BACKGROUND: We aimed to identify the genes of 35 pathogens, including viruses, bacteria, and protozoa that cause waterborne infectious diseases, and to assess the feasibility of a wastewater-based surveillance system. MATERIALS AND METHODS: Wastewater was aseptically sampled once a month from 2 sites. A total of 1 L of wastewater from each site underwent 0.2 microm filtration to generate the sample A. Subsequently, 200 ul of the filtered water was ultra-filtered and concentrated to generate the sample B, which was mixed with sample A in a 1:1 ratio. We performed a Filmarray((R)) Gastrointestinal (GI) panel (BioFire Diagnnostics\', Salt Lake City, UT, USA) test to simultaneously detect 13 enterobacteria, 5 enterovirus, and 4 protozoa. RNA was extracted to assess 18 types of viruses. RESULTS: Severe acute respiratory syndrome coronavirus 2 adenovirus, bocavirus, and rhinovirus was detected at both site. Norovirus GI/GII was continuously detected at both sites. Moreover, adenovirus, group A rotaviruses, and hepatitis A virus were frequently detected; however, hepatitis E virus was absent at either site. Campylobacter, enteroaggregative Escherichia coli, enterotoxigenic E. coli, Shiga toxin-producing E. coli, and Clostridioides difficile toxin A/B were detected at both sites. Giardia lamblia was also detected in both sites. CONCLUSION: We analyze multiple infectious disease pathogens under sample surveillance with incidence. Further in-depth studies on wastewater-based surveillance will be feasible and important.