A small, well-defined watershed was investigated over a 2-year period to determine the prevalence of F-specific RNA coliphage (F + RNA) serotypes as indicators of animal fecal contamination. Sampling sites collected runoff from areas of urban and agricultural land use patterns. F-specific coliphages were concentrated from 2-L freshwater samples by polyethylene glycol precipitation, isolated using the double agar layer (DAL) method, confirmed as F + RNA by RNAse suppression, and serotyped. A subset of serotyped F + RNA were confirmed by genotyping. To determine relative survival, 10 confirmed F + RNA field isolates and 5 prototypic F + RNA were spiked into surface water and incubated at 25 degrees C for 36 days. F-specific coliphage isolation was strongly associated with rainfall events and was infrequent from primarily animal impacted surface waters. Field isolates were predoffiinantly Type I F + RNA (81%) and raw sewage isolates were predominantly Type III F + RNA (57%). Genotyping from either the watershed or raw sewage samples never positively identified Type IV F + RNA. Results from laboratory studies showed that F + RNA differ in their survival in water and that Type IV strains were the least persistent. Type III F + RNA were found to be reliably related to the release of uncontrolled human fecal material in the watershed, but the results of this study suggest that further study is required before utilizing for fecal source identification in natural waters.