Combined application of metagenomics and FEAST to trace sources of microbial eukaryotic contamination in the Pasig–Marikina–San Juan (PAMARISAN) river system in Metro Manila, Philippines

Microbial eukaryotes are vital to global microbial diversity, but there is limited information about their composition and sources in contaminated surface waters. This study examined the pathogens and potential sources of microbial eukaryotic communities in polluted sink environments using the 18S rDNA amplicon sequencing combined with the fast expectation–maximization for microbial source tracking (FEAST) program. Six sampling sites were selected along the Pasig–Marikina–San Juan (PAMARISAN) River System, representing different locations within the waterway and classified as sinks (n = 12), whereas animal fecal samples collected from various farms were classified as sources (n = 29). Taxonomic composition revealed Stramenopila, Alveolata, Rhizaria (SAR), Archaeplastida, and Excavata in the rivers, accounting for 85.1%, 13.2%, and 0.36% mean abundance of microbial sink communities, respectively. Clinically relevant human pathogens were also observed in sink environments. The correlation test demonstrated that dissolved oxygen, total suspended solids, pH, temperature, fecal coliform count, and phosphates were important environmental factors driving community variations. Moreover, FEAST results indicated that sewage (19.6%) was the primary source of microbial eukaryotes, followed by duck (0.644%) and cow (0.566%) feces. Spatio-seasonal variations showed higher contributions at downstream stations and during the wet season, highlighting the role of rainfall in enhancing microbial dispersal. Results from community-based microbial source tracking can be used to explore factors shaping microbial eukaryotes in freshwater environments, assess potential pathogen-related hazards, and inform river conservation and management strategies. Furthermore, this also serves as preliminary data for microbial eukaryotic source tracking in the Philippines, laying groundwork for future research.