Human and animal fecal pollution of the environment presents a risk to human health because of the presence of pathogenic viruses and bacteria. To distinguish between human and animal sources of pollution, we designed specific real-time reverse transcription (RT)-PCR assays for human and animal enteric viruses, including norovirus genogroups I, II, and III; porcine adenovirus types 3 and 5; ovine adenovirus; atadenovirus; and human adenovirus species C and F, which are excreted by infected humans, pigs, cattle, sheep, deer, and goats, and for the detection of F؉ RNA bacteriophage genogroups I to IV, which are associated with human and animal wastes. The sensitivity of this viral toolbox (VTB) was tested against 10-fold dilution series of DNA plasmids that carry the target sequences of the respective viruses and was shown to detect at least 10 plasmid copies for each assay. A panel of human and animal enteric and respiratory viruses showed these assays to be highly sensitive and specific to their respective targets. The VTB was used to detect viruses in fecal and environmental samples, including raw sewage and biosolids from municipal sewage treatment plants, abattoir sewage, and fecally contaminated shellfish and river water, which were likely to contain animal or human viruses.It is important that sources of fecal pollution are rapidly and accurately identified so that environmental managers can develop strategies to eliminate the source of the pollution in an efficient and cost-effective manner. The identification of fecal sources also allows the potential risk to human health following the consumption of or exposure to contaminated food or water to be estimated. For this purpose, numerous fecal source tracking methods have been developed. Among these are library-dependent methods such as antibiotic resistance testing or molecular fingerprinting (e.g., terminal restriction length fragment polymorphism [T-RFLP] or ribotyping) and library-independent approaches, including the detection of human-specific bacteriophages by plaque assay (e.g., Bacteroides fragilis bacteriophages), the detection of certain chemicals associated with human and nonhuman wastes (e.g., caffeine, laundry brighteners, fecal sterols, and stanols), and the detection of specific genetic markers by PCR. The molecular detection of human and animal viruses and bacteriophages falls into the latter category. Each method has advantages and disadvantages depending on the circumstances and the questions at hand. To date, none has been proposed as a standard method (9,34,40,45).The detection and genotyping of Fϩ RNA bacteriophages have been widely used as a tool for microbial source tracking (MST) (5,8,17,18,31,44). Fϩ RNA bacteriophage genogroup I (GI) and GIV have been associated with animal contamination, and genogroups II and III indicate predominantly human fecal contamination or domestic sewage-associated in-
