Within-House Spatial Distribution of Fecal Indicator Bacteria in Poultry Litter.

Land application of poultry litter is often considered to be a major source of water pollutants in poultry-producing regions. However, reported levels of fecal indicator microorganisms in litter vary widely, with considerable variation possible within houses and across farms, depending on management practices. Therefore, a study was conducted to determine the levels and distribution of indicator microorganisms within 12 broiler farms representing three companies. Within each house, litter samples were collected from around the feed line, water line, north wall, cool pad end, middle, and fan end. Litter moisture content was significantly different within the houses, with the litter being driest around the feed line (19.8%) and wettest around the water line (40.7%). Mean levels of total coliforms, , enterococci, and were 3.7, 3.3, 6.4, and 4.0 log colony-forming units g dry litter, respectively. Levels of total coliforms, , and were positively correlated with litter moisture content, but enterococci levels were not. Consequently, levels of total coliforms, , and , as well as enterococci, were highest around the water line and lowest around the feed line. These results indicate that areas with higher litter water content are more likely to contain higher levels of most fecal indicator microorganisms. Approaches to reduce litter water content in these areas would not only benefit the microbial quality of litter for land application but would also likely improve in-house disease control.

Impacts of Heronries on Water Quality as Evaluated through Escherichia coli and Fecal Sterol Analyses.

The authors used fecal sterol analysis to determine the potential contribution of E. coli from heronries to waterbodies in east-central Texas. They analyzed E. coli and fecal sterol concentrations in samples from four heronries during the breeding seasons in 2011-2013. The highest E. coli concentrations were in water samples from the two largest heronries established directly over water. The main sterols in fecal samples were cholesterol and stigmasterol, and in water, cholesterol, coprostanol, and cholestanol. Total sterols ranged 979 to 5838 ng/g dry weight in fecal samples, and 13 to 600 ng/L in water samples. There was a positive correlation between E. coli and the sum of bird sterols in water exposed directly to fecal deposition, but not in water surrounding the heronries. The authors found a strong association between E. coli and stigmasterol, suggesting that the presence of stigmasterol in water could be used for predicting E. coli sources from heronries nesting close to waterbodies.

Microbiome analysis revealed distinct microbial communities occupying different sized nodules in field-grown peanut.

Legume nodulation is the powerhouse of biological nitrogen fixation (BNF) where host-specific rhizobia dominate the nodule microbiome. However, other rhizobial or non-rhizobial inhabitants can also colonize legume nodules, and it is unclear how these bacteria interact, compete, or combinedly function in the nodule microbiome. Under such context, to test this hypothesis, we conducted 16S-rRNA based nodule microbiome sequencing to characterize microbial communities in two distinct sized nodules from field-grown peanuts inoculated with a commercial inoculum. We found that microbial communities diverged drastically in the two types of peanut nodules (big and small). Core microbial analysis revealed that the big nodules were inhabited by Bradyrhizobium, which dominated composition (>99%) throughout the plant life cycle. Surprisingly, we observed that in addition to Bradyrhizobium, the small nodules harbored a diverse set of bacteria (~31%) that were not present in big nodules. Notably, these initially less dominant bacteria gradually dominated in small nodules during the later plant growth phases, which suggested that native microbial communities competed with the commercial inoculum in the small nodules only. Conversely, negligible or no competition was observed in the big nodules. Based on the prediction of KEGG pathway analysis for N and P cycling genes and the presence of diverse genera in the small nodules, we foresee great potential of future studies of these microbial communities which may be crucial for peanut growth and development and/or protecting host plants from various biotic and abiotic stresses.

Isolation and Characterization of Bacterial Endophytes from Small Nodules of Field-Grown Peanut.

It is evident that legume root nodules can accommodate rhizobial and non-rhizobial bacterial endophytes. Our recent nodule microbiome study in peanuts described that small nodules can harbor diverse bacterial endophytes. To understand their functional role, we isolated 87 indigenous endophytes from small nodules of field-grown peanut roots and characterized them at molecular, biochemical, and physiological levels. The amplified 16S rRNA genes and phylogenetic analysis of these isolates revealed a wide variety of microorganisms related to the genera Bacillus, Burkholderia, Enterobacter, Herbaspirillum, Mistsuaria, Pantoea, Pseudomonas, and Rhizobia. It was observed that 37% (100% identity) and 56% (>99% identity) of the isolates matched with the amplified sequence variants (ASVs) from our previous microbiome study. All of these isolates were tested for stress tolerance (high temperature, salinity, acidic pH) and phosphate (P) solubilization along with ammonia (NH3), indole-3-acetic acid (IAA), 1-aminocyclopropane-1-carboxylate deaminase (ACCD), and siderophore production. The majority (78%) of the isolates were found to be halotolerant, thermotolerant, and acidophilic, and a few of them showed a significant positive response to the production of IAA, NH3, siderophore, ACCD, and P-solubilization. To evaluate the plant growth promotion (PGP) activity, plant and nodulation assays were performed in the growth chamber conditions for the selected isolates from both the non-rhizobial and rhizobial groups. However, these isolates appeared to be non-nodulating in the tested conditions. Nonetheless, the isolates 2 (Pantoea), 17 (Burkholderia), 21 (Herbaspirillum), 33o (Pseudomonas), and 77 (Rhizobium sp.) showed significant PGP activity in terms of biomass production. Our findings indicate that these isolates have potential for future biotechnological applications through the development of biologicals for sustainable crop improvement.

Hurricane Harvey Impacts on Water Quality and Microbial Communities in Houston, TX Waterbodies.

Extreme weather events can temporarily alter the structure of coastal systems and generate floodwaters that are contaminated with fecal indicator bacteria (FIB); however, every coastal system is unique, so identification of trends and commonalities in these episodic events is challenging. To improve our understanding of the resilience of coastal systems to the disturbance of extreme weather events, we monitored water quality, FIB at three stations within Clear Lake, an estuary between Houston and Galveston, and three stations in bayous that feed into the estuary. Water samples were collected immediately before and after Hurricane Harvey (HH) and then throughout the fall of 2017. FIB levels were monitored by culturing E. coli and Enterococci. Microbial community structure was profiled by high throughput sequencing of PCR-amplified 16S rRNA gene fragments. Water quality and FIB data were also compared to historical data for these water body segments. Before HH, salinity within Clear Lake ranged from 9 to 11 practical salinity units (PSU). Immediately after the storm, salinity dropped to < 1 PSU and then gradually increased to historical levels over 2 months. Dissolved inorganic nutrient levels were also relatively low immediately after HH and returned, within a couple of months, to historical levels. FIB levels were elevated immediately after the storm; however, after 1 week, E. coli levels had decreased to what would be acceptable levels for freshwater. Enterococci levels collected several weeks after the storm were within the range of historical levels. Microbial community structure shifted from a system dominated by Cyanobacteria sp. before HH to a system dominated by Proteobacteria and Bacteroidetes immediately after. Several sequences observed only in floodwater showed similarity to sequences previously reported for samples collected following Hurricane Irene. These changes in beta diversity corresponded to salinity and nitrate/nitrite concentrations. Differential abundance analysis of metabolic pathways, predicted from 16S sequences, suggested that pathways associated with virulence and antibiotic resistance were elevated in floodwater. Overall, these results suggest that floodwater generated from these extreme events may have high levels of fecal contamination, antibiotic resistant bacteria and bacteria rarely observed in other systems.

Dynamics of microbial community composition and function during in situ bioremediation of a uranium-contaminated aquifer.

A pilot-scale system was established to examine the feasibility of in situ U(VI) immobilization at a highly contaminated aquifer (U.S. DOE Integrated Field Research Challenge site, Oak Ridge, TN). Ethanol was injected intermittently as an electron donor to stimulate microbial U(VI) reduction, and U(VI) concentrations fell to below the Environmental Protection Agency drinking water standard (0.03 mg liter(-1)). Microbial communities from three monitoring wells were examined during active U(VI) reduction and maintenance phases with GeoChip, a high-density, comprehensive functional gene array. The overall microbial community structure exhibited a considerable shift over the remediation phases examined. GeoChip-based analysis revealed that Fe(III)-reducing bacterial (FeRB), nitrate-reducing bacterial (NRB), and sulfate-reducing bacterial (SRB) functional populations reached their highest levels during the active U(VI) reduction phase (days 137 to 370), in which denitrification and Fe(III) and sulfate reduction occurred sequentially. A gradual decrease in these functional populations occurred when reduction reactions stabilized, suggesting that these functional populations could play an important role in both active U(VI) reduction and maintenance of the stability of reduced U(IV). These results suggest that addition of electron donors stimulated the microbial community to create biogeochemical conditions favorable to U(VI) reduction and prevent the reduced U(IV) from reoxidation and that functional FeRB, SRB, and NRB populations within this system played key roles in this process.

Water management impacts on arsenic speciation and iron-reducing bacteria in contrasting rice-rhizosphere compartments.

Rice cultivated on arsenic (As) contaminated-soils will accumulate variable grain-As concentrations, as impacted by varietal differences, soil variables, and crop management. A field-scale experiment was conducted to study the impact of intermittent and continuous flooding on As speciation and microbial populations in rice rhizosphere compartments of soils that were either historically amended with As pesticide or unamended with As. Rhizosphere-soil, root-plaque, pore-water and grain As were quantified and speciated, and microbial populations in rhizosphere soil and root-plaque were characterized. Total-As concentrations in rhizosphere and grain were significantly lower in intermittently flooded compared to the continuously flooded plots (86% lower in pore-water, 55% lower in root-plaque and 41% lower in grain samples). iAs(V), iAs(III), and DMAs(V) were the predominant As species detected in rhizosphere-soil and root-plaque, pore-water and grain samples, respectively. Relative proportions of Archaea and iron-reducing bacteria (FeRB) were higher in rhizosphere soil compared to root-plaque. In rhizosphere soil, the relative abundance of FeRB was lower in intermittently flooded compared to continuously flooded plots, but there were no differences between root-plaque samples. This study has demonstrated that reductions in dissolved As concentrations in the rhizosphere and subsequent decreases in grain-As concentration can be attained through water management.

Niche Differentiation of Arsenic-Transforming Microbial Groups in the Rice Rhizosphere Compartments as Impacted by Water Management and Soil-Arsenic Concentrations.

Arsenic (As) bioavailability in the rice rhizosphere is influenced by many microbial interactions, particularly by metal-transforming functional groups at the root-soil interface. This study was conducted to examine As-transforming microbes and As-speciation in the rice rhizosphere compartments, in response to two different water management practices (continuous and intermittently flooded), established on fields with high to low soil-As concentration. Microbial functional gene composition in the rhizosphere and root-plaque compartments were characterized using the GeoChip 4.0 microarray. Arsenic speciation and concentrations were analyzed in the rhizosphere soil, root-plaque, pore water, and grain samples. Results confirmed several As-biotransformation processes in the rice rhizosphere compartments, and distinct assemblage of As-reducing and methylating bacteria was observed between the root-plaque and rhizosphere. Results confirmed higher potential for microbial As-reduction and As-methylation in continuously flooded, long term As-contaminated fields, which accumulated highest concentrations of AsIII and methyl-As concentrations in pore water and rice grains. Water management treatment significantly altered As-speciation in the rhizosphere, and intermittent flooding reduced methyl-As and AsIII concentrations in the pore water, root-plaque and rice grain. Ordination and taxonomic analysis of detected gene-probes indicated that root-plaque and rhizosphere assembled significantly different microbial functional groups demonstrating niche separation. Taxonomic non-redundancy was evident, suggesting that As-reduction, -oxidation and -methylation processes were performed by different microbial functional groups. It was also evident that As transformation was coupled to different biogeochemical cycling processes (nutrient assimilation, carbon metabolism etc.) in the compartments and between treatments, revealing functional non-redundancy of rice-rhizosphere microbiome in response to local biogeochemical conditions and As contamination. This study provided novel insights on As-biotransformation processes and their implications on As-chemistry at the root-soil interface and their responses to water management, which could be applied for mitigating As-bioavailability and accumulation in rice grains.

Increased Antimicrobial and Multidrug Resistance Downstream of Wastewater Treatment Plants in an Urban Watershed.

Development and spread of antimicrobial resistance (AMR) and multidrug resistance (MDR) through propagation of antibiotic resistance genes (ARG) in various environments is a global emerging public health concern. The role of wastewater treatment plants (WWTPs) as hot spots for the dissemination of AMR and MDR has been widely pointed out by the scientific community. In this study, we collected surface water samples from sites upstream and downstream of two WWTP discharge points in an urban watershed in the Bryan-College Station (BCS), Texas area, over a period of nine months. E. coli isolates were tested for resistance to ampicillin, tetracycline, sulfamethoxazole, ciprofloxacin, cephalothin, cefoperazone, gentamycin, and imipenem using the Kirby-Bauer disc diffusion method. Antimicrobial resistant heterotrophic bacteria were cultured on R2A media amended with ampicillin, ciprofloxacin, tetracycline, and sulfamethoxazole for analyzing heterotrophic bacteria capable of growth on antibiotic-containing media. In addition, quantitative real-time polymerase chain reaction (qPCR) method was used to measure eight ARG - tetA, tetW, aacA, ampC, mecA, ermA, blaTEM, and intI1 in the surface water collected at each time point. Significant associations (p < 0.05) were observed between the locations of sampling sites relative to WWTP discharge points and the rate of E. coli isolate resistance to tetracycline, ampicillin, cefoperazone, ciprofloxacin, and sulfamethoxazole together with an increased rate of isolate MDR. The abundance of antibiotic-resistant heterotrophs was significantly greater (p < 0.05) downstream of WWTPs compared to upstream locations for all tested antibiotics. Consistent with the results from the culture-based methods, the concentrations of all ARG were substantially higher in the downstream sites compared to the upstream sites, particularly in the site immediately downstream of the WWTP effluent discharges (except mecA). In addition, the Class I integron (intI1) genes were detected in high amounts at all sites and all sampling points, and were about ∼20 times higher in the downstream sites (2.5 × 107 copies/100 mL surface water) compared to the upstream sites (1.2 × 106 copies/100 mL surface water). Results suggest that the treated WWTP effluent discharges into surface waters can potentially contribute to the occurrence and prevalence of AMR in urban watersheds. In addition to detecting increased ARG in the downstream sites by qPCR, findings from this study also report an increase in viable AMR (HPC) and MDR (E. coli) in these sites. This data will benefit establishment of improved environmental regulations and practices to help manage AMR/MDR and ARG discharges into the environment, and to develop mitigation strategies and effective treatment of wastewater.

Elevated Incidences of Antimicrobial Resistance and Multidrug Resistance in the Maumee River (Ohio, USA), a Major Tributary of Lake Erie.

Maumee River, the major tributary in the western basin of Lake Erie, serves as one of major sources of freshwater in the area, supplying potable, recreational, and industrial water. In this study we collected water samples from four sites in the Maumee River Bay between 2016-2017 and E. coli was isolated, enumerated, and analyzed for antimicrobial resistance (AMR) and multidrug resistance (MDR). Strikingly, 95% of the total isolates were found to be resistant to at least one antibiotic. A very high resistance to the drugs cephalothin (95.3%), ampicillin (38.3%), tetracycline (8.8%), gentamicin (8.2%), ciprofloxacin (4.2%), cefoperazone (4%), and sulfamethoxazole (1.5%) was observed within isolates from all four sampling sites. Percentages of AMR and MDR was consistently very high in the summer and fall months, whereas it was observed to be lowest in the winter. A remarkably high number of the isolates were detected to be MDR-95% resistant to ≥1 antibiotic, 43% resistant to ≥2 antibiotics, 15% resistant to ≥3 antibiotics, 4.9% resistant to ≥4 antibiotic and 1.2% resistant to ≥5 antibiotics. This data will serve in better understanding the environmental occurrence and dissemination of AMR/MDR in the area and assist in improving and establishing control measures.

Structure and dynamics of the microbial communities underlying the carboxylate platform for biofuel production.

The carboxylate platform utilizes a mixed microbial community to convert lignocellulosic biomass into chemicals and fuels. While much of the platform is well understood, little is known about its microbiology. Mesophilic (40 degrees C) and thermophilic (55 degrees C) fermentations employing a sorghum feedstock and marine sediment inoculum were profiled using 16S rRNA tag-pyrosequencing over the course of a 30-day incubation. The contrasting fermentation temperatures converted similar amounts of biomass, but the mesophilic community was significantly more productive, and the two temperatures differed significantly with respect to propionic and butyric acid production. Pyrotag sequencing revealed the presence of dynamic communities that responded rapidly to temperature and changed substantially over time. Both temperatures were dominated by bacteria resembling Clostridia, but they shared few taxa in common. The species-rich mesophilic community harbored a variety of Bacteroidetes, Actinobacteria, and gamma-Proteobacteria, whereas the thermophilic community was composed mainly of Clostridia and Bacilli. Despite differences in composition and productivity, similar patterns of functional class dynamics were observed. Over time, organisms resembling known cellulose degraders decreased in abundance, while organisms resembling known xylose degraders increased. Improved understanding of the carboxylate platform's microbiology will help refine platform performance and contribute to our growing knowledge regarding biomass conversion and biofuel production processes.

Whole genome sequence analysis reveals high genetic variation of newly isolated Acidithiobacillus ferrooxidans IO-2C.

Acidithiobacillus ferrooxidans, a chemolithoautotrophic bacterium, is well known for its mineral oxidizing properties. The current study combines experimental and whole genome sequencing approaches to investigate an iron oxidizing, extreme acidophilic bacterium, A. ferrooxidans isolate (IO-2C) from an acid seep area near Carlos, TX, USA. Strain IO-2C was capable of oxidizing iron i.e. iron sulphate and iron ammonium sulphate yielding shwertmannite and jarosite minerals. Further, the bacterium's genome was sequenced, assembled and annotated to study its general features, structure and functions. To determine genetic heterogeneity, it was compared with the genomes of other published A. ferrooxidans strains. Pan-genome analysis displayed low gene conservation and significant genetic diversity in A. ferrooxidans species comprising of 6926 protein coding sequences with 23.04% (1596) core genes, 46.13% (3195) unique and 30.82% (2135) accessory genes. Variant analysis showed >75,000 variants, 287 of them with a predicted high impact, in A. ferrooxidans IO-2C genome compared to the reference strain, resulting in abandonment of some important functional key genes. The genome contains numerous functional genes for iron and sulphur metabolism, nitrogen fixation, secondary metabolites, degradation of aromatic compounds, and multidrug and heavy metal resistance. This study demonstrated the bio-oxidation of iron by newly isolated A. ferrooxidans IO-2C under acidic conditions, which was further supported by genomic analysis. Genomic analysis of this strain provided valuable information about the complement of genes responsible for the utilization of iron and tolerance of other metals.

Fungal Community Structural and Microbial Functional Pattern Changes After Soil Amendments by Oilseed Meals of Jatropha curcas and Camelina sativa: A Microcosm Study.

The meals after oil extraction from many oilseed crops have nutrition and biofumigation potential for land application. Oilseed meal (SM) from the dedicated bioenergy crop Jatropha curcas were implicated to contain compounds that have antibacterial properties on some soil pathogens. However, little is known about its effect on non-targeted soil microbial community, especially on fungi. SM from Camelina sativa contains moderate level of glucosinolates (GLS) and was under studied. To investigate soil fungal community responses to jatropha and camelina SMs, we conducted a lab based microcosm study, amending soil with 1% SMs of jatropha, camelina, flax, and biomass of wheat straw. Fungal community abundance and structure were analyzed based on the ITS region using qPCR and tag-pyrosequencing. Microbial functional changes were examined by community level physiological profile (CLPP) using Biolog assay. Both SMs from jatropha and camelina showed biofumigant properties and inhibited fungal proliferation. Jatropha SM significantly altered soil fungal community structures with lower fungal biodiversity and higher Chaetomium composition. Camelina SM amended soil promoted Fusarium proliferation. CLPP indicated sequential hierarchy for C metabolism in the oilseed-amended microcosms was generally complex C > phosphate-associated C > carboxylic acids > carbohydrates > amines > amino acids. No significant difference in CLPP was detected due to the type of SM treatment. Our data indicate that both SMs of jatropha and camelina have biofumigant properties and can differentially impact soil microbial communities, and the changes were relatively persistent over time. Microbial functional patterns on the other side were not impacted by SM type. Our study revealed biofumigant and nutritional influence of SMs from dedicated biofuel plants on soil microbial community. This information will help properly using jatropha and camelina SMs for pathogen control while minimizing their negative impacts on non-target microorganisms. However, further studies in the field are demanded to investigate their influences in real practice.

Diesel degrading bacterial endophytes with plant growth promoting potential isolated from a petroleum storage facility.

Thirteen (13) endophytic bacterial strains were isolated from Echinochloa crus-galli (Cockspur grass) and Cynodon dactylon (Bermuda grass) growing in an oil-contaminated site at a petroleum storage and transportation facility. Of the 13 strains assessed for their potential to degrade monoaromatic compounds (phenol, toluene, and xylene) and diesel and for their plant growth promoting (PGP) ability (phosphate solubilization, siderophores and 1-aminocyclopropane-1-carboxylate (ACC) deaminase production), isolate J10 (identified as Pseudomonas sp. by 16S rRNA gene sequencing) was found to the best diesel biodegrader with the best PGP traits. The Monod model used for Pseudomonas sp. J10 growth kinetics on diesel fuel as the sole carbon source showed that the maximum specific bacterial growth rate was 0.0644 h- 1 and the half velocity constant (K s ) was estimated as 4570 mg L- 1. The overall growth yield coefficient and apparent growth yield were determined to be 0.271 g h- 1 and 0.127 g cells/g substrate, respectively. Pseudomonas sp. J10 removed 69% diesel in four days as determined by gas chromatographic (GC) analysis. These findings could assist in developing an endophyte assisted efficient diesel biodegradation system using Pseudomonas sp. J10 isolated from Echinochloa crus-galli.

Biodegradation of phenol and benzene by endophytic bacterial strains isolated from refinery wastewater-fed Cannabis sativa.

The presence of benzene and phenol in the environment can lead to serious health effects in humans and warrant development of efficient cleanup strategies. The aim of the present work was to assess the potential of indigenous endophytic bacterial strains to degrade benzene and phenol. Seven strains were successfully isolated from Cannabis sativa plants irrigated with oil refinery wastewater. Molecular characterization was performed by 16S rRNA gene sequencing. Phenol was biodegraded almost completely with Achromobacter sp. (AIEB-7), Pseudomonas sp. (AIEB-4), and Alcaligenes sp. (AIEB-6) at 250, 500, and 750 mg L-1; however, the degradation was only 81%, 72%, and 69%, respectively, when exposed to 1000 mg L-1. Bacillus sp. (AIEB-1), Enterobacter sp. (AIEB-3), and Acinetobacter sp. (AIEB-2) degraded benzene significantly at 250, 500, and 750 mg L-1. However, these strains showed 80%, 72%, and 68% benzene removal at 1000 mg L-1 exposure, respectively. Rates of degradation could be modeled with first-order kinetics with rate constant values of 1.86 × 10-2 for Pseudomonas sp. (AIEB-4) and 1.80 × 10-2 h-1 for Bacillus sp. (AIEB-1) and half-lives of 1.5 and 1.6 days, respectively. These results establish a foundation for further testing of the phytoremediation of hydrocarbon-contaminated soils in the presence of these endophytic bacteria.

Soil Bacterial Community Was Changed after Brassicaceous Seed Meal Application for Suppression of Fusarium Wilt on Pepper.

Application of Brassicaceous seed meal (BSM) is a promising biologically based disease-control practice but BSM could directly and indirectly also affect the non-target bacterial communities, including the beneficial populations. Understanding the bacterial response to BSM at the community level is of great significance for directing plant disease management through the manipulation of resident bacterial communities. Fusarium wilt is a devastating disease on pepper. However, little is known about the response of bacterial communities, especially the rhizosphere bacterial community, to BSM application to soil heavily infested with Fusarium wilt pathogen and cropped with peppers. In this study, a 25-day microcosm incubation of a natural Fusarium wilt pathogen-infested soil supplemented with three BSMs, i.e., Camelina sativa 'Crantz' (CAME), Brassica juncea 'Pacific Gold' (PG), and a mixture of PG and Sinapis alba cv. 'IdaGold' (IG) (PG+IG, 1:1 ratio), was performed. Then, a further 35-day pot experiment was established with pepper plants growing in the BSM treated soils. The changes in the bacterial community in the soil after 25 days of incubation and changes in the rhizosphere after an additional 35 days of pepper growth were investigated by 454 pyrosequencing technique. The results show that the application of PG and PG+IG reduced the disease index by 100% and 72.8%, respectively, after 35 days of pepper growth, while the application of CAME did not have an evident suppressive effect. All BSM treatments altered the bacterial community structure and decreased the bacterial richness and diversity after 25 days of incubation, although this effect was weakened after an additional 35 days of pepper growth. At the phylum/class and the genus levels, the changes in specific bacterial populations resulting from the PG and PG+IG treatments, especially the significant increase in Actinobacteria-affiliated Streptomyces and an unclassified genus and the significant decrease in Chloroflexi, were suspected to be one of the microbial mechanisms involved in PG-containing BSM-induced disease suppression. This study is helpful for our understanding of the mechanisms that lead to contrasting plant disease severity after the addition of different BSMs.

Mineralisation of atrazine, metolachlor and their respective metabolites in vegetated filter strip and cultivated soil.

In vegetated filter strips (VFS) the presence of perennial vegetation, rhizodeposition of labile organic substrates and the accumulation of an organic residue thatch layer may enhance microbial numbers and activity, thereby increasing the potential for mineralisation of herbicides and herbicide metabolites retained during run-off events. The objective of this laboratory experiment was to compare the mineralisation of atrazine and metolachlor with that of their respective metabolites in VFS and cultivated soil. With the exception of total bacteria, propagule density of the microbial groups, endogenous soil enzymes and microbial diversity were higher in the VFS soil. This correlated with increased mineralisation of metolachlor and its metabolites in the VFS soil and indicates potential for VFS to curtail the subsequent transport of these compounds. In contrast, the mineralisation of atrazine and the majority of its metabolites was substantially reduced in VFS soil relative to cultivated soil. Consequently, the potential for subsequent transport of atrazine and many of its metabolites may be greater in VFS soil than in cultivated soil if reduced mineralisation is not offset by increased sorption in the VFS.

Hill number as a bacterial diversity measure framework with high-throughput sequence data.

Bacterial diversity is an important parameter for measuring bacterial contributions to the global ecosystem. However, even the task of describing bacterial diversity is challenging due to biological and technological difficulties. One of the challenges in bacterial diversity estimation is the appropriate measure of rare taxa, but the uncertainty of the size of rare biosphere is yet to be experimentally determined. One approach is using the generalized diversity, Hill number (Na), to control the variability associated with rare taxa by differentially weighing them. Here, we investigated Hill number as a framework for microbial diversity measure using a taxa-accmulation curve (TAC) with soil bacterial community data from two distinct studies by 454 pyrosequencing. The reliable biodiversity estimation was obtained when an increase in Hill number arose as the coverage became stable in TACs for a ≥ 1. In silico analysis also indicated that a certain level of sampling depth was desirable for reliable biodiversity estimation. Thus, in order to attain bacterial diversity from second generation sequencing, Hill number can be a good diversity framework with given sequencing depth, that is, until technology is further advanced and able to overcome the under- and random-sampling issues of the current sequencing approaches.

The role of cell bioaugmentation and gene bioaugmentation in the remediation of co-contaminated soils.

Soils co-contaminated with metals and organics present special problems for remediation. Metal contamination can delay or inhibit microbial degradation of organic pollutants such that for effective in situ biodegradation, bioaugmentation is necessary. We monitored the degradation of 2,4-dichlorophenoxyacetic acid (2,4-D) or 3-chlorobenzoate (3-CB) in two different soils with and without cadmium (Cd) contamination. Additionally, we evaluated the ability of bioaugmentation to enhance organic degradation in these co-contaminated soils. Finally, we determined whether enhanced degradation was due to survival of the introduced organism (cell bioaugmentation) or plasmid transfer to indigenous microbial populations (gene bioaugmentation). In Brazito soil, dual inoculation with a Cd-resistant bacterium plus a known 2,4-D-degrading bacterium, Ralstonia eutropha JMP134, enhanced 2,4-D degradation. Escherichia coli D11, which lacks chromosomal genes necessary for complete 2,4-D mineralization, was used for gene bioaugmentation in Madera soil. Significant gene transfer of the plasmid to the indigenous populations was observed, and the rate of 2,4-D degradation was enhanced relative to that of controls. Cell bioaugmentation was further demonstrated when (Comamonas testosteroni was used to enhance biodegradation of 3-CB in Madera soil. In this case no transfer of plasmid pBRC60 to indigenous soil recipients was observed. For the Madera soil, nonbioaugmented samples ultimately showed complete 2,4-D degradation. In contrast, nonbioaugmented Brazito soils showed incomplete 2,4-D degradation. These studies are unique in showing that both cell bioaugmentation and gene bioaugmentation can be effective in enhancing organic degradation in co-contaminated soils. Ultimately, the bioaugmentation strategy may depend on the degree of contamination and the time frame available for remediation.

Real-time PCR quantification of a green fluorescent protein-labeled, genetically engineered Pseudomonas putida strain during 2-chlorobenzoate degradation in soil.

The potential for real-time PCR (RTm-PCR) detection of the genetically engineered strain Pseudomonas putida GN2 was studied during 2-chlorobenzoate (2-CB) degradation in three different soils. The strain contained the constructed plasmid pGN2 which encoded genes for 2-CB oxidation (cbdA) and the green fluorescent protein (gfp). P. putida GN2 numbers were assessed by plating onto 2-CB minimal media and also by RTm-PCR detection of cbdA and gfp. Addition of P. putida GN2 decreased the time required to degrade 2-CB in all tested soils by more than 7 days. The RTm-PCR estimations of P. putida GN2 numbers strongly correlated with those obtained from plate count methods during active 2-CB degradation. However, after 2-CB degradation in the soils had ceased, RTm-PCR estimations of cbdA and gfp genes were generally one order of magnitude lower than those from plate counts. These results indicate the potential for RTm-PCR to rapidly determine degrader numbers in soil following bioaugmentation but also the need to exercise caution when attempting to determine cell numbers of degraders from the RTm-PCR quantification of plasmid encoded genes after substrate is depleted.