Abstract
The occurrence of pathogenic bacteria in drinking water distribution systems (DWDSs) is a major health concern, and our current understanding is mostly related to pathogenic species such as Legionella pneumophila and Mycobacterium avium but not to bacterial species closely related to them. In this study, genomic-based approaches were used to characterize pathogen-related species in relation to their abundance, diversity, potential pathogenicity, genetic exchange, and distribution across an urban drinking water system. Nine draft genomes recovered from 10 metagenomes were identified as Legionella (4 draft genomes), Mycobacterium (3 draft genomes), Parachlamydia (1 draft genome), and Leptospira (1 draft genome). The pathogenicity potential of these genomes was examined by the presence/absence of virulence machinery, including genes belonging to Type III, IV, and VII secretion systems and their effectors. Several virulence factors known to pathogenic species were detected with these retrieved draft genomes except the Leptospira-related genome. Identical clustered regularly interspaced short palindromic repeats-CRISPR-associated proteins (CRISPR-Cas) genetic signatures were observed in two draft genomes recovered at different stages of the studied system, suggesting that the spacers in CRISPR-Cas could potentially be used as a biomarker in the monitoring of Legionella related strains at an evolutionary scale of several years across different drinking water production and distribution systems. Overall, metagenomics approach was an effective and complementary tool of culturing techniques to gain insights into the pathogenic characteristics and the CRISPR-Cas signatures of pathogen-related species in DWDSs.
Highlights
Over 500 waterborne or water-based pathogens of potential concern in drinking water (e.g., Legionella pneumophila, Escherichia coli O157:H7, Mycobacterium avium, and Cryptosporidium parvum) have been included in the Candidate Contaminant List by the US Environmental Protection Agency (EPA; Ashbolt, 2015)
We investigated the possible targets of spacers in CRISPR-Cas arrays within the obtained draft genomes using CRISPRTarget to search against all the available databases (i.e., GenBank-Phage, GenBankEnvironmental, RefSeq-Plasmid, RefSeq-Viral, and RefSeqBacteria), which was combined with the known features of each subtype that had been reported to be essential for target recognition, such as protospacer adjacent motifs (PAMs) and seed regions (Biswas et al, 2013)
The before filtration and chlorination (BC) sample contained the highest level of Legionella spp.: a relative abundance of 4.7 × 10−3 based on 16S rRNA amplicon analysis and a concentration of 40.9 copies/ng-Genomic DNA (gDNA) by digital droplet PCR (ddPCR)
Summary
Over 500 waterborne or water-based pathogens of potential concern in drinking water (e.g., Legionella pneumophila, Escherichia coli O157:H7, Mycobacterium avium, and Cryptosporidium parvum) have been included in the Candidate Contaminant List by the US Environmental Protection Agency (EPA; Ashbolt, 2015). Some secretion systems are dedicated for bacteria-host interaction, such as the type III secretion system (T3SS) in Chlamydia (BettsHampikian and Fields, 2010), the type IVB secretion system (T4BSS, Dot/Icm) in Lg. pneumophila (Voth et al, 2012), and the type VII secretion system (T7SS) in Mycobacterium (Costa et al, 2015). The deletion of these secretion systems could result in a substantial decrease in virulence (Costa et al, 2015). Several other VFs have been reported for pathogens including those facilitating attachment and invasion (e.g., cell wall, type IV pili) and endotoxins
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