High-Throughput Microfluidic Quantitative PCR Platform for the Simultaneous Quantification of Pathogens, Fecal Indicator Bacteria, and Microbial Source Tracking Markers

Contamination of water with bacterial, viral, and protozoanpathogenscan cause human diseases. Both humans and nonhumans can release thesepathogens through their feces. To identify the sources of fecal contaminationin the water environment, microbial source tracking (MST) approacheshave been developed; however, the relationship between MST markersand pathogens is still not well understood most likely due to thelack of comprehensive datasets of pathogens and MST marker concentrations.In this study, we developed a novel microfluidic quantitative PCR(MFQPCR) platform for the simultaneous quantification of 37 previouslyvalidated MST markers, two fecal indicator bacteria (FIB), 22 bacterial,11 viral, and five protozoan pathogens, and three internal amplification/processcontrols in many samples. The MFQPCR chip was applied to analyze pathogenremoval rates during the wastewater treatment processes. In addition,multiple host-specific MST markers, FIB, and pathogens were successfullyquantified in human and avian-impacted surface waters. While the genesfor pathogens were relatively infrequently detected, positive correlationswere observed between some potential pathogens such as Clostridium perfringens and Mycobacterium spp., and human MST markers. The MFQPCR chips developed in thisstudy, therefore, can provide useful information to monitor and improvewater quality. A high-throughput tool wasdeveloped to quantify variouspathogens and microbial source tracking markers to monitor and improvewater quality.