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Sewer overflows and exfiltration can potentially contaminate water bodies with pathogens from wastewater. Microbial source tracking (MST) methods such as the detection of the HF183 gene target of  Bacteroides  have been proposed to monitor human fecal pollution inputs to surface waters; however, the persistence of HF183 and other MST markers in water flushed from soils after contamination events is not well understood. In this study, the persistence and decay of two culture-based fecal indicators,  Escherichia coli  and enterococci, and two molecular MST markers, HF183 and pepper mild mottle virus (PMMoV), were evaluated in riverbank soils spiked with untreated sewage, which were left idle for 1, 14, 28, 60, and 121 days under dark conditions and then flushed with synthetic rainwater. All four microbial indicators were still detected in flush water 4 months after the soil was contaminated. PMMoV persisted much longer and had a slower decay rate than the other microbial indicators, and  E. coli  degraded most rapidly. In consecutive flushing experiments with fresh (1 day) sewage-spiked soils, HF183,  E. coli , and enterococci were all detected after 20 consecutive flushes with rainwater, but PMMoV was not detected after the fifth flush. Our findings indicate that water (e.g., stormwater interflow) flushing through riverbank soils that have previously been contaminated by sewer overflows or sewer exfiltration can potentially be a source of microbial pollution to surface waters, even for several months after the contamination occurs. Results from this study also demonstrate the benefits of using multiple human-associated fecal indicators to distinguish pollution from different microbial groups in water bodies. Graphical abstract

Persistence of Fecal Indicators and Microbial Source Tracking Markers in Water Flushed from Riverbank Soils