Detecting Human Bacterial Pathogens in Wastewater Treatment Plants by a High-Throughput Shotgun Sequencing Technique

Abstract

Human pathogens are one of the major threats to global public health. Wastewater treatment plants (WWTPs) serve as city guts to receive and digest various human pathogens. Several techniques have been developed to detect human pathogens in WWTPs and to assess potential environmental risks. In this study, we employed 24 metagenomic DNA data sets derived from a high-throughput shotgun sequencing technique to more accurately and efficiently detect human bacterial pathogens in influent, activated sludge, and effluent of two Hong Kong WWTPs. Each data set was quality-filtered and normalized to 12,500,000 DNA sequences with a length of 150-190 bp. Then, a BLASTN search against Greengenes general 16S rRNA gene database and human pathogenic bacteria 16S rRNA gene database, a BLASTX search against human pathogenic bacteria virulence factor database, as well as MetaPhlAn analysis were conducted to survey the distribution, diversity, and abundance of human bacterial pathogens. The results revealed that (i) nine bacterial pathogens were detected; (ii) the overall pathogenic bacteria abundance was estimated as 0.06-3.20% in the total bacteria population using 16S rRNA gene fingerprinting; (iii) pathogenic bacteria detected in activated sludge and effluent shared similar profiles but were different from influent based on both 16S rRNA gene and virulence factor fingerprintings; (iv) Mycobacterium tuberculosis -like species may present potential pathogenic risks because it was detected with high abundance in both activated sludge and effluent. These findings provided a comprehensive profile of commonly concerned human pathogens in two Hong Kong WWTPs and demonstrated that the high-throughput shotgun sequencing technique is a feasible and effectual approach for environmental detection of human bacterial pathogens.

Metagenomic analysis of bacterial community composition and antibiotic resistance genes in a wastewater treatment plant and its receiving surface water

“…Compared to activated sludge, more pathogens were detected in both influent and effluent, which was also reported in previous study, and different sample processing in pretreatment (i.e. filtration or centrifugation) might be one of the major reasons causing this result (Cai and Zhang, 2013). Although, most pathogens were eliminated in the WWTP, the presence of the opportunistic pathogens in the effluent still should be concerned.…”

“…However, the incomplete treatment of recalcitrant bacteria, especially some pathogens and bacteria carrying antibiotic resistance genes (ARGs) will put the receiving surface water at risk (Kumaraswamy et al, 2014;Marti et al, 2013). Owing to their high virulence and rapid airborne or water transmission, bacterial pathogens are disastrous and detrimental for human health (Cai and Zhang, 2013;Ahmed et al, 2014;Gomi et al, 2015). On the other hand, due to the overuse and misuse of antibiotics, antibiotic resistance has represented a significant global health problem, causing ARGs as new emerging pollutants (Port et al, 2012).…”

Effect of lake water on algal biomass and microbial community structure in municipal wastewater-based lab-scale photobioreactors

“…Twenty-five of the 35 pathogens examined by Cai and Zhang (2013) were found in the wastewater used in both TWR and LWR. The original list consisted of bacteria that possessed one or more human virulence factors.…”

Anaerobic Membrane Bioreactor Effluent Reuse: A Review of Microbial Safety Concerns

“…These solutions have included monoazide-based staining of DNA to differentiate cells with intact cell walls against those with compromised ones. Furthermore, the merits of molecular detection of pathogenic bacteria are well established and can offer specific insight into various wastewater treatment process strengths for increasing microbial safety [90][91][92][93][94]. These merits include the quantification of non-cultivable pathogens by quantitative PCR and semi-comprehensive comparative analysis of potentially pathogenic bacterial abundance by high-throughput sequencing.…”