Occurrence And Human Infection Potential Of Pathogenic Pathogens In Urban Wastewater Treatment Plant Effluents In Shanghai, China

Cryptosporidium spp., Giardia duodenalis and Enterocytozoon bieneusi are important causes of diarrhea in humans. Due to a fresh water shortage worldwide, reclaimed water from wastewater treatment plant (WWTP) effluents has become a potential water supply for irrigation and recreation. However, in the standard for WWTP effluents in China, the only microbiological indicator is fecal coliforms without monitoring of pathogenic microorganisms, raising concerns about the potential for pathogen transmission through WWTP effluent reuse.To fully investigate the occurrence and the public infection potential of Cryptosporidium spp., G. duodenalis and E. bieneusi in WWTP effluents in Shanghai, China,50 samples were collected and analyzed. All samples were pre-treated by calcium carbonate flocculation, followed by immunofluorescence microscopy and nested PCR for the examination of Cryptosporidium (at small-subuit ribosomal RNA loci) and G. duodenalis (at triose phosphate isomerase, glutamate dehydrogenase and β-giardia loci). Moreover, propidium monoazide (PMA)-PCR was used to assess the viability of oocysts and cysts. Only nested PCR was used in the detection of E. bieneusi (at internal transcribed spacer loci). Furthermore, a 10 mL aliquot of effluent was mixed and used for the determination of fecal coliform by the Colilert Quanti-Tray/2000(?) before pretreatment.The microscopy and PCR-positive rates for Cryptosporidium spp. were 62% and 40%, respectively. The occurrence rates of G. duodenalis were 96% by microscopy and 92-100% by PCR analysis of three genetic loci. Furthermore, E. bieneusi was detected in 70% of samples by PCR. Altogether, ten Cryptosporidium species or genotypes, two G. duodenalis genotypes, and eleven E. bieneusi genotypes were found. Most of the species or genotypes of the pathogens identified were human-pathogenic. Turbidity and pH were quite stable in all the samples. Approximately 93% and 83% of the samples from WWTP1 and WWTP3 failed to meet the Chinese national standard on fecal coliform levels. There were no significant differences in pathogen detection rates between PCR and PMA-PCR. Also we cannot find a consistant relationship between the occurrence of fecal coliform and other three pathogens in this study. The results suggest that urban WWTP effluents in China often contain residual human enteric pathogens and fecal coliform cannot be used to indicate the occurance of the remaining Cryptosporidium spp., G. duodenalis or E. bieneusi in WWTP effluents.Data from this study should be useful for tracking the source of microorganisms in case of outbreaks caused by wastewater discharage and provided basis for improving the standard for urban WWTP efflents.