Selective Detection of Viable Pathogens in Wastewater Treatment Plant by Quantitative PCR Combined with Propidium Monoazide

摘要

@@Introduction Wastewater treatment plants (WWTP) collect wastewater containing varieties of pathogens from different sources. The presence and proliferate of these pathogens may be a great potential threat to public health,indicating the importance of control and monitoring of pathogens in WWTP. Traditional methods of pathogen detection are culture-dependent,but they are labor-intensive, time-consuming and difficult.More importantly,pathogens may enter a viable but non-culturable (VBNC) state,which makes culture-based methods largely underestimate the number of pathogen cells.DNA-based methods,such as polymerase chain reaction (PCR) and microarray,can be good alternatives with high specificity,speed,and sensitivity.Due to the persistence of DNA after cell lost its viability,DNA-based methods may cause false-positives or overestimate the number of infectious target pathogens.Recently,this problem can be solved by treating bacterial samples with propidium monoazide (PMA),which can intercalate with DNA from non-viable cells,prior to DNA extraction.However,validation of this method in practically environmental samples needs to be further investigated.In this study,we analyzed the effectiveness of PMA in real samples in WWTP and applied quantitative PCR combined with PMA (PMA-qPCR) to detect the amount of two typical pathogens,E coli and Salmonella spp.,in WWTP.