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New, non-chemical weed suppression methods are needed to support multi-tactic integrated weed management (IWM) programs that reduce reliance on herbicides. One such approach is to manage arbuscular mycorrhizal (AM) fungal communities to enhance their weed biocontrol. Successful implementation will require a better understanding of AM fungi-crop-weed relationships under field conditions, especially in the face of soil disturbances imposed by agricultural practices. We examined root AM fungal communities of four weed species and one model crop species (i.e., corn) in a field experiment with contrasting tillage and cover cropping treatments in Illinois, USA. Our objectives were to quantify the impact of AM fungi on the growth of corn and weed species, to assess influences of plant species, tillage, and cover cropping on AM fungal community structure, and to characterize the co-occurrence patterns of AM fungal taxa in weed and corn roots. Our results showed that effects of AM fungal taxa on corn and weed growth varied widely depending on plant species, and several AM fungal taxa had species-specific negative effects on weeds but not on corn. In addition, plant identity was the primary factor affecting root AM fungal community structure and co-occurrence patterns, whereas effects of tillage and cover cropping were relatively limited. We propose that AM fungal taxa that have selectively suppressive effects on weeds but not on corn are candidates for developing new strategies for weed biocontrol as part of IWM. Future approaches to weed management based on direct inoculation with AM fungal taxa or manipulation of AM fungal communities through specific management practices, are promising opportunities to increase the density of weed-suppressive AM fungal taxa in agroecosystems, and our methods provide important insights for AM fungal taxa selection.

Cover crops can supply and recycle soil nitrogen (N) in agroecosystems. With increasing interest in cover crop mixtures, farmers have questions about how N cycling is affected by individual species when planted alone or in mixtures. Field trials were conducted on two organic vegetable farms during the spring and summer of 2014–2016 to quantify biomass tissue N provisioned, soil nitrate (NO3-N) scavenged, and soil NO3-N mineralized from 12 species of cover crops (six spring and six summer species) planted in monoculture or five-way mixtures. All cover crops were compared to weedy and weeded (no cover crops or weeds) controls. Cover crops provisioned 28–33% more biomass tissue N than fallow weeds. Among spring cover crops, field pea produced the most biomass tissue N (131 ± 13 kg N ha−1; mean ± 1 SE) but that benefit disappeared when pea abundance was reduced in mixtures. Soil NO3-N to a depth of 0.2 m was reduced by 50–67% during cover crop and weed growth, regardless of treatment. Soil NO3-N availability approximately 1 month after incorporation of cover crop and weedy residues varied by treatment. Post-incorporation soil NO3-N was 17–19% greater in pea and soybean monocultures compared to mixtures; conversely, soil NO3-N was 17–19% lower following incorporation of buckwheat and sudangrass monocultures compared to mixtures. Results of this study demonstrate that cover crop N recycling services in near-surface soil are largely independent of species or mixture, but N provisioning services require careful management and species selection to ensure maximum biomass tissue N and timely N mineralization.

The invasive forest plant garlic mustard (Alliaria petiolata) has been shown to alter soil microbial communities in the northeastern part of its invaded range in the United States, and this disruption of soil communities may contribute to its invasion success. However, garlic mustard allelochemistry can vary with invasion age, and it is not clear whether garlic mustard’s impacts on soil microbes are consistent over its invaded range. Here, we compare the composition and diversity of soil fungal, bacterial, and archaeal communities among garlic mustard present, absent, and removed treatments in replicated blocks across five forests in the midwestern United States with relatively young garlic mustard invasions (approximately 17-26 years old, with consistent management). We collected samples in May and August, corresponding to garlic mustard active and senescent life history stages. While soil fungal and bacterial/ archaeal communities (based on ITS2 region and 16S rRNA gene DNA sequencing, respectively) differed significantly between different blocks/ forests and over time, we found no significant effect of garlic mustard treatment on soil microbial community composition or the relative abundance of mycorrhizal, saprotrophic, or pathogenic fungal guilds. The lack of garlic mustard impacts on the soil microbial community in recently invaded central Illinois forests suggests that these well-documented impacts in the northeastern United States and in older invasions cannot necessarily be generalized across all environmental contexts.

Integrated weed management (IWM) relies upon multiple chemical, physical, or biological weed management techniques to achieve an acceptable level of weed control. Agents that selectively suppress weeds but not crops and that can be manipulated in agriculture will be promising components for inclusion in IWM. We used a meta-analytic approach to investigate the potential of arbuscular mycorrhizal fungi (AMF) to contribute to IWM. We quantified the effect of crop and weed host status (strong and weak AMF hosts are divided in this study by a 10% root length colonization threshold), AMF diversity (single vs. mixed), and soil N and P fertility management on plant mycorrhizal growth responses (MGRs). Our results indicated that weak host weeds had consistently lower MGRs than strong host crops in both controlled and field conditions. Moreover, these differences in MGRs between weak host weeds and strong host crops were more pronounced under mixed AMF inoculum and low N and P nutrient availability. In contrast, MGR of strong host weeds was not different from strong host crops in general. However, we observed a wide range of MGRs among strong host weeds, some of which had much lower MGRs than strong host crops. In addition, in the presence of N and P fertilizers, strong host crops had a stronger positive response to AMF than strong host weeds. Thus, our meta-analysis indicates that AMF have potential to contribute to weed control by direct and indirect pathways: directly suppress weak host weeds, and indirectly suppress some strong host weeds mediating by competitive effects exerted by strong host crops. We suggest that management practices affecting AMF diversity and crop and weed mycorrhizal responses could be chosen to improve the contribution of AMF to IWM. Better understanding is needed of crop–weed–AMF interactions and management practices that enhance this form of weed management.

Managing the soil nitrogen (N) cycle is a major component of agricultural sustainability. Soil functional zone management (zonal management) is a novel agroecological strategy for managing row-crop agroecosystems. It may improve the efficiency of soil N cycling compared with conventional and no-tillage approaches, by managing the timing and location (crop row vs inter-row) of key soil N cycling processes. We compared N mineralization and availability during the period of maize peak N demand in crop rows and inter-rows in zonal management and conventional chisel plow tillage systems at four sites across the US Corn Belt over three growing seasons. Under zonal management, potential N mineralization and N availability during crop peak N demand were significantly greater in crop rows, where the majority of crop roots are found, compared with inter-rows. Averaged across all site-years, plant-available N in zonal management crop rows was 46 mg kg −1 compared with 21 mg kg −1 in inter-rows. In contrast, in conventional tillage, potential N mineralization and N availability were greater in inter-rows compared with crop rows; averaged across all site-years, plant-available N in conventional tillage crop rows was 24 mg kg −1 compared with 51 mg kg −1 in inter-rows. The results demonstrate that the active management of crop residues under zonal management can enhance the spatiotemporal efficiency of soil N cycling processes, by concentrating N mineralization and availability close to crop roots in synchrony with crop developmental needs. Zonal management therefore has potential to increase crop N-use efficiency compared with conventional tillage, and thereby reduce the impacts of row-crop agricultural production on water resources and greenhouse gas emissions that result from N leaching and denitrification.

Zonal tillage (e.g. ridge tillage, RT) separates management of row and inter-row positions, while non-zonal tillage (e.g. chisel plough, CP) applies management uniformly across a field. This may have large effects on soil hydrothermal properties, affecting soil processes and crop development. We examined the effects of RT versus CP on soil hydrothermal conditions under maize (Zea mays L.) at four sites spanning the US Corn Belt over two growing seasons (2012–2013). We also investigated whether RT, as a result of changes in hydrothermal conditions, could stimulate greater soil nitrogen (N) availability during peak maize N demand. We captured wide variation in soil types and climates, allowing us to generalise tillage effects across a large environmental gradient. Continuous hydrothermal measurements were taken in the centre of row and inter-row positions. Soil cores collected shortly after maize six leaf stage (V6) were analysed for plant-available N and potentially mineralisable N (PMN). We hypothesised: 1) in spring CP and RT both produce warm, dry seedbeds with equivalent accumulations of growing degree days (GDD), but later in season RT holds greater soil moisture, providing better conditions for cover or relay crop establishment; 2) Hydrothermal properties of RT rows are distinct from RT inter-rows, while CP rows and inter-rows are indistinguishable; 3) RT promotes greater soil N mineralisation and availability in crop rows compared with CP. Results largely confirmed all hypotheses. In early spring, rows were drier in RT than CP, and both were similar in warmth (i.e. in accumulated GDD). From V6 to tasselling, CP accumulated more GDD than RT in inter-rows, while row positions remained similar; RT maintained greater soil moisture across both positions. From tasselling to harvest, RT inter-rows held greater soil moisture than CP, but accumulated fewer GDD. Both tillage systems showed zonation of soil moisture between planting and harvest (inter-rows moister than rows); the magnitude of zonation was greatest in RT. Plant-available N and PMN were greater in RT compared with CP at V6, suggesting RT increases synchrony of soil N availability with crop requirements. The results demonstrate that zonal tillage can integrate the seedbed benefits of conventional tillage with increased soil moisture retention across a wide range of climates and soil types. Increased moisture retention may help buffer agricultural systems against drought, and improve seedbed conditions for cover and relay crops in late summer and early autumn, thus potentially improving both sustainability and production in these systems.

Particulate organic matter (POM) is considered an “active” source of nitrogen (N) in cultivated soils, responding readily to management and being more physically accessible to decomposers than mineral-associated forms of organic matter. However, there is increasing evidence that mineral-associated organic matter (MAOM) can also exhibit short-term changes to management that may impact plant and microbial N dynamics. In this study, we investigated how N within soil organic matter fractions responded to three years of tillage and cover crop treatments. We collected soils from a row-crop (maize-soybean rotation) field experiment replicated across three sites in the north central and mid-Atlantic United States: a high-soil organic matter site (3.1% soil organic carbon) in Illinois (IL) and two sites in Michigan (MI) and Pennsylvania (PA) with lower soil organic matter content (1.0% and 1.4% soil organic carbon, respectively). Management treatments included two levels of tillage (chisel plow and ridge tillage) and two levels of cover crop (with and without rye cover crop). Using an optimized sonication method coupled with particle size separation, we isolated and analyzed for N content free POM, occluded POM, a coarse silt fraction, and MAOM. Using partial least squares regression, we also explored broad cross-site relationships between soil organic matter (SOM) fractions, soil N availability, and crop performance. Both particulate and fine fractions responded to tillage and cover crop treatments, but patterns varied by site and fraction. In the low-SOM MI and PA soils, ridge tillage and cover cropping both increased N within POM fractions. The response to ridge tillage was most pronounced, with a 76% and 24% increase in occluded POM N content in MI and PA, respectively. In contrast, at the IL site (high-SOM), the inclusion of cover crops led to higher N, specifically within the fine fractions (coarse silt and MAOM). Cover cropping increased MAOM N content in IL by 24%. When analyzing all sites together, variables associated with fine fractions were more closely associated with N mineralization and crop performance. MAOM can be responsive to short-term management practices and, along with POM, may also be potential sources of N for crops.

Lespedeza cuneata is a non-native invasive legume that alters the soil bacterial community, associates promiscuously with rhizobia, and benefits more from rhizobial interactions compared to coexisting native Lespedeza in North America. We tested the hypothesis that native congeners differ in their nodule bacteria associations compared to L. cuneata. Plots with high L. cuneata invasion, low L. cuneata invasion with native Lespedeza species present, and uninvaded plots where native Lespedeza species existed without L. cuneata were sampled. Nodules were collected from all Lespedeza species present, and Chamaecrista fasciculata, a common native annual legume. Bacterial DNA from nodules was isolated and sequenced. Nodule bacterial composition differed significantly between hosts. L. cuneata nodules contained high frequencies of rhizobial DNA and low bacterial diversity, while native Lespedeza nodules contained lower rhizobial frequencies and higher non-rhizobial bacterial diversity. Specific non-rhizobial bacterial groups exhibited strong associations with native legumes and uninvaded sites. Significant differences exist in the nodule bacterial composition between native legumes and an introduced congener. The mechanism(s) and ecological importance of these differences remain unknown. These differences in bacterial associations could influence not only the competitive ability of the invader, but recovery of invaded sites as well.

Invasive plants often benefit from changes that they impose on soil microbes via positive plant–soil feedback, but the mechanisms that underlie these changes, and the legacy of their effects, remain poorly quantified. We investigated the impacts of an invasive annual grass, Microstegium vimineum, on the structure and functioning of soil microbial communities in a multi-year, field-based common garden experiment. Given previous reports that M. vimineum can both elevate nitrification rates in soil and benefit from enhanced nitrate availability, we sought to answer the following questions: 1) Does M. vimineum alter the abundance or composition of soil nitrifying microbial communities (ammonia oxidizing archaea and bacteria, AOA and AOB, respectively)? 2) Are such effects reversible or do soil legacy effects persist after M. vimineum is no longer present? After three years, invaded plots had greater AOA abundances than uninvaded native dominated plots, as well as different AOA community structure. However, after seven years, and following a period of M. vimineum replacement by native plants in the invaded plots, AOA abundances and nitrification rates declined toward levels found in uninvaded plots. Collectively, our results suggest that while the impacts of M. vimineum invasions on nitrogen cycling likely relate to their association with AOA, these effects may not persist if M. vimineum declines over time and native plants and their associated microbes are able to re-establish.

Lespedeza cuneata is an introduced legume that is invasive in the tallgrass prairie system and open woodlands of North America. This system includes native Lespedeza species that coexist with L. cuneata, including L. capitata and L. virginica. Previous research has indicated that L. cuneata exudates have profound influences on soil biological functions. The goal of this current research was to identify and then compare exudate chemicals from L. cuneata, L. capitata, L. virginica, and common tallgrass prairie grasses. Exudates from Lespedeza species were found to be distinct from the grasses. In particular, L. cuneata and, to a lesser extent, a subset of L. virginica could be differentiated based on the presence of five unique compounds. Two low molecular weight compounds were identified via gas chromatography/mass spectrometry (GC/MS) and tentatively identified as benzophenone and 1,4-diacetylbenzene. Three higher molecular weight compounds were identified by liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS), with possible relationships to a 3 sterol, phosphatidylglycerol, and the 19-nor analogues of vitamin D. The relative abundance of these compounds were found to be greater in the invasive species, L. cuneata, than in the native noninvasive species, L. virginica. These results are an integral step in further understanding through chemical exudation how L. cuneata can be benefiting from greater rhizobial associations.

Identification of microbes that actively utilize root exudates is essential to understand plant-microbe interactions. To identify active root exudate-utilizing microorganisms associated with switchgrass - a potential bioenergy crop - plants were labelled in situ with (13) CO2 , and 16S and 18S rRNA genes in the (13) C-labelled rhizosphere DNA were pyrosequenced. Multi-pulse labelling for 5 days produced detectable (13) C-DNA, which was well separated from unlabelled DNA. Methylibium from the order Burkholderiales were the most heavily labelled bacteria. Pythium, Auricularia and Galerina were the most heavily labelled eukaryotic microbes. We also identified a Glomus intraradices-like species; Glomus members are arbuscular mycorrhizal fungi that are able to colonize the switchgrass root. All of these heavily labelled microorganisms were also among the most abundant species in the rhizosphere. Species belonging to Methylibium and Pythium were the most heavily labelled and the most abundant bacteria and eukaryotes in the rhizosphere of switchgrass. Our results revealed that nearly all of the dominant rhizosphere bacterial and eukaryotic microbes were able to utilize root exudates. The enrichment of microbial species in the rhizosphere is selective and mostly due to root exudation, which functions as a nutrition source, promoting the growth of these microbes.

Increasingly, funding agencies are investing in integrated and transdisciplinary research to tackle “grand challenge” priority areas, critical for sustaining agriculture and protecting the envi -ronment. Coordinating multidisciplinary research teams capable of addressing these priority areas, however, presents its own unique set of chal-lenges, ranging from bridging across multiple dis -ciplinary perspectives to achieve common ques-tions and methods to facilitating engagement in holistic and integrative thinking that promotes linkages from scholarship to societal needs. We propose that structural equation modeling (SEM) can provide a powerful framework for synergizing multidisciplinary research teams around grand challenge issues. Structural equation modeling can integrate both visual and statistical expres-sion of complex hypotheses at all stages of the research process, from planning to analysis. Three elements of the SEM framework are par-ticularly beneficial to multidisciplinary research teams; these include (i) a common graphical lan-guage that transcends disciplinary boundaries, (ii) iterative, critical evaluation of complex hypoth -eses involving manifest and latent variables and direct and indirect interactions, and (iii) enhanced opportunities to discover unanticipated interac-tions or causal pathways as empirical data are tested statistically against the model. Using our ongoing multidisciplinary, multisite field inves-tigation of climate change adaptation and miti-gation in annual row crop agroecosystems as a case study, we demonstrate the value of the SEM framework for project design, coordination, and implementation and provide recommendations for its broader application as a means to more effectively engage and address issues of critical societal concern.R.G. Smith, L.W. Atwood, A.B. Daly, and A.S. Grandy, Dep. of Natural Resources and the Environment, Univ. of New Hampshire, Durham, NH 03824; A.S. Davis, USDA-ARS Global Change and Photosynthesis Research Unit, Urbana, IL 61801; N.R. Jordan and P. Ewing, Dep. of Agronomy and Plant Genetics, Univ. of Minnesota, St. Paul, MN 55108; M.C. Hunter, and D.A. Mortensen, Dep. of Plant Science, The Pennsylvania State Univ., University Park, PA 16802; R.T. Koide, Dep. of Biology, Brigham Young Univ., Provo, UT 84602; D. Kane and S.S. Snapp, Dep. of Plant, Soil and Microbial Sciences, Michigan State Univ., East Lansing, MI 48824; M. Li, Dep. of Crop Sciences, Univ. of Illinois, Urbana, IL 61801; Y. Lou and A.C. Yannarell, Dep. of Natural Resources and Environmental Sciences, Univ. of Illinois, Urbana, IL 61801; K.A. Spokas, USDA-ARS Soil and Water Management Research Unit, St. Paul, MN 55108. Received 19 July 2013. *Corresponding author (richard.smith@unh.edu).

Feedback loops involving soil microorganisms can regulate plant populations. Here, we hypothesize that microorganisms are most likely to play a role in plant–soil feedback loops when they possess an affinity for a particular plant and the capacity to consistently affect the growth of that plant for good or ill. We characterized microbial communities using whole-community DNA fingerprinting from multiple "home-and-away" experiments involving giant ragweed (Ambrosia trifida L.) and common sunflower (Helianthus annuus L.), and we looked for affinity–effect relationships in these microbial communities. Using canonical ordination and partial least squares regression, we developed indices expressing each microorganism's affinity for ragweed or sunflower and its putative effect on plant biomass, and we used linear regression to analyze the relationship between microbial affinity and effect. Significant linear affinity–effect relationships were found in 75 % of cases. Affinity–effect relationships were stronger for ragweed than for sunflower, and ragweed affinity–effect relationships showed consistent potential for negative feedback loops. The ragweed feedback relationships indicated the potential involvement of multiple microbial taxa, resulting in strong, consistent affinity–effect relationships in spite of large-scale microbial variability between trials. In contrast, sunflower plant–soil feedback may involve just a few key players, making it more sensitive to underlying microbial variation. We propose that affinity–effect relationship can be used to determine key microbial players in plant–soil feedback against a low "signal-to-noise" background of complex microbial datasets. Electronic supplementary material The online version of this article (doi:10.1007/s00248-013-0349-2) contains supplementary material, which is available to authorized users.

Rhizosphere bacteria have significant contributions to crop health, productivity and carbon sequestration. As maize (Zea mays) is an important economic crop, its rhizosphere bacterial communities have been intensively investigated using various approaches. However, low-resolution profiling methods often make it difficult to understand the complicated rhizosphere bacterial communities and their dynamics. In this study, we analyzed growth-stage related dynamics of bacterial community structures in the rhizosphere of maize using the pyrosequencing method, which revealed an assembly of bacteria enriched in the rhizosphere. Our results revealed that the rhizosphere of maize was preferentially colonized by Proteobacteria, Bacteroidetes and Actinobacteria, and each bacterial phylum was represented by one or two dominating subsets of bacterial groups. Dominant genera enriched in the rhizosphere included Massilia, Burkholderia, Ralstonia, Dyella, Chitinophaga and Sphingobium. Rhizosphere bacterial community structures significantly changed through different growth stages at lower taxonomic ranks (family, genus and OTU levels). Genera Massilia, Flavobacterium, Arenimonas and Ohtaekwangia were relatively abundant at early growth stages, while genera Burkholderia, Ralstonia, Dyella, Chitinophaga, Sphingobium, Bradyrhizobium and Variovorax populations were dominant at later stages. Comparisons of pyrosequencing data collected in Illinois, USA in this study with the available data from Braunschweig, Germany indicated many common bacterial inhabitants but also many differences in the structure of bacterial communities, implying that some site-specific factors, such as soil properties, may play important roles in shaping the structure of rhizosphere bacterial community.

A low-diversity microbial community, dominated by the γ-proteobacterium Halomonas sulfidaeris, was detected in samples of warm saline formation porewater collected from the Cambrian Mt. Simon Sandstone in the Illinois Basin of the North American Midcontinent (1.8 km/5872 ft burial depth, 50°C, pH 8, 181 bars pressure). These highly porous and permeable quartz arenite sandstones are directly analogous to reservoirs around the world targeted for large-scale hydrocarbon extraction, as well as subsurface gas and carbon storage. A new downhole low-contamination subsurface sampling probe was used to collect in situ formation water samples for microbial environmental metagenomic analyses. Multiple lines of evidence suggest that this H. sulfidaeris-dominated subsurface microbial community is indigenous and not derived from drilling mud microbial contamination. Data to support this includes V1-V3 pyrosequencing of formation water and drilling mud, as well as comparison with previously published microbial analyses of drilling muds in other sites. Metabolic pathway reconstruction, constrained by the geology, geochemistry and present-day environmental conditions of the Mt. Simon Sandstone, implies that H. sulfidaeris-dominated subsurface microbial community may utilize iron and nitrogen metabolisms and extensively recycle indigenous nutrients and substrates. The presence of aromatic compound metabolic pathways suggests this microbial community can readily adapt to and survive subsurface hydrocarbon migration.

This study aimed to determine if biotic contaminants originating from pig production farms are disseminated into soil and groundwater microbial communities. A spatial and temporal sampling of soil and groundwater in proximity to pig production farms was conducted, and quantitative PCR (Q-PCR) was utilized to determine the abundances of tetracycline resistance genes (i.e., tetQ and tetZ ) and integrase genes (i.e., intI1 and intI2 ). We observed that the abundances of tetZ , tetQ , intI1 , and intI2 in the soils increased at least 6-fold after manure application, and their abundances remained elevated above the background for up to 16 months. Q-PCR further determined total abundances of up to 5.88 × 10 9 copies/ng DNA for tetZ , tetQ , intI1 , and intI2 in some of the groundwater wells that were situated next to the manure lagoon and in the facility well used to supply water for one of the farms. We further utilized 16S rRNA-based pyrosequencing to assess the microbial communities, and our comparative analyses suggest that most of the soil samples collected before and after manure application did not change significantly, sharing a high Bray-Curtis similarity of 78.5%. In contrast, an increase in Bacteroidetes and sulfur-oxidizing bacterial populations was observed in the groundwaters collected from lagoon-associated groundwater wells. Genera associated with opportunistic human and animal pathogens, such as Acinetobacter , Arcobacter , Yersinia , and Coxiella , were detected in some of the manure-treated soils and affected groundwater wells. Feces-associated bacteria such as Streptococcus , Erysipelothrix , and Bacteroides were detected in the manure, soil, and groundwater ecosystems, suggesting a perturbation of the soil and groundwater environments by invader species from pig production activities.

Summary Biomass production for bioenergy may change soil microbes and influence ecosystem properties. To explore the impact of different bioenergy cropping systems on soil microorganisms, the compositions and quantities of soil microbial communities (16S rRNA gene) and N-cycling functional groups (nifH, bacterial amoA, archaeal amoA and nosZ genes) were assessed under maize, switchgrass and Miscanthus x giganteus at seven sites representing a climate gradient (precipitation and temperature) in Illinois, USA. Overall, the site-to-site variation in community composition surpassed the variation due to plant type, and microbial communities under each crop did not converge on a ‘typical’ species assemblage. Fewer than 5% of archaeal amoA, bacterial amoA, nifH and nosZ OTUs were significantly different among these crops, but the largest differences observed at each site were found between maize and the two perennial grasses. Quantitative PCR revealed that the abundance of the nifH gene was significantly higher in the perennial grasses than in maize, and we also found significantly higher total N in the perennial grass soils than in maize. Thus, we conclude that cultivation of these perennial grasses, instead of maize, as bioenergy feedstocks can improve soil ecosystem nitrogen sustainability by increasing the population size of N-fixing bacteria.

A combination of culture-dependent and culture-independent methods was used to assess bacterial diversity at different depths within a former solid waste dump in Medellin, Colombia. Sampling sites included a densely populated area, which is built upon 40 m of solid waste (domestic, industrial, agricultural, and medical). The soil and leachate contain high levels of contaminants and the natural soil is highly disturbed with solid anthropogenic materials, disrupting natural aggregation and resulting in a loose, porous matrix with irregular aggregate structure. The unusual physical structure and contaminant levels at the site made it unclear if the indigenous bacterial community would possess the complexity commonly observed for natural soils, and thus may limit potential for remediating the site using monitored natural attenuation. Bacterial diversity patterns were determined through 16S-TTGE and T-RFLP at depths of 0, 10, 20 and 30 m. Abundance and diversity patterns, as estimated by number and intensity of terminal restriction fragments and TTGE bands, varied among the 4 different depths, showing more complex patterns in deeper samples (20 and 30 m), which also contained greater concentrations of organic carbon. General diversity patterns were dominated by the phylum Proteobacteria (λ, β, and α divisions). These findings were reinforced by analysis of the culturable fraction able to use n-hexadecane as sole carbon source, in which the genera Acinetobacter sp. (λ-Proteobacteria) was dominant. This research offers new clues regarding bacterial diversity patterns through different depths in polluted environments with unique physicochemical conditions, suggesting that bacterial diversity profiles may be highly influenced by the nature of pollutants present. Additionally, results imply that the culturable fraction at the site has a very important role in the community.

Reverse complementary DNA sequences - sequences that are inadvertently given backwards with all purines and pyrimidines transposed - can affect sequence analysis detrimentally unless taken into account. We present an open-source, high-throughput software tool -v-revcomp (http://www.cmde.science.ubc.ca/mohn/software.html) - to detect and reorient reverse complementary entries of the small-subunit rRNA (16S) gene from sequencing datasets, particularly from environmental sources. The software supports sequence lengths ranging from full length down to the short reads that are characteristic of next-generation sequencing technologies. We evaluated the reliability of v-revcomp by screening all 406 781 16S sequences deposited in release 102 of the curated SILVA database and demonstrated that the tool has a detection accuracy of virtually 100%. We subsequently used v-revcomp to analyse 1 171 646 16S sequences deposited in the International Nucleotide Sequence Databases and found that about 1% of these user-submitted sequences were reverse complementary. In addition, a nontrivial proportion of the entries were otherwise anomalous, including reverse complementary chimeras, sequences associated with wrong taxa, nonribosomal genes, sequences of poor quality or otherwise erroneous sequences without a reasonable match to any other entry in the database. Thus, v-revcomp is highly efficient in detecting and reorienting reverse complementary 16S sequences of almost any length and can be used to detect various sequence anomalies.

In this study, we evaluated the specificity, distribution, and sensitivity of Prevotella strain-based (PF163 and PigBac1) and methanogen-based (P23-2) PCR assays proposed to detect swine fecal pollution in environmental waters. The assays were tested against 222 fecal DNA extracts derived from target and nontarget animal hosts and against 34 groundwater and 15 surface water samples from five different sites. We also investigated the phylogenetic diversity of 1,340 “ Bacteroidales ” 16S rRNA gene sequences derived from swine feces, swine waste lagoons, swine manure pits, and waters adjacent to swine operations. Most swine fecal samples were positive for the host-specific Prevotella- based PCR assays (80 to 87%), while fewer were positive with the methanogen-targeted PCR assay (53%). Similarly, the Prevotella markers were detected more frequently than the methanogen-targeted assay markers in waters historically impacted with swine fecal contamination. However, the PF163 PCR assay cross-reacted with 23% of nontarget fecal DNA extracts, although Bayesian statistics suggested that it yielded the highest probability of detecting pig fecal contamination in a given water sample. Phylogenetic analyses revealed previously unknown swine-associated clades comprised of clones from geographically diverse swine sources and from water samples adjacent to swine operations that are not targeted by the Prevotella assays. While deeper sequencing coverage might be necessary to better understand the molecular diversity of fecal Bacteroidales species, results of sequence analyses supported the presence of swine fecal pollution in the studied watersheds. Overall, due to nontarget cross amplification and poor geographic stability of currently available host-specific PCR assays, development of additional assays is necessary to accurately detect sources of swine fecal pollution.

Antibiotics are used in animal livestock production for therapeutic treatment of disease and at subtherapeutic levels for growth promotion and improvement of feed efficiency. It is estimated that approximately 75% of antibiotics are not absorbed by animals and are excreted in waste. Antibiotic resistance selection occurs among gastrointestinal bacteria, which are also excreted in manure and stored in waste holding systems. Land application of animal waste is a common disposal method used in the United States and is a means for environmental entry of both antibiotics and genetic resistance determinants. Concerns for bacterial resistance gene selection and dissemination of resistance genes have prompted interest about the concentrations and biological activity of drug residues and break-down metabolites, and their fate and transport. Fecal bacteria can survive for weeks to months in the environment, depending on species and temperature, however, genetic elements can persist regardless of cell viability. Phylogenetic analyses indicate antibiotic resistance genes have evolved, although some genes have been maintained in bacteria before the modern antibiotic era. Quantitative measurements of drug residues and levels of resistance genes are needed, in addition to understanding the environmental mechanisms of genetic selection, gene acquisition, and the spatiotemporal dynamics of these resistance genes and their bacterial hosts. This review article discusses an accumulation of findings that address aspects of the fate, transport, and persistence of antibiotics and antibiotic resistance genes in natural environments, with emphasis on mechanisms pertaining to soil environments following land application of animal waste effluent.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms., Microorganisms are vital in mediating the earth's biogeochemical cycles; yet, despite our rapidly increasing ability to explore complex environmental microbial communities, the relationship between microbial community structure and ecosystem processes remains poorly understood. Here, we address a fundamental and unanswered question in microbial ecology: 'When do we need to understand microbial community structure to accurately predict function?' We present a statistical analysis investigating the value of environmental data and microbial community structure independently and in combination for explaining rates of carbon and nitrogen cycling processes within 82 global datasets. Environmental variables were the strongest predictors of process rates but left 44% of variation unexplained on average, suggesting the potential for microbial data to increase model accuracy. Although only 29% of our datasets were significantly improved by adding information on microbial community structure, we observed improvement in models of processes mediated by narrow phylogenetic guilds via functional gene data, and conversely, improvement in models of facultative microbial processes via community diversity metrics. Our results also suggest that microbial diversity can strengthen predictions of respiration rates beyond microbial biomass parameters, as 53% of models were improved by incorporating both sets of predictors compared to 35% by microbial biomass alone. Our analysis represents the first comprehensive analysis of research examining links between microbial community structure and ecosystem function. Taken together, our results indicate that a greater understanding of microbial communities informed by ecological principles may enhance our ability to predict ecosystem process rates relative to assessments based on environmental variables and microbial physiology.

Utilization of dissolved organic matter (DOM) is thought to be the purview of heterotrophic microorganisms, but photoautotrophs can take up dissolved organic nitrogen (DON) and dissolved organic carbon (DOC). This study investigated DOC and DON uptake in a laminated cyanobacterial mat community from hypersaline Salt Pond (San Salvador, Bahamas). The total community uptake of 3H-labeled substrates was measured in the light and in the dark and under conditions of high and low salinity. Salinity was the primary control of DOM uptake, with increased uptake occurring under low-salinity, ‘freshened’ conditions. DOC uptake was also enhanced in the light as compared with the dark and in samples incubated with the photosystem II inhibitor 3(3,4-dichlorophenyl)-1, 1-dimethylurea, suggesting a positive association between photosynthetic activity and DOC uptake. Microautoradiography revealed that some DOM uptake was attributed to cyanobacteria. Cyanobacteria DOM uptake was negatively correlated with that of smaller filamentous microorganisms, and DOM uptake by individual coccoid cells was negatively correlated with uptake by colonial coccoids. These patterns of activity suggest that Salt Pond microorganisms are engaged in resource partitioning, and DOM utilization may provide a metabolic boost to both heterotrophs and photoautrophs during periods of lowered salinity.

To monitor the dissemination of resistance genes into the environment, we determined the occurrence of tetracycline resistance (Tc r ) genes in groundwater underlying two swine confinement operations. Monitoring well networks (16 wells at site A and 6 wells at site C) were established around the lagoons at each facility. Groundwater ( n = 124) and lagoon ( n = 12) samples were collected from the two sites at six sampling times from 2000 through 2003. Total DNA was extracted, and PCR was used to detect seven Tc r genes [ tet (M), tet (O), tet (Q), tet (W), tet (C), tet (H), and tet (Z)]. The concentration of Tc r genes was quantified by real-time quantitative PCR. To confirm the Tc r gene source in groundwater, comparative analysis of tet (W) gene sequences was performed on groundwater and lagoon samples. All seven Tc r genes were continually detected in groundwater during the 3-year monitoring period at both sites. At site A, elevated detection frequency and concentration of Tc r genes were observed in the wells located down-gradient of the lagoon. Comparative analysis of tet (W) sequences revealed that the impacted groundwater contained gene sequences almost identical (99.8% identity) to those in the lagoon, but these genes were not found in background libraries. Novel sequence clusters and unique indigenous resistance gene pools were also found in the groundwater. Thus, antibiotic resistance genes in groundwater are affected by swine manure, but they are also part of the indigenous gene pool.

Trees that fall into lakes from riparian forest become habitat for aquatic organisms, and are potentially an important link between terrestrial and aquatic ecosystems. Coarse woody habitat (CWH) promotes the production of benthic invertebrate prey and offers refuge for prey fishes, which are in turn consumed by piscivorous fishes. We used a simulation model to explore responses of an aquatic food web to changes in CWH caused by lakeshore residential development and windstorm, as well as to harvest of adult piscivores. Residential development had a negative effect on fishes, and could lead to extirpation of benthivorous prey fish species. In contrast, pulsed addition of CWH following a windstorm had little effect on the aquatic food web. Our results suggest that CWH is more important as shelter for prey fishes than as a substrate for benthic invertebrate production. However, piscivore harvest can supersede the role of CWH as prey shelter, leading to piscivore collapse and prey persistence even when CWH levels are low enough to promote piscivore dominance. Thus, the effects of lakeshore residential development on fishes can be masked by angler harvest of top piscivores.

Disturbance and recovery influence microbial community structure and ecosystem functions in most natural environments. This study from a hypersaline Bahamian lagoon details the response of a benthic cyanobacterial mat to disturbance by Hurricane Frances, a category-4 storm. Clone libraries of cyanobacterial small subunit r-RNA genes and nitrogenase genes revealed significant shifts in cyanobacterial and diazotroph community composition following the hurricane. Post-hurricane clone libraries were dominated by sequences that had been rare in pre-hurricane communities. In spite of this dominance shift, re-colonizing mat communities performed nitrogen fixation and photosynthesis at rates within the normal range of variation measured in the mat at similar salinities. There was a tendency for nitrogen fixation rates from mats re-colonizing sites with hurricane-related sand deposition to be higher than those from mats re-colonizing sites without significant sand deposition. This suggests that the altered communities responded to a carbon : nitrogen imbalance that was particularly pronounced in areas subjected to disturbance by sand burial. The post-hurricane dominance of organisms that had been previously rare suggests that pre-hurricane diversity and functional redundancy contributed to the rapid recovery of ecosystem function in the post-disturbance environment.

There is increasing global demand for food, bioenergy feedstocks and a wide variety of bio-based products. In response, agriculture has advanced production, but is increasingly depleting soil regulating and supporting ecosystem services. New production systems have emerged, such as no-tillage, that can enhance soil services but may limit yields. Moving forward, agricultural systems must reduce trade-offs between production and soil services. Soil functional zone management (SFZM) is a novel strategy for developing sustainable production systems that attempts to integrate the benefits of conventional, intensive agriculture and no-tillage. SFZM creates distinct functional zones within crop row and inter-row spaces. By incorporating decimetre-scale spatial and temporal heterogeneity, SFZM attempts to foster greater soil biodiversity and integrate complementary soil processes at the sub-field level. Such integration maximizes soil services by creating zones of ‘active turnover’, optimized for crop growth and yield (provisioning services); and adjacent zones of ‘soil building’, that promote soil structure development, carbon storage and moisture regulation (regulating and supporting services). These zones allow SFZM to secure existing agricultural productivity while avoiding or minimizing trade-offs with soil ecosystem services. Moreover, the specific properties of SFZM may enable sustainable increases in provisioning services via temporal intensification (expanding the portion of the year during which harvestable crops are grown). We present a conceptual model of ‘virtuous cycles’, illustrating how increases in crop yields within SFZM systems could create self-reinforcing feedback processes with desirable effects, including mitigation of trade-offs between yield maximization and soil ecosystem services. Through the creation of functionally distinct but interacting zones, SFZM may provide a vehicle for optimizing the delivery of multiple goods and services in agricultural systems, allowing sustainable temporal intensification while protecting and enhancing soil functioning.

Examination of variation in ecological communities can lead to an understanding of the forces that structure communities, the consequences of change at the ecosystem level, and the relevant scales involved. This study details spatial and seasonal variability in the composition of nitrogen-fixing and cyanobacterial (i.e., oxygenic photosynthetic) functional groups of a benthic, hypersaline microbial mat from Salt Pond, San Salvador Island, Bahamas. This system shows extreme annual variability in the salinity of the overlying water and the extent of water coverage. Analysis of molecular variance and F ST tests of genetic differentiation of nifH and cyanobacterial 16S rRNA gene clone libraries allowed for changes at multiple taxonomic levels (i.e., above, below, and at the species level) to inform the conclusions regarding these functional groups. Composition of the nitrogen-fixing community showed significant seasonal changes related to salinity, while cyanobacterial composition showed no consistent seasonal pattern. Both functional groups exhibited significant spatial variation, changing with depth in the mat and horizontally with distance from the shoreline. The patterns of change suggest that cyanobacterial composition was more insensitive to water stress, and consequently, cyanobacteria dominated the nitrogen-fixing community during dry months but gave way to a more diverse community of diazotrophs in wet months. This seasonal pattern may allow the mat community to respond quickly to water-freshening events after prolonged dry conditions (system recovery) and maintain ecosystem function in the face of disturbance during the wet season (system resilience).

This study used a genetic fingerprinting technique (automated ribosomal intergenic spacer analysis [ARISA]) to characterize microbial communities from a culture-independent perspective and to identify those environmental factors that influence the diversity of bacterial assemblages in Wisconsin lakes. The relationships between bacterial community composition and 11 environmental variables for a suite of 30 lakes from northern and southern Wisconsin were explored by canonical correspondence analysis (CCA). In addition, the study assessed the influences of ARISA fragment detection threshold (sensitivity) and the quantitative, semiquantitative, and binary (presence-absence) use of ARISA data. It was determined that the sensitivity of ARISA was influential only when presence-absence-transformed data were used. The outcomes of analyses depended somewhat on the data transformation applied to ARISA data, but there were some features common to all of the CCA models. These commonalities indicated that differences in bacterial communities were best explained by regional (i.e., northern versus southern Wisconsin lakes) and landscape level (i.e., seepage lakes versus drainage lakes) factors. ARISA profiles from May samples were consistently different from those collected in other months. In addition, communities varied along gradients of pH and water clarity (Secchi depth) both within and among regions. The results demonstrate that environmental, temporal, regional, and landscape level features interact to determine the makeup of bacterial assemblages in northern temperate lakes.

Species diversity and richness, and seasonal population dynamics of phytoplankton, planktonic protozoa, and bacterioplankton sampled from the epilimnion of Crystal Bog in 2000, were examined in order to test the hypothesis that these groups' diversity and abundance patterns might be linked. Crystal Bog, a humic lake in Vilas County, Wisconsin, is part of the North Temperate Lakes Long-Term Ecological Research Site. Phytoplankton and planktonic protozoa were identified and enumerated in a settling chamber with an inverted microscope. Bacterial cells were enumerated with the use of fluorescence 4', 6'-diamidino-2-phenylindole (DAPI)-staining procedures, and automated ribosomal intergenic spacer analysis (ARISA) was used to assess bacterioplankton diversity. Bacterial cell counts showed little seasonal variation and averaged 2.6 x 10(6) cells/mL over the ice-free season. Phytoplankton and planktonic protozoan numbers varied by up to two orders of magnitude and were most numerous in late spring and summer. Dinoflagellates largely dominated Crystal Bog throughout the ice-free period, specifically Peridiniopsis quadridens in the spring, Peridinium limbatum in summer, and Gymnodinium fuscum and P. quadridens in fall. Brief blooms of Cryptomonas, Dinobryon, and Synura occurred between periods of dinoflagellate domination. The dominant dinoflagellate, Peridinium limbatum, was calculated to have a growth rate of 0.065 day(-1) and a doubling time of 10.7 days. Heterotrophic nanoflagellates (HNFs) were a consistent component of the planktonic protozoa; seasonal patterns were determined for three genera of HNFs (Monosiga, Bicosoeca, and Desmarella moniliformis). Three genera of ciliates (Coleps, Strobilidium, and Strombidium) comprised the greater part of the planktonic protozoa in Crystal Bog. The number of species of planktonic protozoa was too low to calculate a diversity index. Shannon-Weaver diversity indices for phytoplankton and bacterioplankton in the epilimnion followed very similar seasonal patterns in this lake, supporting the hypothesis that in freshwaters, diversity patterns of these groups are linked.

Despite considerable attention in recent years, the composition and dynamics of lake bacterial communities over annual time scales are poorly understood. This study used automated ribosomal intergenic spacer analysis (ARISA) to explore the patterns of change in lake bacterial communities in three temperate lakes over 2 consecutive years. The study lakes included a humic lake, an oligotrophic lake, and a eutrophic lake, and the epilimnetic bacterial communities were sampled every 2 weeks. The patterns of change in bacterial communities indicated that seasonal forces were important in structuring the behavior of the bacterial communities in each lake. All three lakes had relatively stable community composition in spring and fall, but summer changes were dramatic. Summertime variability was often characterized by recurrent drops in bacterial diversity. Specific ARISA fragments derived from these lakes were not constant among lakes or from year to year, and those fragments that did recur in lakes in different years did not exhibit the same seasonal pattern of recurrence. Nonetheless, seasonal patterns observed in 2000 were fairly successful predictors of the rate of change in bacterial communities and in the degree of autocorrelation of bacterial communities in 2001. Thus, seasonal forces may be important structuring elements of these systems as a whole even if they are uncoupled from the dynamics of the individual system components.

Microbial communities in the rhizosphere make significant contributions to crop health and nutrient cycling. However, their ability to perform important biogeochemical processes remains uncharacterized. Here, we identified important functional genes that characterize the rhizosphere microbial community to understand metabolic capabilities in the maize rhizosphere using the GeoChip-based functional gene array method. Significant differences in functional gene structure were apparent between rhizosphere and bulk soil microbial communities. Approximately half of the detected gene families were significantly (p

This chapter provides a survey of established molecular approaches as well as emerging ones for studying microbial communities in gastrointestinal systems. Whole-genome typing methods are cost-effective alternatives for assessing genetic relatedness. These molecular techniques provide more specificity than phenotypic identification of culturable microbes and include pulsed-field gel electrophoresis (PFGE), rep-PCR, multilocus sequence typing (MLST), and octamer-based genome scanning (OGBS). The genotyping method used in recent years for Enterobacteriaceae has been a PCR-based technique using primers targeting repetitive sequences (rep-PCR), which also produces numerous DNA fragments that are separable by electrophoresis. MLST is another PCR-enabled genotyping method that has found widespread use for subtyping a variety of pathogens. The technique is an extension of multilocus enzyme electrophoresis and was first described in 1998 using Neisseria meningitidis as an example. OBGS, a powerful method enabled by genome sequence information, was developed as a result of determining the complete genome sequence of Escherichia coli. Researchers using molecular community fingerprinting methods can rapidly infer the genetic composition or community structure following the analysis of a region of amplified DNA. All community fingerprinting methods begin with extraction of total genomic DNA from the sample. Direct DNA extraction followed by PCR amplification and subsequent restriction digestion is the basis for (terminal restriction fragment length polymorphism (T-RFLP). The greatest challenge to microbial community analysis is linking the community structure at the phylogenetic level with its metabolic potential and subsequent functionality in the environment being studied.

Hill prairies are remnant grasslands perched on the bluffs of major river valleys, and because their steep slopes make them unsuitable for traditional row crop agriculture, they have some of the lowest levels of anthropogenic disturbance of any prairie ecosystems in the Midwestern USA. However, many decades of fire suppression have allowed for shrub encroachment from the surrounding forests. While shrub encroachment of grasslands can modify soil respiration rates and nutrient storage, it is not known whether shrubs also alter the community composition of soil microorganisms. We conducted transect sampling of nine different hill prairie remnants showing varying degrees of shrub encroachment, and we used DNA-based community profiling (automated ribosomal intergenic spacer analysis) to characterize the composition of bacterial and fungal communities in the open prairie habitat, the shrub-encroached border, and the surrounding forest. While both bacterial and fungal communities showed statistically significant variation across these habitats, their predominant patterns were different. Bacterial communities of forest soils were distinct from those of the open prairie and the shrub-encroached areas, while fungal communities of the open prairie were distinct from those of the forest and the shrub-encroached border. Shrub encroachment significantly altered the community composition of soil fungal communities. Furthermore, fungal communities of heavily encroached prairie remnants more closely resembled those of the surrounding forest than those of lightly encroached prairies. Thus, shrub encroachment can cause soil fungi to shift from a “grassland” community to a “woody” community, with potential consequences for soil processes and plant-microbe interactions.

Identification of microbes that actively utilize root exudates is essential to understand plant-microbe interactions. To identify active root exudate-utilizing microorganisms associated with switchgrass - a potential bioenergy crop - plants were labelled in situ with (13) CO2 , and 16S and 18S rRNA genes in the (13) C-labelled rhizosphere DNA were pyrosequenced. Multi-pulse labelling for 5 days produced detectable (13) C-DNA, which was well separated from unlabelled DNA. Methylibium from the order Burkholderiales were the most heavily labelled bacteria. Pythium, Auricularia and Galerina were the most heavily labelled eukaryotic microbes. We also identified a Glomus intraradices-like species; Glomus members are arbuscular mycorrhizal fungi that are able to colonize the switchgrass root. All of these heavily labelled microorganisms were also among the most abundant species in the rhizosphere. Species belonging to Methylibium and Pythium were the most heavily labelled and the most abundant bacteria and eukaryotes in the rhizosphere of switchgrass. Our results revealed that nearly all of the dominant rhizosphere bacterial and eukaryotic microbes were able to utilize root exudates. The enrichment of microbial species in the rhizosphere is selective and mostly due to root exudation, which functions as a nutrition source, promoting the growth of these microbes.

Biomass production for bioenergy may change soil microbes and influence ecosystem properties. To explore the impact of different bioenergy cropping systems on soil microorganisms, the compositions and quantities of soil microbial communities (16S rRNA gene) and N-cycling functional groups (nifH, bacterial amoA, archaeal amoA and nosZ genes) were assessed under maize, switchgrass and Miscanthus x giganteus at seven sites representing a climate gradient (precipitation and temperature) in Illinois, USA. Overall, the site-to-site variation in community composition surpassed the variation due to plant type, and microbial communities under each crop did not converge on a 'typical' species assemblage. Fewer than 5% of archaeal amoA, bacterial amoA, nifH and nosZ OTUs were significantly different among these crops, but the largest differences observed at each site were found between maize and the two perennial grasses. Quantitative PCR revealed that the abundance of the nifH gene was significantly higher in the perennial grasses than in maize, and we also found significantly higher total N in the perennial grass soils than in maize. Thus, we conclude that cultivation of these perennial grasses, instead of maize, as bioenergy feedstocks can improve soil ecosystem nitrogen sustainability by increasing the population size of N-fixing bacteria.

The spatial scale on which microbial communities respond to plant invasions may provide important clues as to the nature of potential invader-microbe interactions. Lespedeza cuneata (Dum. Cours.) G. Don is an invasive legume that may benefit from associations with mycorrhizal fungi; however, it has also been suggested that the plant is allelopathetic and may alter the soil chemistry of invaded sites through secondary metabolites in its root exudates or litter. Thus, L. cuneata invasion may interact with soil microorganisms on a variety of scales. We investigated L. cuneata-related changes to soil bacterial and fungal communities at two spatial scales using multiple sites from across its invaded N. American range. Using whole community DNA fingerprinting, we characterized microbial community variation at the scale of entire invaded sites and at the scale of individual plants. Based on permutational multivariate analysis of variance, soil bacterial communities in heavily invaded sites were significantly different from those of uninvaded sites, but bacteria did not show any evidence of responding at very local scales around individual plants. In contrast, soil fungi did not change significantly at the scale of entire sites, but there were significant differences between fungal communities of native versus exotic plants within particular sites. The differential scaling of bacterial and fungal responses indicates that L. cuneata interacts differently with soil bacteria and soil fungi, and these microorganisms may play very different roles in the invasion process of this plant.

Reverse complementary DNA sequences - sequences that are inadvertently given backwards with all purines and pyrimidines transposed - can affect sequence analysis detrimentally unless taken into account. We present an open-source, high-throughput software tool -v-revcomp (http://www.cmde.science.ubc.ca/mohn/software.html) - to detect and reorient reverse complementary entries of the small-subunit rRNA (16S) gene from sequencing datasets, particularly from environmental sources. The software supports sequence lengths ranging from full length down to the short reads that are characteristic of next-generation sequencing technologies. We evaluated the reliability of v-revcomp by screening all 406 781 16S sequences deposited in release 102 of the curated SILVA database and demonstrated that the tool has a detection accuracy of virtually 100%. We subsequently used v-revcomp to analyse 1 171 646 16S sequences deposited in the International Nucleotide Sequence Databases and found that about 1% of these user-submitted sequences were reverse complementary. In addition, a nontrivial proportion of the entries were otherwise anomalous, including reverse complementary chimeras, sequences associated with wrong taxa, nonribosomal genes, sequences of poor quality or otherwise erroneous sequences without a reasonable match to any other entry in the database. Thus, v-revcomp is highly efficient in detecting and reorienting reverse complementary 16S sequences of almost any length and can be used to detect various sequence anomalies.

The effect of parenteral penicillin treatment on the intestinal microbiota was determined by monitoring the phenotypic antimicrobial resistance among Escherichia coli in 19 calves (15 calves received treatment and four calves were healthy controls) and by examining changes in the fecal microbial community structure using molecular fingerprinting techniques in a subset of eight calves (five treated calves and three control calves). After five days of penicillin treatment an increased resistance to multiple unrelated antimicrobial agents, including non-β-lactams, was seen in E. coli from treated calves, and this was not seen in the controls. Automated ribosomal intergenic spacer analysis (ARISA) and terminal restriction fragment length polymorphism (TRFLP) revealed that penicillin treatment causes a significant variation in the microbial structure within an individual calf. The study shows that parenteral administration of penicillin has an impact on the composition of the fecal microbiota in calves, and on the antimicrobial resistance pattern of their fecal E. coli.

Microbial communities are often organized in surficial laminated matrices called mats, which provide both a substrate for attachment and chemical gradients. Within the mats are distinct microhabitats in which specific members of the community are able to optimize biogeochemical cycling and growth. These gradients also provide protection against external environmental factors, including extremely high irradiance (including UV), nutrient deprivation, and periodic desiccation, which potentially inhibit growth and reproduction. Hypersalinity (>45‰) represents a particular challenge to microbes, as this condition can adversely affect microbial community function and structure. Benthic microbial mats provide a protective habitat in systems experiencing large fluctuations in salinity, including hypersalinity, by enabling microbial taxa to produce and exchange osmotically active metabolites, including intracellular and extracellular polymers that can counter such fluctuations; these polymers can also form part of the mat matrix. This structure enables a taxonomically diverse, productive community to thrive under conditions that would otherwise be too hostile for life to flourish, as in the overlying water column. As such, microbial production, nutrient cycling, and growth are often optimized in mats in aquatic ecosystems experiencing large swings in salinity. Microbial communities in these mats are surprisingly diverse and exhibit a high degree of redundancy. They include a wide spectrum of metabolically flexible prokaryotes (Eubacteria—including Cyanobacteria, photosynthetic and heterotrophic bacteria, and Archaea) and eukaryotic microalgae and micrograzers. Together, these organisms carry out primary production, nutrient cycling, and trophic transfer in lakes, lagoons, and other shallow-water ecosystems under both fluctuating and extreme conditions. We examined a highly dynamic and diverse community in a benthic mat system of hypersaline Salt Pond, located on San Salvador Island, the Bahamas. This mat community exhibits remarkable diversity in the face of extreme variations in salinity, brought about by long periods of drought and desiccation (leading to salinities in excess of 200%) and periodic tropical storms and hurricanes that can reduce the salinity to less than 40%. The high microbial diversity serves to provide a rapid response to environmental variability, in that specific members of this community can assume key production and nutrient cycling functions depending on their salinity optima. This mat system illustrates the functional roles and significance of a high degree of taxonomic and metabolic diversity in response to either sort of long-term climatic change, which characterized past and present environmental conditions on Earth.

The effect of oral amoxicillin treatment on fecal microbiota of seven healthy adult dogs was determined with a focus on the prevalence of bacterial antibiotic resistance and changes in predominant bacterial populations. After 4-7 days of exposure to amoxicillin, fecal Escherichia coli expressed resistance to multiple antibiotics when compared with the pre-exposure situation. Two weeks postexposure, the susceptibility pattern had returned to pre-exposure levels in most dogs. A shift in bacterial populations was confirmed by molecular fingerprinting of fecal bacterial populations using denaturing gradient gel electrophoresis (PCR-DGGE) of the 16S V3 rRNA gene region. Much of the variation in DGGE profiles could be attributed to dog-specific factors. However, permutation tests indicated that amoxicillin exposure significantly affected the DGGE profiles after controlling for the dog effect (P=0.02), and pre-exposure samples were clearly separated from postexposure samples. Sequence analysis of DGGE bands and real-time PCR quantification indicated that amoxicillin exposure caused a shift in the intestinal ecological balance toward a Gram-negative microbiota including resistant species in the family Enterobacteriaceae.

RNA methylase genes are common antibiotic resistance determinants for multiple drugs of the macrolide, lincosamide, and streptogramin B (MLS(B)) families. We used molecular methods to investigate the diversity, distribution, and abundance of MLS(B) methylases in waste lagoons and groundwater wells at two swine farms with a history of tylosin (a macrolide antibiotic structurally related to erythromycin) and tetracycline usage. Phylogenetic analysis guided primer design for quantification of MLS(B) resistance genes found in tylosin-producing Streptomyces (tlr(B), tlr(D)) and commensal/pathogenic bacteria (erm(A), erm(B), erm(C), erm(F), erm(G), erm(Q)). The near absence of tlr genes at these sites suggested a lack of native antibiotic-producing organisms. The gene combination erm(ABCF) was found in all lagoon samples analyzed. These four genes were also detected with high frequency in wells previously found to be contaminated by lagoon leakage. A weak correlation was found between the distribution of erm genes and previously reported patterns of tetracycline resistance determinants, suggesting that dissemination of these genes into the environment is not necessarily linked. Considerations of gene origins in history (i.e., phylogeny) and gene distributions in the landscape provide a useful "molecular ecology" framework for studying environmental spread of antibiotic resistance.