Human infective potential of Cryptosporidium spp., Giardia duodenalis and Enterocytozoon bieneusi in urban wastewater treatment plant effluents

Cryptosporidiosis, giardiasis, and microsporidiosis are important waterborne diseases. In the standard for wastewater treatment plant (WWTP) effluents in China and other countries, the fecal coliform count is the only microbial indicator, raising concerns about the potential for pathogen transmission through WWTP effluent reuse. In this study, we collected 50 effluent samples (30 L/sample) from three municipal WWTPs in Shanghai, China, and analyzed for Cryptosporidium spp., Giardia duodenalis and Enterocytozoon bieneusi by microscopy and/or polymerase chain reaction (PCR). Moreover, propidium monoazide (PMA)-PCR was used to assess the viability of oocysts/cysts. 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% (35/50) of samples by PCR. Altogether, 10 Cryptosporidium species or genotypes, two G. duodenalis genotypes, and 11 E. bieneusi genotypes were found, most of which were human-pathogenic. The chlorine dioxide disinfection employed in WWTP1 and WWTP3 failed to inactivate the residual pathogens; 93% of the samples from WWTP1 and 83% from WWTP3 did not meet the national standard on fecal coliform levels. Thus, urban WWTP effluents often contain residual waterborne human pathogens.