Abstract
BackgroundHospital wastewaters contain fecal material from a large number of individuals, of which many are undergoing antibiotic therapy. It is, thus, plausible that hospital wastewaters could provide opportunities to find novel carbapenemases and other resistance genes not yet described in clinical strains. Our aim was therefore to investigate the microbiota and antibiotic resistome of hospital effluent collected from the city of Mumbai, India, with a special focus on identifying novel carbapenemases.ResultsShotgun metagenomics revealed a total of 112 different mobile antibiotic resistance gene types, conferring resistance against almost all classes of antibiotics. Beta-lactamase genes, including encoding clinically important carbapenemases, such as NDM, VIM, IMP, KPC, and OXA-48, were abundant. NDM (0.9% relative abundance to 16S rRNA genes) was the most common carbapenemase gene, followed by OXA-58 (0.84% relative abundance to 16S rRNA genes). Among the investigated mobile genetic elements, class 1 integrons (11% relative abundance to 16S rRNA genes) were the most abundant. The genus Acinetobacter accounted for as many as 30% of the total 16S rRNA reads, with A. baumannii accounting for an estimated 2.5%. High throughput sequencing of amplified integron gene cassettes identified a novel functional variant of an IMP-type (proposed IMP-81) carbapenemase gene (eight aa substitutions) along with recently described novel resistance genes like sul4 and blaRSA1. Using a computational hidden Markov model, we detected 27 unique metallo-beta-lactamase (MBL) genes in the shotgun data, of which nine were novel subclass B1 genes, one novel subclass B2, and 10 novel subclass B3 genes. Six of the seven novel MBL genes were functional when expressed in Escherichia coli.ConclusionBy exploring hospital wastewater from India, our understanding of the diversity of carbapenemases has been extended. The study also demonstrates that the microbiota of hospital wastewater can serve as a reservoir of novel resistance genes, including previously uncharacterized carbapenemases with the potential to spread further.
Highlights
Hospital wastewaters contain fecal material from a large number of individuals, of which many are undergoing antibiotic therapy
Environmental and commensal microbiota serves as sources for antibiotic resistance genes (ARGs) that emerge over time in pathogens
A total of 193,098 reads (0.11% of the total reads) matched to ARGs. These accounted for 0.844 copies of ARGs per 16S rRNA gene
Summary
Hospital wastewaters contain fecal material from a large number of individuals, of which many are undergoing antibiotic therapy. HMM accurately predicts the gene fragments belonging to specific gene classes, based on evolutionarily conserved domains [11] This leads to accurate detection of both known and previously undescribed resistance genes in genomic and metagenomic sequence data. Functional metagenomics is another strategy that has the ability to identify novel ARGs without apparent similarities to known ARGs as it is based on a functional selection of DNA fragments expressed in a surrogate host like Escherichia coli [12]. Several studies, using functional metagenomics, have reported novel resistance genes from a variety of environments like human gut, soil, and seawater [14,15,16,17,18]
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