Search

Spontaneous preterm birth (sPTB) is a leading cause of maternal and neonatal morbidity and mortality, yet its prevention and early risk stratification are limited. Previous investigations have suggested that vaginal microbes and metabolites may be implicated in sPTB. Here we performed untargeted metabolomics on 232 second-trimester vaginal samples, 80 from pregnancies ending preterm. We find multiple associations between vaginal metabolites and subsequent preterm birth, and propose that several of these metabolites, including diethanolamine and ethyl glucoside, are exogenous. We observe associations between the metabolome and microbiome profiles previously obtained using 16S ribosomal RNA amplicon sequencing, including correlations between bacteria considered suboptimal, such as Gardnerella vaginalis, and metabolites enriched in term pregnancies, such as tyramine. We investigate these associations using metabolic models. We use machine learning models to predict sPTB risk from metabolite levels, weeks to months before birth, with good accuracy (area under receiver operating characteristic curve of 0.78). These models, which we validate using two external cohorts, are more accurate than microbiome-based and maternal covariates-based models (area under receiver operating characteristic curve of 0.55–0.59). Our results demonstrate the potential of vaginal metabolites as early biomarkers of sPTB and highlight exogenous exposures as potential risk factors for prematurity.

Natural bacterial populations can display enormous genomic diversity, primarily in the form of gene content variation caused by the frequent exchange of DNA with the local environment. However, the ecological drivers of genomic variability and the role of selection remain controversial. Here, we address this gap by developing a nationwide atlas of 1,854 Listeria isolates, collected systematically from soils across the contiguous United States. We found that Listeria was present across a wide range of environmental parameters, being mainly controlled by soil moisture, molybdenum and salinity concentrations. Whole-genome data from 594 representative strains allowed us to decompose Listeria diversity into 12 phylogroups, each with large differences in habitat breadth and endemism. ‘Cosmopolitan’ phylogroups, prevalent across many different habitats, had more open pangenomes and displayed weaker linkage disequilibrium, reflecting higher rates of gene gain and loss, and allele exchange than phylogroups with narrow habitat ranges. Cosmopolitan phylogroups also had a large fraction of genes affected by positive selection. The effect of positive selection was more pronounced in the phylogroup-specific core genome, suggesting that lineage-specific core genes are important drivers of adaptation. These results indicate that genome flexibility and recombination are the consequence of selection to survive in variable environments. A population genomic analysis of 1,854 Listeria soil isolates collected across the contiguous United States identifies geographically prevalent phylogroups with increased pangenome openness and recombination, as a result of adaptation to variable environments.

CRISPR-Cas systems provide their prokaryotic hosts with sequence-specific immunity to foreign genetic elements, including bacteriophages and plasmids. While most interfere with phage infection though cleavage of viral DNA, type VI CRISPR systems use the RNA-guided nuclease Cas13 to recognize mRNA targets. Upon engaging with target RNA, Cas13 cleaves both phage and host transcripts nonspecifically, leading to a state of cell dormancy that is incompatible with phage propagation. However, whether and how infected cells recover from dormancy is not clear. Here we show that type VI CRISPR systems frequently co-occur with DNA-cleaving restriction modification (RM) systems. Using genetics and microscopy, we show that Cas13 and RM systems synergize in anti-phage defense in the natural type VI CRISPR host Listeria seeligeri. Cleavage of the phage genome by RM removes the source of phage transcripts, enabling cells to recover from Cas13-induced cellular dormancy. We find that Cas13 and RM systems operating simultaneously eliminate phage DNA and neutralize infection more effectively than either defense alone. Thus, cells harboring both defense systems exhibit robust anti-phage immunity and survive infection. Our work therefore reveals that type VI CRISPR immunity is cell-autonomous and non-abortive, if paired with RM or similar DNA-targeting defenses. The ability of an abortive response to be resolved by the actions of another anti-phage defense has implications for the roles of diverse host-directed immune systems in bacteria.

A potential host-derived probiotic, Bacillus subtilis 6-3-1, was successfully screened from 768 isolates from the intestines of healthy hybrid grouper (Epinephelus fuscoguttatus ♀ × Epinephelus lanceolatus ♂) based on multiple probiotic characteristics in vitro assays, such as, non-hemolytic activity, extracellular enzyme activity, inhibitory activity against pathogens, tolerance to gastrointestinal stress, cell surface hydrophobicity, autoaggregation, and antibiotic susceptibility. Eight weeks of feeding trial revealed that dietary supplementation of B. subtilis 6-3-1 at all three concentrations (1 × 106 CFU g−1 as BS6; 1 × 107 CFU g−1 as BS7; 1 × 108 CFU g−1 as BS8) could promote the growth performance of hybrid groupers to a certain extent at different time points. At the end of 8th week, BS6 and BS8 significantly promoted the weight gain rate (WGR), specific growth rate (SGR) of hybrid groupers. The digestive enzyme activities were also increased in BS6 and BS8 groups comparing with those in control group, except that the increase of amylase activities in BS6 was not significant (P > 0.05). However, BS7 showed the best non-specific immunity stimulating effects among the three concentration groups. While BS7 significantly boosted serum total protein contents, lysozyme (LZM), total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT), and acid phosphatase (ACP) levels, BS6 significantly enhanced serum total protein, LZM activity, and BS8 significantly improved LZM, respiratory bursts activity. B. subtilis 6-3-1 up-regulated the expression of MyD88 in head kidney and intestine and increased villi length (VL) in intestine of BS7 group. It also up-regulated the expression of IgM in head kidney in BS6 group and IgM and TLR1 in intestine of BS8 group. Though all B. subtilis 6-3-1 supplemented groups reduced the cumulative mortality rate post-Vibro harveyi-challenge, BS7 showed the best protection effects among the three concentration groups. In conclusion, with its immune promoting, intestine health enhancing, and V. harveyi resisting effects, BS7 show great potential to be used as a probiotic in hybrid grouper culture.

State-of-the-art nanopore sequencing enables rapid and real-time identification of novel pathogens, which has wide application in various research areas and is an emerging diagnostic tool for infectious diseases including COVID-19. Nanopore translocation enables de novo sequencing with long reads (> 10 kb) of novel genomes, which has advantages over existing short-read sequencing technologies. Biological nanopore sequencing has already achieved success as a technology platform but it is sensitive to empirical factors such as pH and temperature. Alternatively, ångström- and nano-scale solid-state nanopores, especially those based on two-dimensional (2D) membranes, are promising next-generation technologies as they can surpass biological nanopores in the variety of membrane materials, ease of defining pore morphology, higher nucleotide detection sensitivity, and facilitation of novel and hybrid sequencing modalities. Since the discovery of graphene, atomically-thin 2D materials have shown immense potential for the fabrication of nanopores with well-defined geometry, rendering them viable candidates for nanopore sequencing membranes. Here, we review recent progress and future development trends of 2D materials and their ångström- and nano-scale pore-based nucleic acid (NA) sequencing including fabrication techniques and current and emerging sequencing modalities. In addition, we discuss the current challenges of translocation-based nanopore sequencing and provide an outlook on promising future research directions.

Soil samples collected in the Great Smoky Mountains National Park yielded aListeriaisolate that could not be classified to the species level. Whole-genome sequence-based average nucleotide identity BLAST andin silicoDNA-DNA Hybridization analyses confirmed this isolate to be a novelListeria sensu strictospecies with the highest similarity toL. marthii(ANI=93.9%, isDDH=55.9%). Additional whole-genome-based analysis using the Genome Taxonomy Database Toolkit, an automated program for classifying bacterial genomes, further supported delineation as a novelListeria sensu strictospecies, as this tool failed to assign a species identification but identifiedL. marthiias the closest match. Phenotypic and genotypic characterization results indicate that this species is nonpathogenic. Specifically, the novelListeriaspecies described here is phenotypically (i) non-hemolytic and (ii) negative for phosphatidylinositol-specific phospholipase C activity; the draft genome lacks all virulence genes found in theListeriapathogenicity island 1 (LIPI-1), as well as the internalin genesinlAandinlB. While the type strain for the new species is phenotypically catalase-negative (an unusual characteristic forListeria sensu strictospecies), its genome contained an apparently intact catalase gene (kat); hence assessment of this phenotype with future isolates will be important. Rapid species identification systems (ListeriaAPI, VITEK 2, VITEK MS) misidentified this novel species as eitherL. monocytogenes, L. innocua, orL. marthii. We propose the nameL. swaminathanii, and the type strain is FSL L7-0020T(=ATCC TSD-239T).IMPORTANCEL. swaminathaniiis a novelsensu strictospecies that originated from a US National Park, and its place of origin is ultimately preventing this species from achieving valid status. The US National Park Service restricts strain accessibility and open access is currently a prerequisite for species validation. Essentially the only criteria that was not met forL. swaminathaniivalidation is accessibility of the type strain, therefore nomenclature status should not negate the significance of this discovery. As a novelsensu strictospecies,L. swaminathaniiexpands the group of species whose presence is associated with an increased risk of anL. monocytogenescontamination, and therefore could play an important role in public health. While developers ofListeriaspp. detection methods historically only included validly publish species in their validation studies,L. swaminathaniiis unequivocally asensu strictospecies and should be included as well.

Biomechanical and molecular processes of premature cervical remodeling preceding spontaneous preterm birth (sPTB) likely result from interactions between the cervicovaginal microbiota and host immune responses. A non-optimal cervicovaginal microbiota confers increased risk of sPTB. The cervicovaginal space is metabolically active in pregancy; microbiota can produce, modify, and degrade metabolites within this ecosystem. We establish that cervicovaginal metabolomic output clusters by microbial community in pregnancy among Black individuals, revealing increased metabolism within the amino acid and dipeptide pathways as hallmarks of a non-optimal microbiota. Few differences were detected in metabolomic profiles when stratified by birth outcome. The study raises the possibility that metabolites could distinguish women with greater risk of sPTB among those with similar cervicovaginal microbiota, and that metabolites within the amino acid and carbohydrate pathways may play a role in this distinction.

Background The emergence of antimicrobial-resistant (AMR) strains of the important human and animal pathogen Salmonella enterica poses a growing threat to public health. Here, we studied the genome-wide evolution of 90 S. enterica AMR isolates, representing one host adapted serotype (S. Dublin) and two broad host range serotypes (S. Newport and S. Typhimurium). Results AMR S. Typhimurium had a large effective population size, a large and diverse genome, AMR profiles with high diversity, and frequent positive selection and homologous recombination. AMR S. Newport showed a relatively low level of diversity and a relatively clonal population structure. AMR S. Dublin showed evidence for a recent population bottleneck, and the genomes were characterized by a larger number of genes and gene ontology terms specifically absent from this serotype and a significantly higher number of pseudogenes as compared to other two serotypes. Approximately 50% of accessory genes, including specific AMR and putative prophage genes, were significantly over- or under-represented in a given serotype. Approximately 65% of the core genes showed phylogenetic clustering by serotype, including the AMR gene aac (6′)-Iaa. While cell surface proteins were shown to be the main target of positive selection, some proteins with possible functions in AMR and virulence also showed evidence for positive selection. Homologous recombination mainly acted on prophage-associated proteins. Conclusions Our data indicates a strong association between genome content of S. enterica and serotype. Evolutionary patterns observed in S. Typhimurium are consistent with multiple emergence events of AMR strains and/or ecological success of this serotype in different hosts or habitats. Evolutionary patterns of S. Newport suggested that antimicrobial resistance emerged in one single lineage, Lineage IIC. A recent population bottleneck and genome decay observed in AMR S. Dublin are congruent with its narrow host range. Finally, our results suggest the potentially important role of positive selection in the evolution of antimicrobial resistance, host adaptation and serotype diversification in S. enterica. Electronic supplementary material The online version of this article (10.1186/s12862-019-1457-5) contains supplementary material, which is available to authorized users.

Spontaneous preterm birth (sPTB) is a leading cause of maternal and neonatal morbidity and mortality, yet both its prevention and early risk stratification are limited. The vaginal microbiome has been associated with PTB risk, possibly via metabolic or other interactions with its host. Here, we performed untargeted metabolomics on 232 vaginal samples, in which we have previously profiled the microbiota using 16S rRNA gene sequencing. Samples were collected at 20-24 weeks of gestation from women with singleton pregnancies, of which 80 delivered spontaneously before 37 weeks of gestation. We find that the vaginal metabolome correlates with the microbiome and separates into six clusters, three of which are associated with spontaneous preterm birth (sPTB) in Black women. Furthermore, while we identify five metabolites that associate with sPTB, another five associate with sPTB only when stratifying by race. We identify multiple microbial correlations with metabolites associated with sPTB, including intriguing correlations between vaginal bacteria that are considered sub-optimal and metabolites that were enriched in women who delivered at term. We propose that several sPTB-associated metabolites may be exogenous, and investigate another using metabolic models. Notably, we use machine learning models to predict sPTB risk using metabolite levels, weeks to months in advance, with high accuracy. We show that these predictions are more accurate than microbiome-based and maternal covariates-based models. Altogether, our results demonstrate the potential of vaginal metabolites as early biomarkers of sPTB and highlight exogenous exposures as potential risk factors for prematurity.

A total of 27 Listeria isolates that could not be classified to the species level were obtained from soil samples from different locations in the contiguous United States and an agricultural water sample from New York. Whole-genome sequence-based average nucleotide identity blast (ANIb) showed that the 27 isolates form five distinct clusters; for each cluster, all draft genomes showed ANI values of

A total of 27 Listeria isolates that could not be classified to the species level were obtained from soil samples from different locations in the contiguous United States and an agricultural water sample from New York. Whole-genome sequence-based average nucleotide identity blast (ANIb) showed that the 27 isolates form five distinct clusters; for each cluster, all draft genomes showed ANI values of Listeria species, indicating that each cluster represents a novel species. Of the five novel species, three cluster with the Listeria sensu stricto clade and two cluster with sensu lato. One of the novel sensu stricto species, designated L. cossartiae sp. nov., contains two subclusters with an average ANI similarity of 94.9%, which were designated as subspecies. The proposed three novel sensu stricto species (including two subspecies) are Listeria farberi sp. nov. (type strain FSL L7-0091T=CCUG 74668T=LMG 31917T; maximum ANI 91.9 % to L. innocua ), Listeria immobilis sp. nov. (type strain FSL L7-1519T=CCUG 74666T=LMG 31920T; maximum ANI 87.4 % to L. ivanovii subsp. londoniensis ) and Listeria cossartiae sp. nov. [subsp. cossartiae (type strain FSL L7-1447T=CCUG 74667T=LMG 31919T; maximum ANI 93.4 % to L. marthii ) and subsp. cayugensis (type strain FSL L7-0993T=CCUG 74670T=LMG 31918T; maximum ANI 94.7 % to L. marthii ). The two proposed novel sensu lato species are Listeria portnoyi sp. nov. (type strain FSL L7-1582T=CCUG 74671T=LMG 31921T; maximum ANI value of 88.9 % to L. cornellensis and 89.2 % to L. newyorkensis ) and Listeria rustica sp. nov. (type strain FSL W9-0585T=CCUG 74665T=LMG 31922T; maximum ANI value of 88.7 % to L. cornellensis and 88.9 % to L . newyorkensis ). L. immobilis is the first sensu stricto species isolated to date that is non-motile. All five of the novel species are non-haemolytic and negative for phosphatidylinositol-specific phospholipase C activity; the draft genomes lack the virulence genes found in Listeria pathogenicity island 1 (LIPI-1), and the internalin genes inlA and inlB, indicating that they are non-pathogenic.

High-quality habitats for wildlife (e.g., forest) provide essential ecosystem services while increasing species diversity and habitat connectivity. Unfortunately, the presence of such habitats adjacent to produce fields may increase risk for contamination of fruits and vegetables by enteric bacteria, including Escherichia coli. E. coli survives in extrahost environments (e.g., soil) and could be dispersed across landscapes by wildlife. Understanding how terrestrial landscapes impact the distribution of soil E. coli strains is of importance in assessing the contamination risk of agricultural products. Here, using multilocus sequence typing, we characterized 938 E. coli soil isolates collected from two watersheds with different landscape patterns in New York State, USA, and compared the distribution of E. coli and the influence that environmental selection and dispersal have on the distribution between these two watersheds. Results showed that for the watershed with widespread produce fields, sparse forests, and limited interaction between the two land use types, E. coli composition was significantly different between produce field sites and forest sites; this distribution appears to be shaped by relatively strong environmental selection, likely from soil phosphorus, and slight dispersal limitation. For the watershed with more forested areas and stronger interaction between produce field sites and forest sites, E. coli composition between these two land use types was relatively homogeneous; this distribution appeared to be a consequence of wildlife-driven dispersal, inferred by competing models. Collectively, our results suggest that terrestrial landscape attributes could impact the biogeographic pattern of enteric bacteria by adjusting the importance of environmental selection and dispersal. IMPORTANCE Understanding the ecology of enteric bacteria in extrahost environments is important for the development and implementation of strategies to minimize preharvest contamination of produce with enteric pathogens. Our findings suggest that watershed landscape is an important factor influencing the importance of ecological drivers and dispersal patterns of E. coli. Agricultural areas in such watersheds may have a higher risk of produce contamination due to fewer environmental constraints and higher potential of dispersal of enteric bacteria between locations. Thus, there is a perceived trade-off between priorities of environmental conservation and public health in on-farm food safety, with limited ecological data supporting or refuting the role of wildlife in dispersing pathogens under normal operating conditions. By combining field sampling and spatial modeling, we explored ecological principles underlying the biogeographic pattern of enteric bacteria at the regional level, which can benefit agricultural, environmental, and public health scientists who aim to reduce the risk of food contamination by enteric bacteria while minimizing negative impacts on wildlife habitats.

Natural bacterial populations can display enormous genomic diversity, primarily in the form of gene content variation caused by the frequent exchange of DNA with the local environment. However, the ecological drivers of genomic variability and the role of selection remain controversial. Here, we address this gap by developing a nationwide atlas of 1,854 Listeria isolates, collected systematically from soils across the contiguous United States. We found that Listeria was present across a wide range of environmental parameters, being mainly controlled by soil moisture, molybdenum and salinity concentrations. Whole-genome data from 594 representative strains allowed us to decompose Listeria diversity into 12 phylogroups, each with large differences in habitat breadth and endemism. 'Cosmopolitan' phylogroups, prevalent across many different habitats, had more open pangenomes and displayed weaker linkage disequilibrium, reflecting higher rates of gene gain and loss, and allele exchange than phylogroups with narrow habitat ranges. Cosmopolitan phylogroups also had a large fraction of genes affected by positive selection. The effect of positive selection was more pronounced in the phylogroup-specific core genome, suggesting that lineage-specific core genes are important drivers of adaptation. These results indicate that genome flexibility and recombination are the consequence of selection to survive in variable environments.

High-quality habitats for wildlife (e.g., forest) provide essential ecosystem services while increasing species diversity and habitat connectivity. Unfortunately, presence of such habitats adjacent to produce fields may increase risk for contamination of fruits and vegetables by enteric bacteria, including Escherichia coli. E. coli survives in extra-host environments (e.g., soil) and could disperse across landscapes by wildlife. Understanding how terrestrial landscapes impact the distribution of soil E. coli is of importance in assessing the contamination risk of agricultural products. Here, using multi-locus sequence typing, we characterized 938 E. coli soil isolates collected from two watersheds with different landscape patterns in New York state, USA, and compared the distribution of E. coli and the influence of two ecological forces (environmental selection and dispersal) on the distribution between these two watersheds. Results showed that for the watershed with widespread produce fields, sparse forests, and limited interaction between the two land-use types, E. coli composition was significantly different between produce field sites and forest sites; this distribution was shaped by relatively strong environmental selection likely from soil phosphorus and slight dispersal limitation. For the watershed with more forested areas and stronger interaction between produce field sites and forest sites, E. coli composition between these two land-use types was relatively homogeneous; this distribution appeared to a consequence of wildlife-driven dispersal, inferred by competing models. Collectively, our results suggest that terrestrial landscape attributes could impact the biogeographic pattern of enteric bacteria by adjusting the importance of environmental selection and dispersal.IMPORTANCEUnderstanding the ecology of enteric bacteria in extra-host environments is important to allow for development and implementation of strategies to minimize pre-harvest contamination of produce with enteric pathogens. Our findings suggest that watershed landscape is an important factor influencing the importance of ecological drivers and dispersal patterns of E. coli. For watersheds with widespread produce fields, E. coli appears to experience local adaptation, possibly due to exposure to environmental stresses associated with agricultural activities. In contrast, for watersheds with high forest coverage we found evidence for wildlife-driven dispersal of E. coli, which might facilitate more frequent genetic exchange in this environment. Agricultural areas in such watersheds may have a higher risk of produce contamination due to less environmental constraints and higher potential of dispersal of enteric bacteria between locations. The significance of our research lies in exploring ecological principles underlying the biogeographic pattern of enteric bacteria at the regional level, which can inform agricultural, environmental and public health scientists that aim to reduce the risk of food contamination by enteric bacteria.

An infallible inhibitor of Cas13 CRISPR-Cas13 protects bacterial populations from viral infections by indiscriminately destroying the RNA of the cell and its invader, simultaneously arresting the growth of infected hosts and the spread of the virus. This response is mediated by the Cas13 nuclease, which unleashes massive RNA degradation after recognition of viral transcripts that are complementary to its guide RNA. Meeske et al. discovered AcrVIA1, a viral-encoded inhibitor that binds to Cas13 to occlude the RNA guide and prevent the activation of the nuclease (see the Perspective by Barrangou and Sontheimer). As opposed to inhibitors of DNA-cleaving CRISPR-Cas systems, which require multiple infections to neutralize all Cas nucleases of the host, production of AcrVIA1 by a single virus is sufficient to overcome the CRISPR-Cas13 response. Science , this issue p. 54 ; see also p. 31

SummaryGenetic variation in a pathogen, including the causative agent of salmonellosis, Salmonella enterica, can occur as a result of eco-evolutionary forces triggered by dissimilarities of ecological niches. Here, we applied comparative genomics to study 90 antimicrobial resistant (AMR) S. enterica isolates from bovine and human hosts in New York state and Washington state to understand host- and geographic-associated population structure. Results revealed distinct presence/absence profiles of functional genes and pseudogenes (e.g., virulence genes) associated with bovine and human isolates. Notably, bovine isolates contained significantly more transposase genes but fewer transposase pseudogenes than human isolates, suggesting the occurrence of large-scale transposition in genomes of bovine and human isolates at different times. The high correlation between transposase genes and AMR genes, as well as plasmid replicons, highlights the potential role of horizontally transferred transposons in promoting adaptation to antibiotics. By contrast, a number of potentially geographic-associated single-nucleotide polymorphisms (SNPs), rather than geographic-associated genes, were identified. Interestingly, 38% of these SNPs were in genes annotated as cell surface protein-encoding genes, including some essential for antibiotic resistance and host colonization. Overall, different evolutionary forces and limited recent inter-population transmission appear to shape AMR S. enterica population structure in different hosts and geographic origins.Originality/Significance StatementSalmonella enterica, which is the causative agent of salmonellosis, poses a growing public health concern due to the emergence and spread of antimicrobial resistant (AMR) strains. The mechanisms underlying the population structure associated with different hosts and geographic origins of AMR S. enterica are underexplored due to limited genome-wide studies assessing the impact of ecological niches on genetic variations. By employing comparative genomics, our study provided insights into the genomic profiles of AMR S. enterica associated with two distinct hosts and two distant geographic locations, improving the mechanistic understanding of how bacterial population structure is shaped by different ecological niches. Our findings have broad implications for elucidating the impact of ecological and evolutionary forces on the adaptation, antimicrobial resistance, and pathogenicity of bacteria. Also, specific genetic markers we identified may help predict host or geographic origin of AMR Salmonella isolates, which could benefit the source tracking (e.g., host and geographic origins) of human disease cases and contamination events caused by AMR S. enterica.

The revolution of molecular techniques in the field of microbial ecology not only allows an in-depth exploration of abundant bacteria, but also confirms the existence of “rare biosphere.” However, the mechanisms underlying the assembly process of abundant and rare bacteria in plateau lakes are still poorly understood. In the present study, distribution of abundant and rare bacteria in 21 lakes on Yungui Plateau, China and the relative importance of deterministic and neutral factors to their biogeographic patterns were examined. Results surprisingly showed that similar biogeographic patterns of rare and abundant taxa were generally shaped via similar assembly mechanisms. For both bacterial groups, deterministic process strongly influenced the distribution, while neutral process was not at play. In addition, they both exhibited a weak distance–decay relationship. Nevertheless, environmental drivers of rare and abundant taxa and their importance were not exactly the same. Water physicochemical property was the most dominant driver shaping the biogeographic pattern of rare taxa, followed by lake morphology and watershed land-use, while for abundant taxa, watershed land-use was the most dominant driver and lake morphology did not play a significant role. This study suggested that rare bacteria in high-altitude aquatic ecosystems might experience equally low extinction risk and respond to environmental changes in a similar manner to abundant ones, but their ecological niches and functions were not identical. To fully understand the assembly mechanism of bacterial community, it is necessary to differentiate the community by traits of taxa.

Figure S1. The distribution of AMR gene numbers of AMR S. Dublin, AMR S. Newport, and AMR S. Typhimurium isolates. The AMR gene number of AMR S. Dublin is significantly different from that of AMR S. Newport and AMR S. Typhimurium. Figure S2. The distribution of PPG gene numbers of AMR S. Dublin, AMR S. Newport, and AMR S. Typhimurium isolates. Figure S3. The distribution of pseudogene numbers in S. Dublin, S. Newport, and S. Typhimurium. Figure S4. Gene tree of AAC (6′)-Iaa inferred by maximum likelihood method. Tree is rooted by midpoint. Bootstrap values > 70% are presented on the tree. S. Dublin is indicated by blue, S. Newport by blue, and S. Typhimurium by red. Figure S5. Maximum likelihood tree of AMR S. Newport isolates, and Lineage II (sub-lineages - IIA, IIB and IIC) and Lineage III reference isolates. Tree is rooted by midpoint. Bootstrap values of major clades are presented on the tree. Reference isolates are indicated by red. (DOCX 681 kb)

Excessive nutrient loads resulted in cascading trophic effects and ecosystem responses. Aims of this study were to determine if the thresholds in nutrient gradient related to phytoplankton community composition could be identified in eutrophic lake, and further to analyze the change of phytoplankton assemblage along the nutrient concentration based on Threshold Indicator Taxa ANalysis (TITAN). The results presented the significant community thresholds estimate for negative taxa declining at 1.650 mg/L TN and 131.5 μg/L TP, as well as simultaneously increasing for positive taxa at 1.665 mg/L TN and 151.5 μg/L TP along nutrient enrichment gradient. However, there was unremarkable change point determined for TN:TP ratios in Lake Dianchi. Elevated TN and TP altered the phytoplankton assemblage, even may induce the fade of algal blooms across the threshold in the hypertrophic lake. The findings could provide implications for deeply deciphering abrupt transitions for phytoplankton assemblage and developing nutrient tactics to protect the lake ecosystems.

Aeromonas schubertii is a conditional pathogen widely distributed in aquatic environments, yet it was only recently reported as a causative pathogen in fish. This study investigated antioxidant capacity, non-specific immunity, histopathology and immune-related gene expression profiles in Nile tilapia Oreochromis niloticus injected with A. schubertii. The results show that A. schubertii infection stimulated the activation of antioxidant activity and non-specific immunity in Nile tilapia. Histopathological analysis revealed that inflammation existed in the liver, spleen, head kidney, brain and intestine of the infected fish. Quantitative real-time PCR was performed to measure mRNA expression levels of six immune-related genes (interleukin-1β, tumor necrosis factor-α, cyclooxygenase-2, transforming growth factor beta, interleukin-10 and heat shock protein 70) in the intestine and head kidney at different timepoints after experimental infection. The results demonstrated clear transcriptional activation of these immune-related genes in the intestine and head-kidney tissues. To our knowledge, this is the first study to investigate the host response to A. schubertii infection in a fish. These results will contribute to further understanding of the pathogenesis and host defense system in fishes following infection with A. schubertii.

Naphthenic acids (NAs) have been gaining recognition in recent years as potentially harmful environmental contaminants. Few studies have focused on the potential ecotoxicity of NAs to terrestrial environment. In this study, the responses of antioxidant system and lipid peroxidation and DNA damage were investigated after exposing Eisenia fetida to soil contaminated with NAs. The results indicated that NAs induced a significant increase (p < 0.05) in superoxide dismutase and catalase enzyme activities. The glutathione peroxidase enzyme activities were significantly inhibited (p < 0.05) in the medium and high dose treatments. An increase in malondialidehyde indicated that NAs could cause cellular lipid peroxidation in the tested earthworms. The percentage of DNA in the tail of comet assay of coelomocytes as an indication of DNA damage increased after treatment with different doses of NAs, and a dose-dependent DNA damage of coelomocytes was found. In conclusion, oxidative stress caused by NAs exposure induces physiological responses and genotoxicity on earthworms. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1337-1343, 2016.

Geographic location and physicochemical properties are thought to represent major factors that shape soil bacterial community abundance and diversity. Crude oil contamination is becoming a notable concern with respect to soil property variation; however, the quantifiable influences of geographic location, physicochemical properties, and oil contamination are still poorly understood. In this study, the 16S ribosomal RNA genes of bacteria in the four oil fields in China were analyzed by using pyrosequencing. Results showed that physicochemical properties were the most dominant factor of bacterial community distribution, followed by geographical location. Oil contamination was a driving factor whose indirect influence was stronger than its direct influence. Under the impact of these three factors, different oil fields presented diversified and distinguishable bacterial community features. The soil of sites with the highest total petroleum hydrocarbon content (HB), nitrogen content (DQ), and phosphorus content (XJ) contained the largest proportion of functional groups participating in hydrocarbon degradation, nitrogen turnover, and phosphorus turnover, respectively. The first dominant phylum of the site with loam soil texture (HB) was Actinobacteria instead of Proteobacteria in other sites with sandy or sandy loam soil texture (DQ, SL, XJ). The site with the highest salinization and alkalization (SL) exhibited the largest proportion of unique local bacteria. The site that was located in the desert with extremely low precipitation (XJ) had the most diversified bacteria distribution. The bacterial community diversity was strongly influenced by soil physicochemical properties.

Naphthenic acids (NAs) are a toxic complex mixture of carboxylic acids occurring naturally in petroleum. Due to the serious potential risks of NAs towards terrestrial ecology during crude oil exploration and production processes, and the lack of efficient methodologies for extraction and analysis of these compounds, the goal of this study is to detail the development of a routine method for extraction, and quantitative and qualitative analysis of NAs in oil contaminated soils. Solid phase extraction using a MAX cartridge was employed in combination with GC/MS. Ethyl acetate with 2% formic acid used as an elution solvent showed the best recoveries of NAs (98.36–112.35%). The total NA concentration and NA profiles of oil contaminated soil samples from 4 oil fields in China were examined. High concentrations of NAs (maximum, 132.91 mg kg−1) were detected in soils, which implied toxic and estrogenic risks for human and terrestrial organisms. Different profiles of NA mixtures were observed in soils from 4 different oil fields, and evaporation and biodegradation could influence the compounds of NAs. The authors present the first feasible method for analysis of NAs in soil.

Sequencing of single genes remains an important tool that allows the rapid classification of bacteria. Sequencing of a portion of sigB , which encodes a stress-responsive alternative sigma factor, has emerged as a commonly used molecular tool for the initial characterization of diverse Listeria isolates. In this study, evolutionary approaches were used to assess the validity of sigB allelic typing for Listeria . For a data set of 4,280 isolates, sigB allelic typing showed a Simpson's index of diversity of 0.96. Analyses of 164 sigB allelic types (ATs) found among the 6 Listeria sensu stricto species, representing these 4,280 isolates, indicate that neither frequent homologous recombination nor positive selection significantly contributed to the evolution of sigB , confirming its genetic stability. The molecular clock test provided evidence for unequal evolution rates across clades; Listeria welshimeri displayed the lowest sigB diversity and was the only species in which sigB evolved in a clocklike manner, implying a unique natural history. Among the four L. monocytogenes lineages, sigB evolution followed a molecular clock only in lineage IV. Moreover, sigB displayed a significant negative Tajima D value in lineage II, suggesting a recent population bottleneck followed by lineage expansion. The absence of positive selection along with the violation of the molecular clock suggested a nearly neutral mechanism of Listeria sensu stricto sigB evolution. While comparison with a whole-genome sequence-based phylogeny revealed that the sigB phylogeny did not correctly reflect the ancestry of L. monocytogenes lineage IV, the availability of a large sigB AT database allowed accurate species classification. IMPORTANCE sigB allelic typing has been widely used for species delineation and subtyping of Listeria . However, an informative evaluation of this method from an evolutionary perspective was missing. Our data indicate that the genetic stability of sigB is affected by neither frequent homologous recombination nor positive selection, which supports that sigB allelic typing provides reliable subtyping and classification of Listeria sensu stricto strains. However, multigene data are required for accurate phylogeny reconstruction of Listeria . This study thus contributes to a better understanding of the evolution of sigB and confirms the robustness of the sigB subtyping system for Listeria .

Sequencing of single genes remains an important tool that allows the rapid classification of bacteria. Sequencing of a portion of sigB, which encodes a stress-responsive alternative sigma factor, has emerged as a commonly used molecular tool for the initial characterization of diverse Listeria isolates. In this study, evolutionary approaches were used to assess the validity of sigB allelic typing for Listeria. For a data set of 4,280 isolates, sigB allelic typing showed a Simpson's index of diversity of 0.96. Analyses of 164 sigB allelic types (ATs) found among the 6 Listeria sensu stricto species, representing these 4,280 isolates, indicate that neither frequent homologous recombination nor positive selection significantly contributed to the evolution of sigB, confirming its genetic stability. The molecular clock test provided evidence for unequal evolution rates across clades; Listeria welshimeri displayed the lowest sigB diversity and was the only species in which sigB evolved in a clocklike manner, implying a unique natural history. Among the four L. monocytogenes lineages, sigB evolution followed a molecular clock only in lineage IV. Moreover, sigB displayed a significant negative Tajima D value in lineage II, suggesting a recent population bottleneck followed by lineage expansion. The absence of positive selection along with the violation of the molecular clock suggested a nearly neutral mechanism of Listeria sensu stricto sigB evolution. While comparison with a whole-genome sequence-based phylogeny revealed that the sigB phylogeny did not correctly reflect the ancestry of L. monocytogenes lineage IV, the availability of a large sigB AT database allowed accurate species classification.

Aquatic ecosystems are ecologically important, but continuously threatened by a growing number of human induced changes. This study evaluates the research trends of "aquatic ecosystem" between 1992 and 2011 in journals of all subject categories of the science citation index and social sciences citation index. The analyzed parameters include publication output, cited publication, document type, language, distributions of journal, author, country and institutes, and analysis of author keywords and keywords plus. The results showed that over the past two decades, there was a consistent growth in publication output with involvement of increasing number of countries and institutions, and North America was still the leading region in the subject. Classification of the top 30 author keywords indicated that more research attentions were paid to the study on aquatic organism, water environment and aquatic ecosystem condition. Aquatic ecosystem, water quality, and fish were the top three most frequently used author keywords. In addition, owing to its significant impact on aquatic ecosystems, climate change has been placed crucial emphasis recently. Aquatic ecosystem research trend was shifting from water environment to aquatic ecosystem wide issues.

Plateau lakes are important ecosystems with diverse ecological functions. Cyanobacteria play a key role in plateau lakes as primary producers. However, they are threatening when dense blooms occur. Identifying cyanobacteiral biogeography and the mechanism of assembly processes shaping the distribution of cyanobacteria in plateau lakes is critical for understanding cyanobacterial ecology and applying it to lake management. In the present study, the biogeographic pattern and importance of neutral and niche processes in assembly of cyanobacteria in 21 lakes on Yungui Plateau, China were examined. Results showed that cyanobacteria exhibit unique biogeographic pattern, and most of them have a narrow habitat preference in plateau lakes. They were assembled via niche processes driven by water physicochemical property, lake morphology and watershed land-use, which explained 62.4% of the biological variation. Neutral processes were not at play. Water physicochemical property (key variables - dissolved oxygen, salinity, trophic status and pH) was the most dominant driver shaping its unique biogeographic pattern. Watershed land-use especially urban land, water body and agricultural land also exhibited a strong impact on cyanobacterial distribution, followed by lake morphology. As most of the cyanobacteiral genus detected in these plateau lakes were potential toxin-producers, this study indicated that in order to protect waters from toxic-bloom in the future, reducing nutrient loading and land-use practices are two practical approaches in plateau lake management.

The mechanisms of community assembly are a central focus in the field of microbial ecology. However, to what extent these mechanisms differ in importance by traits of groups is poorly understood. Here we quantified the importance of neutral and niche processes in community assembly for bacteria, habitat specialists and generalists in 21 plateau lakes of China. Results showed that both neutral and niche processes played a critical role in the assembly of entire bacterial communities, shaping a unique biogeographical pattern. A few habitat generalists and many specialists were identified. Interestingly, habitat specialists were only governed by niche process, with seven significant environmental variables—salinity, dissolved oxygen, water transparency, total phosphorus, ammonium-nitrogen, temperature and total nitrogen—independently explaining 40.3% of the biological variation. By contrast, habitat generalists were strongly driven by neutral process, with 50.9% of the variation of detection frequency explained in neutral community model. Only three environmental variables—salinity, total nitrogen and dissolved oxygen—significantly affected the distribution of habitat generalists, independently explaining 13.6% of the variation. Governed by different assembly mechanisms, habitat specialists and generalists presented disparate biogeographical patterns. Our result emphasizes the importance of investigating the bacterial community assembly at more refined levels than entire communities.

Elevated environmental pressures negatively affect the bacterial community structure. However, little knowledge about the nonlinear responses of spatially related environmental variable across multiple plateau lake ecosystems on bacterioplankton communities has been gathered. Here, we used 454 pyrosequencing of 16S rRNA genes to study the associations of bacterial communities in terms of environmental characteristics as well as the potentially ecological threshold-inducing shifts of the bacterial community structure along the key environmental variables based on hypothesized structural equation models and the SEGMENTED method in 21 plateau lakes. Our results showed that water transparency was the major driving force and that total nitrogen was more significant than total phosphorus in determining the taxon composition of the bacterioplankton community. Significant community threshold estimates for bacterioplankton were observed at 7.36 for pH and 25.6% for the percentage of the agricultural area, while the remarkable change point of the cyanobacteria community structure responding to pH was at 7.74. Furthermore, the findings indicated that increasing nutrient loads can induce a distinct shift in dominance from Proteobacteria to Cyanobacteria, as well as a sharp decrease and adjacent increase when crossing the change point for Actinobacteria and Bacteroidetes along the gradient of the agricultural area.

Since total petroleum hydrocarbons (TPH) are toxic and persistent in environments, studying the impact of oil contamination on microbial communities in different soils is vital to oil production engineering, effective soil management and pollution control. This study analyzed the impact of oil contamination on the structure, activity and function in carbon metabolism of microbial communities of Chernozem soil from Daqing oil field and Cinnamon soil from Huabei oil field through both culture-dependent techniques and a culture-independent technique-pyrosequencing. Results revealed that pristine microbial communities in these two soils presented disparate patterns, where Cinnamon soil showed higher abundance of alkane, (polycyclic aromatic hydrocarbons) PAHs and TPH degraders, number of cultivable microbes, bacterial richness, bacterial biodiversity, and stronger microbial activity and function in carbon metabolism than Chernozem soil. It suggested that complicated properties of microbes and soils resulted in the difference in soil microbial patterns. However, the changes of microbial communities caused by oil contamination were similar in respect of two dominant phenomena. Firstly, the microbial community structures were greatly changed, with higher abundance, higher bacterial biodiversity, occurrence of Candidate_division_BRC1 and TAO6, disappearance of BD1-5 and Candidate_division_OD1, dominance of Streptomyces, higher percentage of hydrocarbon-degrading groups, and lower percentage of nitrogen-transforming groups. Secondly, microbial activity and function in carbon metabolism were significantly enhanced. Based on the characteristics of microbial communities in the two soils, appropriate strategy for in situ bioremediation was provided for each oil field. This research underscored the usefulness of combination of culture-dependent techniques and next-generation sequencing techniques both to unravel the microbial patterns and understand the ecological impact of contamination.

Naphthenic acids (NAs) have been gaining recognition in recent years as potentially harmful environmental contaminants. Few studies have focused on the potential ecotoxicity of NAs to terrestrial environment. In this study, the responses of antioxidant system and lipid peroxidation and DNA damage were investigated after exposing Eisenia fetida to soil contaminated with NAs. The results indicated that NAs induced a significant increase (p 0.05) in superoxide dismutase and catalase enzyme activities. The glutathione peroxidase enzyme activities were significantly inhibited (p 0.05) in the medium and high dose treatments. An increase in malondialidehyde indicated that NAs could cause cellular lipid peroxidation in the tested earthworms. The percentage of DNA in the tail of comet assay of coelomocytes as an indication of DNA damage increased after treatment with different doses of NAs, and a dose-dependent DNA damage of coelomocytes was found. In conclusion, oxidative stress caused by NAs exposure induces physiological responses and genotoxicity on earthworms. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1337-1343, 2016.