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In this paper we present a quantitative sanitary survey of the Middle Santa Ana River, in southern California, utilizing a variety of source tracking tools, including traditional culture-dependent fecal markers (Enterococcus and Escherichia coli by IDEXX), speciation of enterococci isolates, culture-independent fecal markers (human-specific HF183 Bacteroides and Enterococcus by quantitative polymerase chain reaction, qPCR), and chemical markers of sewage and wastewater (nutrients, enantiomeric fraction (EF) of propranolol and ethylenediaminetetraacetic acid). To facilitate comparison of these different methods, data are interpreted in a loading (i.e., mass per time) framework that enables a quantitative apportionment of fecal markers and nutrients to specific source waters in the Middle Santa Ana River. Multiple lines of evidence support the hypothesis that Enterococcus in the Middle Santa Ana River originates primarily from in situ growth in streambed sediments, not from significant and persistent sources of untreated human waste. The EF of propranolol of tertiary treated wastewater effluent is in the range (0.42 to 0.71) previously reported for raw sewage, making EF of propranolol an unsuitable marker for fecal pollution, at least at this site. The human fecal marker HF183 Bacteroides was detected at a few sites, although not in a source of disinfected and tertiary treated wastewater effluent Based on the results presented here and prior experience at other sites in southern California, HF183 Bacteroides would appear to be a candidate marker of fecal contamination for inland waters, although more qPCR measurements in disinfected wastewater effluent are needed to account for variations due to treatment plant performance and other factors. More generally, our results support the notion that regrowth of fecal indicator bacteria (FIB) in river sediments may lead to a decoupling between FIB and pathogen concentrations in the water column and thus limit the utility of FIB as an indicator of recreational waterborne illness in inland waters. [ABSTRACT FROM AUTHOR]