Background: The diverse microbial populations that inhabit pristine aquifers are known to catalyze critical in situ biogeochemical reactions, yet little is known about how the structure and diversity of this subsurface community correlates with and impacts upon groundwater chemistry. Herein we examine 8,786 bacterial and 8,166 archaeal 16S rRNA gene sequences from an array of monitoring wells in the Mahomet aquifer of east-central Illinois. Using multivariate statistical analyses we provide a comparative analysis of the relationship between groundwater chemistry and the microbial communities attached to aquifer sediment along with those suspended in groundwater.Results: Statistical analyses of 16S rRNA gene sequences showed a clear distinction between attached and suspended communities; with iron-reducing bacteria far more abundant in attached samples than suspended, while archaeal clones related to groups associated with anaerobic methane oxidation and deep subsurface gold mines (ANME-2D and SAGMEG-1, respectively) distinguished the suspended community from the attached. Within the attached bacterial community, cloned sequences most closely related to the sulfate-reducingDesulfobacterandDesulfobulbusgenera represented 20% of the bacterial community in wells where the concentration of sulfate in groundwater was high (> 0.2 mM), compared to only 3% in wells with less sulfate. Sequences related to the genusGeobacter, a genus containing ferric-iron reducers, were of nearly equal abundance (15%) to the sulfate reducers under high sulfate conditions, however their relative abundance increased to 34% when sulfate concentrations were < 0.03 mM. Also, in areas where sulfate concentrations were 0.2 mM sulfate. In the suspended fraction of wells where the concentration of sulfate was between 0.03 and 0.2 mM, the archaeal community was dominated by sequences most closely related to the ANME-2D, a group of archaea known for anaerobically oxidizing methane. Based on available energy (∆G_A) estimations, results varied little for both sulfate reduction and methanogenesis throughout all wells studied, but could favor anaerobic oxidation of methane (AOM) in wells containing minimal sulfate and dihydrogen, suggesting AOM coupled with H₂-oxidizing organisms such as sulfate or iron reducers could be an important pathway occurring in the Mahomet aquifer.Conclusions: Overall, the results show several distinct factors control the composition of microbial communities in the Mahomet aquifer. Bacteria that respire insoluble substrates such as iron oxides, i.e.Geobacter, comprise a greater abundance of the attached community than the suspended regardless of groundwater chemistry. Differences in community structure driven by the concentration of sulfate point to a clear link between the availability of substrate and the abundance of certain functional groups, particularly iron reducers, sulfate reducers, methanogens, and methanotrophs. Integrating both geochemical and microbiological observations suggest that the relationships between these functional groups could be driven in part by mutualism, especially between ferric-iron and sulfate reducers.

Although the source of drinking water (DW) used in hospitals is commonly disinfected, biofilms forming on water pipelines are a refuge for bacteria, including possible pathogens that survive different disinfection strategies. These biofilm communities are only beginning to be explored by culture-independent techniques that circumvent the limitations of conventional monitoring efforts. Hence, theories regarding the frequency of opportunistic pathogens in DW biofilms and how biofilm members withstand high doses of disinfectants and/or chlorine residuals in the water supply remain speculative. The aim of this study was to characterize the composition of microbial communities growing on five hospital shower hoses using both 16S rRNA gene sequencing of bacterial isolates and whole-genome shotgun metagenome sequencing. The resulting data revealed aMycobacterium-like population, closely related toMycobacterium rhodesiaeandMycobacterium tusciae, to be the predominant taxon in all five samples, and its nearly complete draft genome sequence was recovered. In contrast, the fraction recovered by culture was mostly affiliated withProteobacteria, including members of the generaSphingomonas,Blastomonas, andPorphyrobacter.The biofilm community harbored genes related to disinfectant tolerance (2.34% of the total annotated proteins) and a lower abundance of virulence determinants related to colonization and evasion of the host immune system. Additionally, genes potentially conferring resistance to β-lactam, aminoglycoside, amphenicol, and quinolone antibiotics were detected. Collectively, our results underscore the need to understand the microbiome of DW biofilms using metagenomic approaches. This information might lead to more robust management practices that minimize the risks associated with exposure to opportunistic pathogens in hospitals.

The detection of environmental enterococci has been determined primarily by using culture-based techniques that might exclude some enterococcal species as well as those that are nonculturable. To address this, the relative abundances of enterococci were examined by challenging fecal and water samples against a currently available genus-specific assay (Entero1). To determine the diversity of enterococcal species, 16S rRNA gene-based group-specific quantitative PCR (qPCR) assays were developed and evaluated against eight of the most common environmental enterococcal species. Partial 16S rRNA gene sequences of 439 presumptive environmental enterococcal strains were analyzed to study further the diversity of enterococci and to confirm the specificities of group-specific assays. The group-specific qPCR assays showed relatively high amplification rates with targeted species (>98%), although some assays cross-amplified with nontargeted species (1.3 to 6.5%). The results with the group-specific assays also showed that different enterococcal species co-occurred in most fecal samples. The most abundant enterococci in water and fecal samples were Enterococcus faecalis and Enterococcus faecium , although we identified more water isolates as Enterococcus casseliflavus than as any of the other species. The prevalence of the Entero1 marker was in agreement with the combined number of positive signals determined by the group-specific assays in most fecal samples, except in gull feces. On the other hand, the number of group-specific assay signals was lower in all water samples tested, suggesting that other enterococcal species are present in these samples. While the results highlight the value of genus- and group-specific assays for detecting the major enterococcal groups in environmental water samples, additional studies are needed to determine further the diversity, distributions, and relative abundances of all enterococcal species found in water.