Draft genome sequences of five Stenotrophomonas indicatrix strains isolated from soil.

Here, we present genome sequences of five Stenotrophomonas indicatrix strains, isolated from agricultural soil. Stenotrophomonas strains are commonly associated with the rhizosphere and are well-known for their ability to degrade xenobiotics. Yet, to date, knowledge about S. indicatrix is limited.

Limited impact of weathered residues from the Deepwater Horizon oil spill on the gut-microbiome and foraging behavior of sheepshead minnows (Cyprinodon variegatus).

The Deepwater Horizon disaster of April 2010 was the largest oil spill in U.S. history and exerted catastrophic effects on several ecologically important fish species in the Gulf of Mexico (GoM). Within fish, the microbiome plays a key symbiotic role in maintaining host health and aids in acquiring nutrients, supporting immune function, and modulating behavior. The aim of this study was to examine if exposure to weathered oil might produce significant shifts in fish gut-associated microbial communities as determined from taxa and genes known for hydrocarbon degradation, and whether foraging behavior was affected. The gut microbiome (16S rRNA and shotgun metagenomics) of sheepshead minnow (Cyprinodon variegatus) was characterized after fish were exposed to oil in High Energy Water Accommodated Fractions (HEWAF; tPAH = 81.1 ± 12.4 µg/L) for 7 days. A foraging behavioral assay was used to determine feeding efficiency before and after oil exposure. The fish gut microbiome was not significantly altered in alpha or beta diversity. None of the most abundant taxa produced any significant shifts as a result of oil exposure, with only rare taxa showing significant shifts in abundance between treatments. However, several bioindicator taxa known for hydrocarbon degradation were detected in the oil treatment, primarily Sphingomonas and Acinetobacter. Notably, the genus Stenotrophomonas was detected in high abundance in 16S data, which previously was not described as a core member of fish gut microbiomes. Data also demonstrated that behavior was not significantly affected by oil exposure. Potential low bioavailability of the oil may have been a factor in our observation of minor shifts in taxa and no behavioral effects. This study lays a foundation for understanding the microbiome of captive sheepshead minnows and indicates the need for further research to elucidate the responses of the fish gut-microbiome under oil spill conditions.

Sugarcane mill mud-induced putative host (soybean (Glycine max))-rhizobia symbiosis in sandy loam soil.

Domestic production of controlled-release, compost-based, and microbe-enhanced fertilizers is being expanded in the U.S. as a part of rural development. Sugarcane mill mud is a sterilized (≈90°C) agricultural byproduct in surplus that has received interests as a soil amendment in several Southern states, because of its high phosphorus and organic carbon contents. Addition of mill mud to sandy loam significantly increased the nodule formation compared to fertilized and unfertilized controls. Mill mud addition also resulted in pod yields similar to the fertilized control. Though not found in mill mud itself, mill mud additions correlated with an increase in soil Rhizobia as determined by deep 16S rRNA gene sequencing. We hypothesize that Firmicutes in sterilized mill mud induced Rhizobia that in turn enhanced soybean (Glycine max) growth. Collectively, mill mud enhanced the plant growth promoting bacteria when applied to a silt loam, although the relative influence of mill mud-derived bacteria, organic carbon, and nutrients is yet to be determined.

Metagenome-assembled genomes from sugarcane mill mud.

The genomes of 11 bacteria and 3 archaea were assembled from metagenomic DNA extracted from sugarcane mill mud. These metagenome-assembled genomes ranged from 1.79 to 6.45 Mb, with 2,263 to 5,551 predicted proteins, 80.65% to 100% genome completeness, and 43.19% to 68.02% G+C content.

Soil Properties Correlate with Microbial Community Structure in Qatari Arid Soils.

This is the first detailed characterization of the microbiota and chemistry of different arid habitats from the State of Qatar. Analysis of bacterial 16S rRNA gene sequences showed that in aggregate, the dominant microbial phyla were Actinobacteria (32.3%), Proteobacteria (24.8%), Firmicutes (20.7%), Bacteroidetes (6.3%), and Chloroflexi (3.6%), though individual soils varied widely in the relative abundances of these and other phyla. Alpha diversity measured using feature richness (operational taxonomic units [OTUs]), Shannon's entropy, and Faith's phylogenetic diversity (PD) varied significantly between habitats (P = 0.016, P = 0.016, and P = 0.015, respectively). Sand, clay, and silt were significantly correlated with microbial diversity. Highly significant negative correlations were also seen at the class level between both classes Actinobacteria and Thermoleophilia (phylum Actinobacteria) and total sodium (R = -0.82 and P = 0.001 and R = -0.86, P = 0.000, respectively) and slowly available sodium (R = -0.81 and P = 0.001 and R = -0.8 and P = 0.002, respectively). Additionally, class Actinobacteria also showed significant negative correlation with sodium/calcium ratio (R = -0.81 and P = 0.001). More work is needed to understand if there is a causal relationship between these soil chemical parameters and the relative abundances of these bacteria. IMPORTANCE Soil microbes perform a multitude of essential biological functions, including organic matter decomposition, nutrient cycling, and soil structure preservation. Qatar is one of the most hostile and fragile arid environments on earth and is expected to face a disproportionate impact of climate change in the coming years. Thus, it is critical to establish a baseline understanding of microbial community composition and to assess how soil edaphic factors correlate with microbial community composition in this region. Although some previous studies have quantified culturable microbes in specific Qatari habitats, this approach has serious limitations, as in environmental samples, approximately only 0.5% of cells are culturable. Hence, this method vastly underestimates natural diversity within these habitats. Our study is the first to systematically characterize the chemistry and total microbiota associated with different habitats present in the State of Qatar.

Chemical and microbial characterization of sugarcane mill mud for soil applications.

Sugarcane mill mud/filter cake is an activated sludge-like byproduct from the clarifier of a raw sugar production factory, where cane juice is heated to ≈90°C for 1-2 hr, after the removal of bagasse. Mill mud is enriched with organic carbon, nitrogen, and nutrient minerals; no prior report utilized 16S rRNA gene sequencing to characterize the microbial composition. Mill mud could be applied to agricultural fields as biofertilizer to replace or supplement chemical fertilizers, and as bio-stimulant to replenish microorganisms and organic carbon depleted by erosion and post-harvest field burning. However, mill mud has historically caused waste management challenges in the United States. This study reports on the chemical and microbial (16S rRNA) characteristics for mill muds of diverse origin and ages. Chemical signature (high phosphorus) distinguished mill mud from bagasse (high carbon to nitrogen (C/N) ratio) and soil (high pH) samples of diverse geographical/environmental origins. Bacterial alpha diversity of all sample types (mill mud, bagasse, and soil) was inversely correlated with C/N. Firmicutes dominated the microbial composition of fresh byproducts (mill mud and bagasse) as-produced within the operating factory. Upon aging and environmental exposure, the microbial community of the byproducts diversified to resemble that of soils, and became dominated by varying proportions of other phyla such as Acidobacteria, Chloroflexi, and Planctomyces. In summary, chemical properties allowed grouping of sample types (mill mud, bagasse, and soil-like), and microbial diversity analyses visualized aging caused by outdoor exposures including soil amendment and composting. Results suggest that a transient turnover of microbiome by amendments shifts towards more resilient population governed by the chemistry of bulk soil.

Effects of Environmental and Spatial Variables on Bacteria in Zhanjiang Mangrove Sediments.

The bottom mud of mangroves contains numerous microbial groups that play an important role in the main ecological functions of the mangrove ecosystem. The diversity and functional and environmental factors related to microbial communities, in terms of the assembly process and in environmental adaptation of the abundance and rare bacterial communities in the mangrove ecosystem, have not been fully explored. We used 16S high-throughput sequencing and operational taxonomic unit analysis to compare the diversity and composition of bacterial communities in different tidal zones in the sediments of the Zhanjiang Gaoqiao Mangrove Nature Reserve, compare the ecological adaptation thresholds and phylogenetic signals of bacterial communities under different environmental gradients, and examine the factors affecting the composition of the bacterial community. The diversity of microbial species and structure and function of the mangrove sediments were affected by the environment, showing the trend: mid tide zone > climax zone > low tide zone. Organic matter content, oxygen content, pH, and total phosphorus were identified as important environmental factors determining the functional diversity of bacterial communities and survival, while pH influences species evolution. The abundant taxa showed a wider response threshold and stronger phylogenetic signals of ecological preference across environmental gradients compared to rare taxa. The abundant bacterial groups have broader environmental adaptability than rare bacterial groups, and different environmental factors affect different communities and functions in the mangrove ecological environment. These results elucidate the mechanism underlying the generation and maintenance of bacterial diversity in response to global environmental changes.

Pumping supplies alter the microbiome of pumped human milk: An in-home, randomized, crossover trial.

BACKGROUND: The human milk microbiome may contribute to the benefits of breastfeeding by providing bacteria to the infant gastrointestinal tract. Many women pump their milk, but the effect of pumping on the milk microbiome is unknown. OBJECTIVES: Our objective was to determine the effects of pumping supplies on the pumped human milk microbiome. METHODS: This was an in-home, randomized, crossover trial of 2 collection methods. Women (n = 52) pumped twice within 3.5 h, once with their own breast pumps and milk collection supplies (OWN SUPP) and once with a hospital-grade pump and sterile collection supplies (STER SUPP). Pumping order was randomized. The milk microbiome was characterized by aerobic culturing and 16S ribosomal RNA gene sequencing. RESULTS: Milk collected with OWN SUPP yielded more total aerobic and gram-negative bacteria than milk collected with STER SUPP, reflecting a 6.6 (adjusted OR; 95% CI: 1.7, 25; P = 0.006) higher odds of containing >104 total aerobic CFU/mL and 19 (adjusted OR; 95% CI: 4.1, 88; P < 0.0001) higher odds of yielding culturable gram-negative bacteria. Milk collected with OWN SUPP yielded more Proteobacterias , including higher relative abundances of Acinetobacter and Stenotrophomonas, compared to milk collected with STER SUPP. Results were consistent across pumping-order groups. CONCLUSIONS: We demonstrated that pumping supplies altered the milk microbiome. On average, milk collected with OWN SUPP resulted in elevated levels of culturable total and gram-negative bacteria and proteobacterial DNA compared to milk collected with STER SUPP. More research is needed to assess implications for infant health.

Genome Sequences of Four Strains of Acinetobacter bereziniae Isolated from Human Milk Pumped with a Personal Breast Pump and Hand-Washed Milk Collection Supplies.

Acinetobacter bereziniae, formerly Acinetobacter genomospecies 10, is an opportunistic pathogen possessing resistance to multiple antibiotics, and it has been reported to be responsible for hospital-associated infections in immunocompromised individuals. We report the draft genome sequences of four Acinetobacter bereziniae strains that were isolated from a single human milk sample collected with a personal breast pump and a hand-washed milk collection kit.

Magnetic biochar derived from biosolids via hydrothermal carbonization: Enzyme immobilization, immobilized-enzyme kinetics, environmental toxicity.

Magnetic and nonmagnetic biochar (MBC & BC) were produced from biosolids under hydrothermal conditions and characterized in order to understand surface chemistry impacts on enzyme immobilization and activity. Peak surface pore size of MBC was 180 nm and that of BC was 17 nm. Despite similar surface area (≈ 49 m2/g) MBC immobilized more laccase (99 mg/g) than biochar (31 mg/g). For horseradish peroxidase (HRP), the two biochars had similar immobilization capacity (≈ 65 mg/g). Laccase and HRP on MBC had 47.1 and 18.0% higher specific activity than on BC, respectively. The matrix activity of MBC-laccase (33.3 U/mg support) was 3.7-fold higher than BC-laccase (8.8 U/mg support) and higher than the same amount of free laccase (30.2 U) at pH 3.0 (P < 0.05). Although MBC had its own peroxide oxidation activity (104.1 and 165.9 U/mg biochar at pHs 5&6) this only accounted for 16.7 and 20.4% of the total MBC-H RP activity respectively. After 10 wash cycles, MBC still retained 79.3% and 60.3% of laccase and HRP activity, respectively. Additionally, MBC had lower acute toxicity, suggesting that it is relative benign from an environmental perspective.

Fomesafen impacts bacterial communities and enzyme activities in the rhizosphere.

Fomesafen, a long-lived protoporphyrinogen-oxidase inhibitor, specially developed for post-emergence control of broad-leaf weeds, is used widely in soybean fields in northern China (Dayan and Duke, 2010). The impact of fomesafen on microbial communities in rhizosphere soils, however, is unknown. In this study we examined fomesafen degradation as well as its effects in the rhizosphere of soybean plants grown in a greenhouse. Fomesafen had shorter half-life in rhizosphere soil than previously reported for bulk soil from the same location (87 vs 120 days). The enzyme activity of soil extracts and the microbial community composition of 16S rRNA genes (16S) amplified from soil DNA were also investigated. Although not immediately apparent, both the high (37.5 mg kg-1) and low (18.75 mg kg-1) doses of fomesafen significantly decreased urease and invertase activities in the rhizosphere soil from days 30 and 45 respectively until the end of the experiment (90 days). Analysis of 16S amplicons demonstrated that fomesafen had a dose dependent effect, decreasing alpha diversity and altering beta diversity. Significant phylum level decreases were observed in five of the ten phyla that were most abundant in the control. Proteobacteria was the only phylum whose relative abundance increased in the presence of fomesafen, driven by increases in the genera Methylophilacaea, Dyella, and Sphingomonas. The functional implications of changes in 16S abundance as predicted using PICRUSt suggested that fomesafen enriched for enzymes involved in xenobiotic metabolism and detoxification (cytochrome P450s and glutathione metabolism). Our data suggest that, despite being degraded more rapidly in the rhizosphere than in bulk soil, fomesafen had long-lasting functional impacts on the soil microbial community.

Curli production enhances clay-E. coli aggregation and sedimentation.

Curli are amyloid fibrils that polymerize extracellularly from curlin, a protein that is secreted by many enteric bacteria and is important for biofilm formation. Presented here is a systematic study of the effects of curli on bacteria-clay interactions. The aggregation trends of curli-producing and curli-deficient bacteria with clay minerals were followed using gradient-sedimentation experiments, Lumisizer measurements, bright-field and electron microscopy. The results revealed that curli-producing bacteria auto-aggregated into high-density flocs (1.23 g/cm3), ranging in size from 10 to 50 µm, that settle spontaneously. In contrast, curli-deficient bacteria remained relatively stable in solution as individual cells (1-2 µm, 1.18 g/cm3), even at high ionic strength (350 mM). The stability of clay suspensions mixed with curli-deficient bacteria depended on clay type and ionic strength, the general trends being consistent with the classic DLVO theory. However, suspensions of curli-producing bacteria mixed with clays were highly unstable regardless of clay type and solution chemistry, suggesting extensive interactions between the clays and the bacteria-curli aggregates. SEM measurements revealed interesting differences in morphologies of the aggregates; montmorillonite particles coated the bacterial auto-aggregates whereas the kaolinite platelets were embedded within the larger curli-bacteria aggregates. These new observations regarding the densities, aggregation trends, and morphologies of bacteria-curli and bacteria-curli-clay complexes make it clear that production of surface appendages, such as curli, need to be considered when addressing the fate, activity and transport of bacteria - particularly in aquatic environments.

Absence of genetic selection in a pathogenic Escherichia coli strain exposed to the manure-amended soil environment.

Escherichia coli that express curli are more common in subsurface soil drainage when manure is surface applied. However, it is unknown whether this arises from mutations in individual strains leading to curli expression or by selection for individuals already expressing higher levels of curli. To test this, we examined curli production in pathogenic E. coli O157:H7 EDL933 as a function of manure management. Five treatments were investigated: (1) soil only, (2) soil with surface-applied E. coli O157:H7 EDL933 Δstx1-2 (EcO157), (3) soil with incorporated EcO157, (4) soil with surface-applied EcO157-inoculated manure, and (5) soil with incorporated EcO157-inoculated manure. EcO157 was reisolated from soils immediately after application and weekly thereafter for 8 weeks. EcO157 in the surface-applied treatments died faster than their incorporated treatment counterparts. Phenotypic assays revealed differences between treatments as well. Half of surface-applied manure reisolates from week 6 developed a mixed red and white colony morphology on Congo Red plates, indicating changes in curli production that were not seen in other treatments or times. In 37°C growth tests, week 6 reisolates from all treatments except soil surface-applied EcO157 left the lag phase at a significantly greater rate than week 0 isolates. We applied whole genome sequencing technology to interrogate the genetic underpinnings of these phenotypes. Surprisingly, we only found single-nucleotide polymorphisms in two of the 94 resequenced isolates from the different treatments, neither of which correlated with curli phenotype. Likewise, we found no differences in other genomic characteristics that might account for phenotypic differences including the count of gaps and the origin of discarded reads that failed to map to the parental strain. These results suggest there were no systematic genomic differences (i.e. individual-level selection) that correlated with time or treatment. We recommend future research focus on population-level selection of E. coli strains in the manure-amended soil environment.

Detection of glyphosate residues in companion animal feeds.

The widespread adoption of genetically modified, glyphosate-tolerant corn and soybean varieties in US crop production has led to a dramatic increase in glyphosate usage. Though present at or below regulatory limits currently set for human foodstuffs, the concentration of glyphosate in companion animal feed is currently unknown. In the present study, 18 commercial companion animal feeds from eight manufacturers were analyzed for glyphosate residues using ELISA. Every product contained detectable glyphosate residues in the range of 7.83 × 101-2.14 × 103 µg kg-1 dry weight, with the average and medians being 3.57 × 102 and 1.98 × 102 µg kg-1 respectively. Three products were tested for within-bag variation and six were tested for lot to lot variation. Little within-bag variation was found, but the concentration of glyphosate varied by lot in half of the products tested. Glyphosate concentration was significantly correlated with crude fiber content, but not crude fat or crude protein. Average daily intakes by animals consuming feeds containing the median glyphosate concentration are estimated to result in exposures that are 0.68-2.5% of the Allowable Daily Intake (ADI) for humans in the US and EU, which are 1750 and 500 µg kg-1 respectively. Consumption of the most contaminated feed, however, would result in exposure to 7.3% and 25% of the above ADIs, though the relevance of such an exposure to companion animals is currently unknown. Companion animal feeds contained 7.83 × 101-2.14 × 103 µg kg-1 glyphosate which is likely to result in pet exposure that is 4-12 times higher than that of humans on a per Kg basis.

Biochar does not attenuate triclosan's impact on soil bacterial communities.

Triclosan, a broad-spectrum antimicrobial, has been widely used in pharmaceutical and personal care products. It undergoes limited degradation during wastewater treatment and is present in biosolids, most of which are land applied in the United States. This study assessed the impact of triclosan (0-100 mg kg-1) with and without biochar on soil bacterial communities. Very little 14C-triclosan was mineralized to 14CO2 (<7%) over the course of the study (42 days). While biochar (1%) significantly lowered mineralization of triclosan, analysis of 16S rRNA gene sequences revealed that biochar impacted very few OTUs and did not alter the overall structure of the community. Triclosan, on the other hand, significantly affected bacterial diversity and community structure (alpha diversity, ANOVA, p < 0.001; beta diversity, AMOVA, p < 0.01). Dirichlet multinomial mixtures (DMM) modeling and complete linkage clustering (CLC) revealed a dose-dependent impact of triclosan. Non-Parametric Metastats (NPM) analysis showed that 150 of 734 OTUs from seven main phyla were significantly impacted by triclosan (adjusted p < 0.05). Genera harboring opportunistic pathogens such as Flavobacterium were enriched in the presence of triclosan, as was Stenotrophomonas. The latter has previously been implicated in triclosan degradation via stable isotope probing. Surprisingly, Sphingomonads, which include well-characterized triclosan degraders were negatively impacted by even low doses of triclosan. Analyses of published genomes showed that triclosan resistance determinants were rare in Sphingomonads which may explain why they were negatively impacted by triclosan in our soil.

A microbial community snapshot of windrows from a commercial composting facility.

The effect of depth on compost microbial communities is unclear but could be relevant to the management of windrows at commercial facilities. DNA extracted from 64 compost samples from seven windrows at a commercial facility were analyzed via deep 16S rRNA gene sequencing. The relative abundance of eight to nine genera was affected by depth during the transition from cooling to maturation phases between 4 and 6 months, whereas very few genera (0-1) showed a depth dependence in young, actively managed windrows or in mature windrows older than 10 months. Seven novel bacterial operational taxonomic units (OTUs) were detected in compost DNA and also in publicly available compost metagenomes. A compost metagenome was used to construct a metagenome-assembled genome for most of the abundant uncharacterized OTU in our samples and suggests its involvement in carbon cycling.

The NAG Sensor NagC Regulates LEE Gene Expression and Contributes to Gut Colonization by Escherichia coli O157:H7.

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 are human pathogens responsible for bloody diarrhea and renal failures. EHEC employ a type 3 secretion system to attach directly to the human colonic epithelium. This structure is encoded by the locus of enterocyte effacement (LEE) whose expression is regulated in response to specific nutrients. In this study, we show that the mucin-derived sugars N-acetylglucosamine (NAG) and N-acetylneuraminic acid (NANA) inhibit EHEC adhesion to epithelial cells through down-regulation of LEE expression. The effect of NAG and NANA is dependent on NagC, a transcriptional repressor of the NAG catabolism in E. coli. We show that NagC is an activator of the LEE1 operon and a critical regulator for the colonization of mice intestine by EHEC. Finally, we demonstrate that NAG and NANA as well as the metabolic activity of Bacteroides thetaiotaomicron affect the in vivo fitness of EHEC in a NagC-dependent manner. This study highlights the role of NagC in coordinating metabolism and LEE expression in EHEC and in promoting EHEC colonization in vivo.

Microbial Characterization of Qatari Barchan Sand Dunes.

This study represents the first characterization of sand microbiota in migrating barchan sand dunes. Bacterial communities were studied through direct counts and cultivation, as well as 16S rRNA gene and metagenomic sequence analysis to gain an understanding of microbial abundance, diversity, and potential metabolic capabilities. Direct on-grain cell counts gave an average of 5.3 ± 0.4 x 105 cells g-1 of sand. Cultured isolates (N = 64) selected for 16S rRNA gene sequencing belonged to the phyla Actinobacteria (58%), Firmicutes (27%) and Proteobacteria (15%). Deep-sequencing of 16S rRNA gene amplicons from 18 dunes demonstrated a high relative abundance of Proteobacteria, particularly enteric bacteria, and a dune-specific-pattern of bacterial community composition that correlated with dune size. Shotgun metagenome sequences of two representative dunes were analyzed and found to have similar relative bacterial abundance, though the relative abundances of eukaryotic, viral and enterobacterial sequences were greater in sand from the dune closer to a camel-pen. Functional analysis revealed patterns similar to those observed in desert soils; however, the increased relative abundance of genes encoding sporulation and dormancy are consistent with the dune microbiome being well-adapted to the exceptionally hyper-arid Qatari desert.

Draft Whole-Genome Sequence of Urease-Producing Sporosarcina koreensis.

Urease-producing microbes are of significance due to their potential application in biocement production. Sporosarcina koreensis Q1 is a urease-producing bacterium belonging to the phylum Firmicutes. Here, we present the draft whole-genome sequence of S. koreensis Q1, isolated from a barchan sand dune in Qatar.