The Effects of Phyllosphere Bacteria on Plant Physiology and Growth of Soybean Infected with Pseudomonas syringae.

Phyllosphere bacteria are an important determinant of plant growth and resistance to pathogens. However, the efficacy of phyllosphere bacteria in regulating infection of Pseudomonas syringae pv. glycinea (Psg) and its influence on soybean growth and physiology is unknown. In a greenhouse study, we assessed the influence of a phyllosphere bacterial consortium (BC) of 13 species isolated from field-grown soybean leaves on uninfected and deliberately Psg infected soybean plants. We measured Psg density on infected leaves with and without the application of the BC. The BC application resulted in a significant reduction in Psg cells. We also measured plant biomass, nodule mass and number, gas exchange, and leaf chlorophyll and nitrogen in four treatment groups: control plants, plants with a BC and no infection (BC), plants with BC and infected with Psg (BC + Psg), and plants infected with Psg alone. For all variables, plants infected with Psg alone showed significant reduction in measured variables compared to both BC treatments. Therefore, the bacterial consortium was effective in controlling the negative effects of Psg on growth and physiology. The BC treatment sometimes resulted in increases in measured variables such as plant biomass, nodule numbers, and leaf chlorophyll as compared to control and BC + Psg treatments. Overall, the positive influence of BC treatment on plant growth and physiology highlights its potential applications to increase crop yield and control bacterial pathogens.

Soybean Root Nodule and Rhizosphere Microbiome: Distribution of Rhizobial and Nonrhizobial Endophytes.

Soybean root nodules are known to contain a high diversity of both rhizobial endophytes and nonrhizobial endophytes (NREs). Nevertheless, the variation of these bacteria among different root nodules within single plants has not been reported. So far, it is unclear whether the selection of NREs among different root nodules within single plants is a random process or is strictly controlled by the host plant to favor a few specific NREs based on their beneficial influence on plant growth. As well, it is also unknown if the relative frequency of NREs within different root nodules is consistent or if it varies based on the location or size of a root nodule. We assessed the microbiomes of 193 individual soybean root nodules from nine plants using high-throughput DNA sequencing. Bradyrhizobium japonicum strains occurred in high abundance in all root nodules despite the presence of other soybean-compatible rhizobia, such as Ensifer, Mesorhizobium, and other species of Bradyrhizobium in soil. Nitrobacter and Tardiphaga were the two nonrhizobial genera that were uniformly detected within almost all root nodules, though they were in low abundance. DNA sequences related to other NREs that have frequently been reported, such as Bacillus, Pseudomonas, Flavobacterium, and Variovorax species, were detected in a few nodules. Unlike for Bradyrhizobium, the low abundance and inconsistent occurrence of previously reported NREs among different root nodules within single plants suggest that these microbes are not preferentially selected as endophytes by host plants and most likely play a limited part in plant growth as endophytes.IMPORTANCE Soybean (Glycine max L.) is a valuable food crop that also contributes significantly to soil nitrogen by developing a symbiotic association with nitrogen-fixing rhizobia. Bacterial endophytes (both rhizobial and nonrhizobial) are considered critical for the growth and resilience of the legume host. In the past, several studies have suggested that the selection of bacterial endophytes within root nodules can be influenced by factors such as soil pH, nutrient availability, host plant genotype, and bacterial diversity in soil. However, the influence of size or location of root nodules on the selection of bacterial endophytes within soybean roots is unknown. It is also unclear whether the selection of nonrhizobial endophytes within different root nodules of a single plant is a random process or is strictly regulated by the host. This information can be useful in identifying potential bacterial species for developing bioinoculants that can enhance plant growth and soil nitrogen.

Tailored Bioactive Compost from Agri-Waste Improves the Growth and Yield of Chili Pepper and Tomato.

An extensive use of chemical fertilizers has posed a serious impact on food and environmental quality and sustainability. As the organic and biofertilizers can satisfactorily fulfill the crop's nutritional requirement, the plants require less chemical fertilizer application; hence, the food is low in chemical residues and environment is less polluted. The agriculture crop residues, being a rich source of nutrients, can be used to feed the soil and crops after composting and is a practicable approach to sustainable waste management and organic agriculture instead of open-field burning of crop residues. This study demonstrates a feasible strategy to convert the wheat and rice plant residues into composted organic fertilizer and subsequent enrichment with plant-beneficial bacteria. The bioactive compost was then tested in a series of in vitro and in vivo experiments for validating its role in growing organic vegetables. The compost was enriched with a blend of micronutrients, such as zinc, magnesium, and iron, and a multi-trait bacterial consortium AAP (Azospirillum, Arthrobacter, and Pseudomonas spp.). The bacterial consortium AAP showed survival up to 180 days post-inoculation while maintaining their PGP traits. Field emission scanning electron microscopic analysis and fluorescence in situ hybridization (FISH) of bioactive compost further elaborated the morphology and confirmed the PGPR survival and distribution. Plant inoculation of this bioactive compost showed significant improvement in the growth and yield of chilies and tomato without any additional chemical fertilizer yielding a high value to cost ratio. An increase of ≈35% in chlorophyll contents, ≈25% in biomass, and ≈75% in yield was observed in chilies and tomatoes. The increase in N was 18.7 and 25%, while in P contents were 18.5 and 19% in chilies and tomatoes, respectively. The application of bioactive compost significantly stimulated the bacterial population as well as the phosphatase and dehydrogenase activities of soil. These results suggest that bioactive compost can serve as a source of bioorganic fertilizer to get maximum benefits regarding vegetable yield, soil quality, and fertilizer saving with the anticipated application for other food crops. It is a possible win-win situation for environmental sustainability and food security.

Diazotrophs Show Signs of Restoration in Amazon Rain Forest Soils with Ecosystem Rehabilitation.

Biological nitrogen fixation can be an important source of nitrogen in tropical forests that serve as a major CO2 sink. Extensive deforestation of the Amazon is known to influence microbial communities and the biogeochemical cycles they mediate. However, it is unknown how diazotrophs (nitrogen-fixing microorganisms) respond to deforestation and subsequent ecosystem conversion to agriculture, as well as whether they can recover in secondary forests that are established after agriculture is abandoned. To address these knowledge gaps, we combined a spatially explicit sampling approach with high-throughput sequencing of nifH genes. The main objectives were to assess the functional distance decay relationship of the diazotrophic bacterial community in a tropical forest ecosystem and to quantify the roles of various factors that drive the observed changes in the diazotrophic community structure. We observed an increase in local diazotrophic diversity (α-diversity) with a decrease in community turnover (ß-diversity), associated with a shift in diazotrophic community structure as a result of the forest-to-pasture conversion. Both diazotrophic community turnover and structure showed signs of recovery in secondary forests. Changes in the diazotrophic community were primarily driven by the change in land use rather than differences in geochemical characteristics or geographic distances. The diazotroph communities in secondary forests resembled those in primary forests, suggesting that at least partial recovery of diazotrophs is possible following agricultural abandonment.IMPORTANCE The Amazon region is a major tropical forest region that is being deforested at an alarming rate to create space for cattle ranching and agriculture. Diazotrophs (nitrogen-fixing microorganisms) play an important role in supplying soil N for plant growth in tropical forests. It is unknown how diazotrophs respond to deforestation and whether they can recover in secondary forests that establish after agriculture is abandoned. Using high-throughput sequencing of nifH genes, we characterized the response of diazotrophs' ß-diversity and identified major drivers of changes in diazotrophs from forest-to-pasture and pasture-to-secondary-forest conversions. Studying the impact of land use change on diazotrophs is important for a better understanding of the impact of deforestation on tropical forest ecosystem functioning, and our results on the potential recovery of diazotrophs in secondary forests imply the possible restoration of ecosystem functions in secondary forests.

Illumina sequencing of 16S rRNA tag shows disparity in rhizobial and non-rhizobial diversity associated with root nodules of mung bean (Vigna radiata L.) growing in different habitats in Pakistan.

In Rhizobium-legume symbiosis, the nodule is the most frequently studied compartment, where the endophytic/symbiotic microbiota demands critical investigation for development of specific inocula. We identified the bacterial diversity within root nodules of mung bean from different growing areas of Pakistan using Illumina sequencing of 16S rRNA gene. We observed specific OTUs related to specific site where Bradyrhizobium was found to be the dominant genus comprising of 82-94% of total rhizobia in nodules with very minor fraction of sequences from other rhizobia at three sites. In contrast, Ensifer (Sinorhizobium) was single dominant genus comprising 99.9% of total rhizobial sequences at site four. Among non-rhizobial sequences, the genus Acinetobacter was abundant (7-18% of total sequences), particularly in Bradyrhizobium-dominated nodule samples. Rhizobia and non-rhizobial PGPR isolated from nodule samples include Ensifer, Bradyrhizobium, Acinetobacter, Microbacterium and Pseudomonas strains. Co-inoculation of multi-trait PGPR Acinetobacter sp. VrB1 with either of the two rhizobia in field exhibited more positive effect on nodulation and plant growth than single-strain inoculation which favors the use of Acinetobacter as an essential component for development of mung bean inoculum. Furthermore, site-specific dominance of rhizobia and non-rhizobia revealed in this study may contribute towards decision making for development and application of specific inocula in different habitats.

Impact of soil salinity on the microbial structure of halophyte rhizosphere microbiome.

The rhizosphere microbiome plays a significant role in the life of plants in promoting plant survival under adverse conditions. However, limited information is available about microbial diversity in saline environments. In the current study, we compared the composition of the rhizosphere microbiomes of the halophytes Urochloa, Kochia, Salsola, and Atriplex living in moderate and high salinity environments (Khewra salt mines; Pakistan) with that of the non-halophyte Triticum. Soil microbiomes analysis using pyrosequencing of 16S rRNA gene indicated that Actinobacteria were dominant in saline soil samples whereas Proteobacteria predominated in non-saline soil samples. Firmicutes, Acidobacteria, Bacteriodetes and Thaumarchaeota were predominant phyla in saline and non-saline soils, whereas Cyanobacteria, Verrucomicrobia, Gemmatimonadetes and the unclassified WPS-2 were less abundant. Sequences from Euryarchaeota, Ignavibacteriae, and Nanohaloarchaeota were identified only from the rhizosphere of halophytes. Dominant halophilic bacteria and archaea identified in this study included Agrococcus, Armatimonadetes gp4, Halalkalicoccus, Haloferula and Halobacterium. Our analysis showed that increases in soil salinity correlated with significant differences in the alpha and beta diversity of the microbial communities across saline and non-saline soil samples. Having a complete inventory of the soil bacteria from different saline environments in Pakistan will help in the discovery of potential inoculants for crops growing on salt-affected land.

Diversity of Bacillus-like bacterial community in the rhizospheric and non-rhizospheric soil of halophytes (Salsola stocksii and Atriplex amnicola), and characterization of osmoregulatory genes in halophilic Bacilli.

Salinity is one of the major abiotic stresses; a total of 3% of the world's land mass is affected by salinity. Approximately 6.3 million hectares of land in Pakistan is affected by salinity to varying degrees, and most of the areas are arid to semiarid with low annual precipitation. The aim of the present study is to identify and characterize Bacillus and Bacillus-derived bacterial genera from the rhizospheric and non-rhizospheric soil samples from the Khewra Salt Mine, Pakistan, by using culture-independent and -dependent methods. Seven Bacillus-like bacterial genera, Bacillus, Halobacillus, Virgibacillus, Brevibacillus, Paenibacillus, Tumebacillus, and Lysinibacillus, were detected by using pyrosequencing analysis, whereas only four genera, Bacillus, Halobacillus, Oceanobacillus, and Virgibacillus, were identified by culture-dependent methods. Most of the Bacillus-like isolates identified in this study were moderately halophilic, alkaliphilic, and mesophilic bacteria and were considered a good source of hydrolytic enzymes because of their ability to degrade proteins, carbohydrates, and lipids. Eight Bacillus-like strains from the genera Bacillus, Halobacillus, Oceanobacillus, and Virgibacillus showed positive results for the presence of ectABC gene cluster (ectoine), six strains could synthesize betaine from choline, and six strains tested positive for the synthesis of proline from either glutamate or ornithine by using proline dehydrogenase enzyme.

Iron limitation effects on nitrogen-fixing organisms with possible implications for cyanobacterial blooms.

Cyanobacteria-dominated harmful algal blooms are increasing in occurrence. Many of the taxa contributing to these blooms are capable of fixing atmospheric nitrogen and should be favored under conditions of low nitrogen availability. Yet, synthesizing nitrogenase, the enzyme responsible for nitrogen fixation, is energetically expensive and requires substantial concentrations of iron. Phosphorus addition to nitrogen poor streams should promote nitrogen fixation, but experimental results so far have been inconclusive, suggesting that other factors may be involved in controlling this process. With iron potentially limited in many streams, we examined the influence of phosphorus-iron colimitation on the community structure of nitrogen-fixing organisms. In stream microcosms, using microscopic and molecular sequence data, we observed: (i) the greatest abundance of heterocyst forming nitrogen-fixing cyanobacteria in low nitrogen treatments with high phosphorus and iron and (ii) greater abundance of non-photosynthetic nitrogen-fixing bacteria in treatments with nitrogen compared to those without it. We also found that comparisons between molecular results and those obtained from microscopic identification provided complementary information about cyanobacterial communities. Our investigation indicates the potential for phosphorus-iron colimitation of stream nitrogen-fixing organisms.

Retrieved 16S rRNA and nifH sequences reveal co-dominance of Bradyrhizobium and Ensifer (Sinorhizobium) strains in field-collected root nodules of the promiscuous host Vigna radiata (L.) R. Wilczek.

In the present study, the relative distribution of endophytic rhizobia in field-collected root nodules of the promiscuous host mung bean was investigated by sequencing of 16S ribosomal RNA (rRNA) and nifH genes, amplified directly from the nodule DNA. Co-dominance of the genera Bradyrhizobium and Ensifer was indicated by 32.05 and 35.84% of the total retrieved 16S rRNA sequences, respectively, and the sequences of genera Mesorhizobium and Rhizobium comprised only 0.06 and 2.06% of the recovered sequences, respectively. Sequences amplified from rhizosphere soil DNA indicated that only a minor fraction originated from Bradyrhizobium and Ensifer strains, comprising about 0.46 and 0.67% of the total retrieved sequences, respectively. 16S rRNA gene sequencing has also identified the presence of several non-rhizobial endophytes from phyla Proteobacteria, Actinobacteria, Bacteroides, and Firmicutes. The nifH sequences obtained from nodules also confirmed the co-dominance of Bradyrhizobium (39.21%) and Ensifer (59.23%) strains. The nifH sequences of the genus Rhizobium were absent, and those of genus Mesorhizobium comprised only a minor fraction of the sequences recovered from the nodules and rhizosphere soil samples. Two bacterial isolates, identified by 16S rRNA gene sequence analysis as Bradyrhizobium strain Vr51 and Ensifer strain Vr38, successfully nodulated the original host (mung bean) plants. Co-dominance of Bradyrhizobium and Ensifer strains in the nodules of mung bean indicates the potential role of the host plant in selecting specific endophytic rhizobial populations. Furthermore, successful nodulation of mung bean by the isolates showed that strains of both the genera Bradyrhizobium and Ensifer can be used for production of inoculum.

Dehalococcoides and general bacterial ecology of differentially trichloroethene dechlorinating flow-through columns.

The diversity of Dehalococcoides mccartyi (Dhc) and/or other organohalide respiring or associated microorganisms in parallel, partial, or complete trichloroethene (TCE) dehalogenating systems has not been well described. The composition of Dhc populations and the associated bacterial community that developed over 7.5 years in the top layer (0-10 cm) of eight TCE-fed columns were examined using pyrosequencing. Columns biostimulated with one of three carbon sources, along with non-stimulated controls, developed into complete (ethene production, whey amended), partial (cis-dichloroethene (DCE) and VC, an emulsified oil with nonionic surfactant), limited (<5 % cis-DCE and 95 % TCE, an emulsified oil), and non- (controls) TCE dehalogenating systems. Bioaugmentation of one column of each treatment with Bachman Road enrichment culture did not change Dhc populations nor the eventual degree of TCE dehalogenation. Pyrosequencing revealed high diversity among Dhc strains. There were 13 OTUs that were represented by more than 1000 sequences each. Cornell group-related populations dominated in complete TCE dehalogenating columns, while Pinellas group related Dhc dominated in all other treatments. General microbial communities varied with biostimulation, and three distinct microbial communities were established: one each for whey, oils, and control treatments. Bacterial genera, including Dehalobacter, Desulfitobacterium, Sulfurospirillum, Desulfuromonas, and Geobacter, all capable of partial TCE dehalogenation, were abundant in the limited and partial TCE dehalogenating systems. Dhc strain diversity was wider than previously reported and their composition within the community varied significantly depending on the nature of the carbon source applied and/or changes in the Dhc associated partners that fostered different biogeochemical conditions across the columns.

Conversion of Amazon rainforest to agriculture alters community traits of methane-cycling organisms.

Land use change is one of the greatest environmental impacts worldwide, especially to tropical forests. The Amazon rainforest has been subject to particularly high rates of land use change, primarily to cattle pasture. A commonly observed response to cattle pasture establishment in the Amazon is the conversion of soil from a methane sink in rainforest, to a methane source in pasture. However, it is not known how the microorganisms that mediate methane flux are altered by land use change. Here, we use the deepest metagenomic sequencing of Amazonian soil to date to investigate differences in methane-cycling microorganisms and their traits across rainforest and cattle pasture soils. We found that methane-cycling microorganisms responded to land use change, with the strongest responses exhibited by methane-consuming, rather than methane-producing, microorganisms. These responses included a reduction in the relative abundance of methanotrophs and a significant decrease in the abundance of genes encoding particulate methane monooxygenase. We also observed compositional changes to methanotroph and methanogen communities as well as changes to methanotroph life history strategies. Our observations suggest that methane-cycling microorganisms are vulnerable to land use change, and this vulnerability may underlie the response of methane flux to land use change in Amazon soils.

New Arsenate Reductase Gene (arrA) PCR Primers for Diversity Assessment and Quantification in Environmental Samples.

The extent of arsenic contamination in drinking water and its potential threat to human health have resulted in considerable research interest in the microbial species responsible for arsenic reduction. The arsenate reductase gene (arrA), an important component of the microbial arsenate reduction system, has been widely used as a biomarker to study arsenate-reducing microorganisms. A new primer pair was designed and evaluated for quantitative PCR (qPCR) and high-throughput sequencing of the arrA gene, because currently available PCR primers are not suitable for these applications. The primers were evaluated in silico and empirically tested for amplification of arrA genes in clones and for amplification and high-throughput sequencing of arrA genes from soil and groundwater samples. In silico, this primer pair matched (≥90% DNA identity) 86% of arrA gene sequences from GenBank. Empirical evaluation showed successful amplification of arrA gene clones of diverse phylogenetic groups, as well as amplification and high-throughput sequencing of independent soil and groundwater samples without preenrichment, suggesting that these primers are highly specific and can amplify a broad diversity of arrA genes. The arrA gene diversity from soil and groundwater samples from the Cache Valley Basin (CVB) in Utah was greater than anticipated. We observed a significant correlation between arrA gene abundance, quantified through qPCR, and reduced arsenic (AsIII) concentrations in the groundwater samples. Furthermore, we demonstrated that these primers can be useful for studying the diversity of arsenate-reducing microbial communities and the ways in which their relative abundance in groundwater may be associated with different groundwater quality parameters. IMPORTANCE: Arsenic is a major drinking water contaminant that threatens the health of millions of people worldwide. The extent of arsenic contamination and its potential threat to human health have resulted in considerable interest in the study of microbial species responsible for the reduction of arsenic, i.e., the conversion of AsV to AsIII In this study, we developed a new primer pair to evaluate the diversity and abundance of arsenate-reducing microorganisms in soil and groundwater samples from the CVB in Utah. We observed significant arrA gene diversity in the CVB soil and groundwater samples, and arrA gene abundance was significantly correlated with the reduced arsenic (AsIII) concentrations in the groundwater samples. We think that these primers are useful for studying the ecology of arsenate-reducing microorganisms in different environments.

Association of plant growth-promoting Serratia spp. with the root nodules of chickpea.

Serratia species-affiliated DNA sequences have recently been discovered in the root nodules of two chickpea cultivars; however, little is known about their potential influence on chickpea plant growth. All Serratia-affiliated sequences (1136) could be grouped into two clusters at 98% DNA similarity. The major cluster, represented by 96% of sequences, was closely associated with Serratia marcescens sequences from GenBank. In the current study, we isolated two Serratia strains, 5D and RTL100, from root nodules of a field-grown Desi cultivar from Faisalabad and Thal areas, respectively. In vitro, strain 5D showed significantly higher phosphate (P) solubilization and lactic acid production than RTL100, whereas a comparable concentration of phytohormone was produced by both isolates. The application of Serratia strain 5D as an inoculum resulted in 25.55% and 30.85% increases in the grain yield of crops grown on fertile soil in irrigated areas and nutrient-deficient soil in rainfed areas, respectively, compared to the non-inoculated control. Results of plant inoculations indicated that Serratia sp. 5D and RTL100 can serve as effective microbial inoculants, particularly in nutrient-deficient soils in rainfed areas, where chickpea is the only major crop grown during the entire year.

Dehalococcoides abundance and alternate electron acceptor effects on large, flow-through trichloroethene dechlorinating columns.

Trichloroethene (TCE) in groundwater is a major health concern and biostimulation/bioaugmentation-based strategies have been evaluated to achieve complete reductive dechlorination with varying success. Different carbon sources were hypothesized to stimulate different extents of TCE reductive dechlorination. Ecological conditions that developed different dechlorination stages were investigated by quantitating Dehalococcoides 16S rRNA (Dhc) and reductive dehalogenase gene abundance, and by describing biogeochemical properties of laboratory columns in response to this biostimulation. Eight large columns (183 cm × 15.2 cm), packed with aquifer material from Hill AFB, Utah, that were continuously fed TCE for 7.5 years. Duplicate columns were biostimulated with whey or one of two different Newman Zone® emulsified oil formulations containing either nonionic surfactant (EOLN) or standard surfactant (EOL). Two columns were non-stimulated controls. Complete (whey amended), partial (EOLN amended), limited (EOL), and non-TCE dehalogenating systems (controls) developed over the course of the study. Bioaugmentation of half of the columns with Bachman Road culture 3 years prior to dismantling did not influence the extent of TCE dehalogenation. Multivariate analysis clustered samples by biostimulation treatments and extent of TCE dehalogenation. Dhc, tceA, and bvcA gene concentrations did not show a consistent relationship with TCE dehalogenation but the vcrA gene was more abundant in completely dehalogenating, whey-treated columns. The whey columns developed strongly reducing conditions producing Fe(II), sulfide, and methane. Biostimulation with different carbon and energy sources can support high concentrations of diverse Dhc, but carbon addition has a major influence on biogeochemical processes effecting the extent of TCE dehalogenation.

Land use change alters functional gene diversity, composition and abundance in Amazon forest soil microbial communities.

Land use change in the Amazon rainforest alters the taxonomic structure of soil microbial communities, but whether it alters their functional gene composition is unknown. We used the highly parallel microarray technology GeoChip 4.0, which contains 83,992 probes specific for genes linked nutrient cycling and other processes, to evaluate how the diversity, abundance and similarity of the targeted genes responded to forest-to-pasture conversion. We also evaluated whether these parameters were reestablished with secondary forest growth. A spatially nested scheme was employed to sample a primary forest, two pastures (6 and 38 years old) and a secondary forest. Both pastures had significantly lower microbial functional genes richness and diversity when compared to the primary forest. Gene composition and turnover were also significantly modified with land use change. Edaphic traits associated with soil acidity, iron availability, soil texture and organic matter concentration were correlated with these gene changes. Although primary and secondary forests showed similar functional gene richness and diversity, there were differences in gene composition and turnover, suggesting that community recovery was not complete in the secondary forest. Gene association analysis revealed that response to ecosystem conversion varied significantly across functional gene groups, with genes linked to carbon and nitrogen cycling mostly altered. This study indicates that diversity and abundance of numerous environmentally important genes respond to forest-to-pasture conversion and hence have the potential to affect the related processes at an ecosystem scale.

Arsenic(V) reduction in relation to Iron(III) transformation and molecular characterization of the structural and functional microbial community in sediments of a basin-fill aquifer in Northern Utah.

Basin-fill aquifers of the Southwestern United States are associated with elevated concentrations of arsenic (As) in groundwater. Many private domestic wells in the Cache Valley Basin, UT, have As concentrations in excess of the U.S. EPA drinking water limit. Thirteen sediment cores were collected from the center of the valley at the depth of the shallow groundwater and were sectioned into layers based on redoxmorphic features. Three of the layers, two from redox transition zones and one from a depletion zone, were used to establish microcosms. Microcosms were treated with groundwater (GW) or groundwater plus glucose (GW+G) to investigate the extent of As reduction in relation to iron (Fe) transformation and characterize the microbial community structure and function by sequencing 16S rRNA and arsenate dissimilatory reductase (arrA) genes. Under the carbon-limited conditions of the GW treatment, As reduction was independent of Fe reduction, despite the abundance of sequences related to Geobacter and Shewanella, genera that include a variety of dissimilatory iron-reducing bacteria. The addition of glucose, an electron donor and carbon source, caused substantial shifts toward domination of the bacterial community by Clostridium-related organisms, and As reduction was correlated with Fe reduction for the sediments from the redox transition zone. The arrA gene sequencing from microcosms at day 54 of incubation showed the presence of 14 unique phylotypes, none of which were related to any previously described arrA gene sequence, suggesting a unique community of dissimilatory arsenate-respiring bacteria in the Cache Valley Basin.

Links between plant and fungal communities across a deforestation chronosequence in the Amazon rainforest.

Understanding the interactions among microbial communities, plant communities and soil properties following deforestation could provide insights into the long-term effects of land-use change on ecosystem functions, and may help identify approaches that promote the recovery of degraded sites. We combined high-throughput sequencing of fungal rDNA and molecular barcoding of plant roots to estimate fungal and plant community composition in soil sampled across a chronosequence of deforestation. We found significant effects of land-use change on fungal community composition, which was more closely correlated to plant community composition than to changes in soil properties or geographic distance, providing evidence for strong links between above- and below-ground communities in tropical forests.

Response of free-living nitrogen-fixing microorganisms to land use change in the Amazon rainforest.

The Amazon rainforest, the largest equatorial forest in the world, is being cleared for pasture and agricultural use at alarming rates. Tropical deforestation is known to cause alterations in microbial communities at taxonomic and phylogenetic levels, but it is unclear whether microbial functional groups are altered. We asked whether free-living nitrogen-fixing microorganisms (diazotrophs) respond to deforestation in the Amazon rainforest, using analysis of the marker gene nifH. Clone libraries were generated from soil samples collected from a primary forest, a 5-year-old pasture originally converted from primary forest, and a secondary forest established after pasture abandonment. Although diazotroph richness did not significantly change among the three plots, diazotroph community composition was altered with forest-to-pasture conversion, and phylogenetic similarity was higher among pasture communities than among those in forests. There was also 10-fold increase in nifH gene abundance following conversion from primary forest to pasture. Three environmental factors were associated with the observed changes: soil acidity, total N concentration, and C/N ratio. Our results suggest a partial restoration to initial levels of abundance and community structure of diazotrophs following pasture abandonment, with primary and secondary forests sharing similar communities. We postulate that the response of diazotrophs to land use change is a direct consequence of changes in plant communities, particularly the higher N demand of pasture plant communities for supporting aboveground plant growth.

Conversion of the Amazon rainforest to agriculture results in biotic homogenization of soil bacterial communities.

The Amazon rainforest is the Earth's largest reservoir of plant and animal diversity, and it has been subjected to especially high rates of land use change, primarily to cattle pasture. This conversion has had a strongly negative effect on biological diversity, reducing the number of plant and animal species and homogenizing communities. We report here that microbial biodiversity also responds strongly to conversion of the Amazon rainforest, but in a manner different from plants and animals. Local taxonomic and phylogenetic diversity of soil bacteria increases after conversion, but communities become more similar across space. This homogenization is driven by the loss of forest soil bacteria with restricted ranges (endemics) and results in a net loss of diversity. This study shows homogenization of microbial communities in response to human activities. Given that soil microbes represent the majority of biodiversity in terrestrial ecosystems and are intimately involved in ecosystem functions, we argue that microbial biodiversity loss should be taken into account when assessing the impact of land use change in tropical forests.

Development of a direct isolation procedure for free-living diazotrophs under controlled hypoxic conditions.

Free-living diazotrophs are diverse and ubiquitous in soil, contributing the nitrogen pool in natural ecosystems. The isolation of nitrogen-fixing microorganisms has relied on semisolid nitrogen-free medium enrichment, followed by multiple subculturing steps. These procedures limit the diversity of recovered isolates. In the current study, we investigated three different isolation strategies for free-living diazotrophs using a soil sample from the Amazon forest. The methods were (i) direct plating on solid nitrogen-free medium under a 2% O(2) concentration, (ii) enrichment in semisolid nitrogen-free medium before plating on solid nitrogen-free medium under 2% O(2), and (iii) enrichment followed by subculturing in the semisolid nitrogen-free medium before plating on nitrogen containing medium under a 21% O(2) concentration. A total of 794 isolates were differentiated by their genomic fingerprinting patterns, and strains with unique profiles were identified on the basis of sequencing of their 16S rRNA gene. Isolates belonged to four bacterial phyla: Proteobacteria, Firmicutes, Actinobacteria, and Bacteriodetes. The novel strategy of combining a solid N-free medium and hypoxic conditions showed an increase of 62.6% in the diversity of diazotrophs in comparison to that obtained by the conventional semisolid medium-based methods. All isolates grew on the nitrogen-free medium under a 2% O(2) concentration, 78% of them showed the presence of the nifH gene, and 39% tested positive for acetylene reduction activity. Our results suggest that direct plating of soil dilutions on nitrogen-free solid medium under a 2% O(2) concentration is a useful strategy for the isolation of the diverse diazotrophic communities.