites that had pH values below 3, 24tons of lime per 1000tons of soil would be required to raise soil pH to 6.5. Lime-facilitated pH increases would be transitory because S(0) would continue to oxidize to H2SO4 until the S(0) is depleted. This study demonstrates that biodegradable trap and skeet targets can acidify soil, which has implications for increasing the mobility of Pb from shotgun pellets.

Consequences of seed origin and biological invasion for early establishment in restoration of a North American grass species.

Local, wild-collected seeds of native plants are recommended for use in ecological restoration to maintain patterns of adaptive variation. However, some environments are so drastically altered by exotic, invasive weeds that original environmental conditions may no longer exist. Under these circumstances, cultivated varieties selected for improved germination and vigor may have a competitive advantage at highly disturbed sites. This study investigated differences in early establishment and seedling performance between wild and cultivated seed sources of the native grass, Poa secunda, both with and without competition from the invasive exotic grass, Bromus tectorum. We measured seedling survival and above-ground biomass at two experimental sites in western Montana, and found that the source of seeds selected for restoration can influence establishment at the restoration site. Cultivars had an overall advantage when compared with local genotypes, supporting evidence of greater vigor among cultivated varieties of native species. This advantage, however, declined rapidly in the presence of B. tectorum and most accessions were not significantly different for growth and survival in competition plots. Only one cultivar had a consistent advantage despite a strong decline in its performance when competing with invasive plants. As a result, cultivated varieties did not meet expectations for greater establishment and persistence relative to local genotypes in the presence of invasive, exotic species. We recommend the use of representative local or regional wild seed sources in restoration to minimize commercial selection, and a mix of individual accessions (wild, or cultivated when necessary) in highly invaded settings to capture vigorous genotypes and increase the odds native plants will establish at restoration sites.

Spatial characterization of arbuscular mycorrhizal fungal molecular diversity at the submetre scale in a temperate grassland.

Although arbuscular mycorrhizal fungi (AMF) form spatially complex communities in terrestrial ecosystems, the scales at which this diversity manifests itself is poorly understood. This information is critical to the understanding of the role of AMF in plant community composition. We examined small-scale (submetre) variability of AMF community composition (terminal restriction fragment length polymorphism fingerprinting) and abundance (extraradical hyphal lengths) in two 1 m(2) plots situated in a native grassland ecosystem of western Montana. Extraradical AMF hyphal lengths varied greatly between samples (14-89 m g soil(-1)) and exhibited spatial structure at scales <30 cm. The composition of AMF communities was also found to exhibit significant spatial autocorrelation, with correlogram analyses suggesting patchiness at scales <50 cm. Supportive of overall AMF community composition analyses, individual AMF ribotypes corresponding to specific phylogenetic groups exhibited distinct spatial autocorrelation. Our results demonstrate that AMF diversity and abundance can be spatially structured at scales of <1 m. Such small-scale heterogeneity in the soil suggests that establishing seedlings may be exposed to very different, location dependent AMF communities. Our results also have direct implications for representative sampling of AMF communities in the field.

Evaluation of LSU rRNA-gene PCR primers for analysis of arbuscular mycorrhizal fungal communities via terminal restriction fragment length polymorphism analysis.

The efficacy of the LSU rDNA PCR primers FLR3 and FLR4 for discrimination of arbuscular mycorrhizal fungi communities via T-RFLP analysis was examined. Analysis of both public database and site-specific derived DNA sequences suggesting LSU rDNA-based T-RFLP analysis represents a valuable alternative for analysis of AMF communities.

Small-scale spatial heterogeneity of arbuscular mycorrhizal fungal abundance and community composition in a wetland plant community.

Although it has become increasingly clear that arbuscular mycorrhizal fungi (AMF) play important roles in population, community, and ecosystem ecology, there is limited information on the spatial structure of the community composition of AMF in the field. We assessed small-scale spatial variation in the abundance and molecular diversity of AMF in a calcareous fen, where strong underlying environmental gradients such as depth to water table may influence AMF. Throughout an intensively sampled 2 x 2 m plot, we assessed AMF inoculum potential at a depth of 0-6 and 6-12 cm and molecular diversity of the AMF community using terminal restriction fragment length polymorphism of 18S rDNA. Inoculum potential was only significantly spatially autocorrelated at a depth of 6-12 cm and was significantly positively correlated with depth to water table at both depths. Molecular diversity of the AMF community was highly variable within the plot, ranging from 2-14 terminal restriction fragments (T-RFs) per core, but the number of T-RFs did not relate to water table or plant species richness. Plant community composition was spatially autocorrelated at small scales, but AMF community composition showed no significant spatial autocorrelation. Saturated soils of calcareous fens contain many infective AMF propagules and the abundance and diversity of AMF inoculum is patchy over small spatial scales.

Phylogeny of arbuscular mycorrhizal fungi predicts community composition of symbiosis-associated bacteria.

Many physicochemical and biotic aspects of the soil environment determine the community composition of bacteria. In this study, we examined the effects of arbuscular mycorrhizal fungi, common symbionts of higher plants, on the composition of bacterial communities after long-term (7-8 years) enrichment culture in the presence of a plant host. We showed that the phylogeny of arbuscular mycorrhizal fungal isolates was a highly significant predictor of bacterial community composition, as assessed by cluster analysis, redundancy analysis and linear discriminant analysis of phospholipid fatty acid patterns. Numerous phospholipid fatty acids differed between the phylogenetic groupings; this pattern also held for fungal-origin phospholipid fatty acids and in a combined bacterial/fungal analysis, suggesting that categorizing phospholipid fatty acids into predominantly bacterial and fungal origin did not affect the overall outcome. The mechanisms underlying this observation could include substrate quality (and quantity) effects, interactions mediated by the host plant (e.g. rhizodeposition) and direct biotic interactions between arbuscular mycorrhizal fungi and bacterial populations. Our results suggest that aspects of arbuscular mycorrhizal fungal functions may be partially explained by the symbiosis-accompanying bacterial communities, a possibility that should be explicitly considered in studies examining the roles of arbuscular mycorrhizal fungal species diversity in soil and ecosystem processes.

Mycorrhizas and soil structure.

In addition to their well-recognized roles in plant nutrition and communities, mycorrhizas can influence the key ecosystem process of soil aggregation. Here we review the contribution of mycorrhizas, mostly focused on arbuscular mycorrhizal fungi (AMF), to soil structure at various hierarchical levels: plant community; individual root; and the soil mycelium. There are a suite of mechanisms by which mycorrhizal fungi can influence soil aggregation at each of these various scales. By extension of these mechanisms to the question of fungal diversity, it is recognized that different species or communities of fungi can promote soil aggregation to different degrees. We argue that soil aggregation should be included in a more complete 'multifunctional' perspective of mycorrhizal ecology, and that in-depth understanding of mycorrhizas/soil process relationships will require analyses emphasizing feedbacks between soil structure and mycorrhizas, rather than a uni-directional approach simply addressing mycorrhizal effects on soils. We finish the discussion by highlighting new tools, developments and foci that will probably be crucial in further understanding mycorrhizal contributions to soil structure.

Culture-independent analysis of midgut microbiota in the arbovirus vector Culicoides sonorensis (Diptera: Ceratopogonidae).

Differences in midgut microbial communities inhabiting Culicoides spp., insect vectors of virus pathogens, may affect the variation observed in the ability of these biting midges to propagate arthropod-borne viruses. As a first step toward addressing this hypothesis, midgut bacterial communities were compared between Culicoides species expected to be efficient and inefficient vectors of virus pathogens. We used 16S rDNA sequence and restriction fragment information to provisionally identify 36 bacterial genera from guts of wild adult female biting midges, Culicoides sonorensis Wirth and Jones and Culicoides variipennis (Coquillet), from two geographical locations. Bacterial identification was made by sequence analysis of 16S rDNA fragments and by terminal restriction fragment length polymorphism analysis of polymerase chain reaction-amplified 16S rDNA fragments from adult guts. Of 36 bacterial genera identified, 12 had been previously identified in other insects: Comomonas, Enterobacter, Klebsiella, Acinetobacter, Pseudomonas, Stenotrophomonas, Staphylococcus, Chryseobacterium, Moraxella, Acholeplasma, Flavobacterium, and Rickettsia, Significant differences in bacterial community composition were found between all three groups of wild adult females analyzed: live-trapped C. sonorensis, laboratory-emerged C. sonorensis, and laboratory-emerged C. variipennis.

Candidate division BD: phylogeny, distribution and abundance in soil ecosystems.

Oligonucleotide primers were designed and used to amplify partial 16S rDNA sequences of the recently identified bacterial group BD from four diverse soils. Phylogenetic analysis of 34 BD group sequences supports division-level status for the group and also indicates that the BD group consists of at least 3 subdivision-level groups. Sequence divergence (21%) amongst these BD group sequences was found to be near the average for bacterial division-level lineages. An intercalating dye-based quantitative PCR (qPCR) assay was used to quantify BD phylogenetic group 3 16S rDNA in Wyoming shortgrass steppe soils. Although BD phylogenetic group 3 16S rDNA sequence numbers were high, averaging 3 x 10(8) copies per g soil, no significant correlations were found between their abundance and soil organic matter content, inorganic N concentration, or pH. Based on microscopically estimated cell numbers and the range of rRNA operons per genome in the bacterial domain, we estimate that BD group 3 represents between 0.75% and 10.7% of the microbial population in a shortgrass steppe soil. Our results indicate that the BD group is widely distributed in the environment and present in significant numbers in Wyoming shortgrass steppe soils.

Spatial and temporal variability of bacterial 16S rDNA-based T-RFLP patterns derived from soil of two Wyoming grassland ecosystems.

Abstract Spatial and temporal variability of soil bacterial 16S rDNA terminal restriction fragment (TRF) size variation was evaluated in a homogeneous grassland (HG) dominated by the turf-forming grass Bouteloua gracilis and in a shrubland (SL) dominated by Artemisia tridentata (Wyoming big sagebrush). Temporal variability was also evaluated on the HG site over a growing season. No trends toward dissimilarity were detected with temporal (180 days) or spatial (up to 100 m) distance in the HG system. Terminal-restriction fragment length polymorphism (T-RFLP) profiles of the SL site exhibited pronounced small-scale spatial variability (<70 cm), although spatial analysis indicated weak spatial autocorrelation to distances greater than 36 cm. While shrub-induced nutrient localization was shown to significantly influence T-RFLP profiles, very little of the variability could be accounted for on the basis of spatial characteristics, suggesting that soil bacterial 16S rDNA composition of this site is predominantly controlled at scales other than those measured. Average dissimilarity values differed greatly between the two sites (0.27 and 0.59 for HG and SL sites, respectively). These results suggest that plant community structure strongly influences bacterial community composition in these semiarid ecosystems, highlighting the importance of considering spatial variability when designing field studies related to bacterial diversity in ecosystems having patchy or heterogeneous plant cover.