b F-specific RNA bacteriophages (FRNAPH) have been widely studied as tools for evaluating fecal or viral pollution in water. It has also been proposed that they can be used to differentiate human from animal fecal contamination. While FRNAPH subgroup I (FRNAPH-I) and FRNAPH-IV are often associated with animal pollution, FRNAPH-II and -III prevail in human wastewater. However, this distribution is not absolute, and variable survival rates in these subgroups lead to misinterpretation of the original distribution. In this context, we studied FRNAPH distribution in urban wastewater and animal feces/wastewater. To increase the specificity, we partially sequenced the genomes of phages of urban and animal origins. The persistence of the genomes and infectivity were also studied, over time in wastewater and during treatment, for each subgroup. FRNAPH-I genome sequences did not show any specific urban or animal clusters to allow development of molecular tools for differentiation. They were the most resistant and as such may be used as fecal or viral indicators. FRNAPH-II's low prevalence and low sequence variability in animal stools, combined with specific clusters formed by urban strains, allowed differentiation between urban and animal pollution by using a specific reverse transcription-PCR (RT-PCR) method. The subgroup's resistance over time was comparable to that of FRNAPH-I, but its surface properties allowed higher elimination rates during activated-sludge treatment. FRNAPH-III's low sequence variability in animal wastewater and specific cluster formation by urban strains also allowed differentiation by using a specific RT-PCR method. Nevertheless, its low resistance restricted it to being used only for recent urban pollution detection. FRNAPH-IV was too rare to be used. Water contamination by pathogens is a major public health problem. Microorganisms affecting water quality are mainly excreted in feces, and water contamination is caused by discharge of fecal waste of animal origin (e.g., from runoff after rain, farms, and slaughterhouses [SH]) or human origin (e.g., from raw or treated urban wastewater) into the environment. Enteric pathogens include bacteria, protozoa, and enteric viruses (1). The last have been identified in many cases of waterborne and food-borne diseases in developed countries (2).The virological quality of water is usually indirectly estimated from the number of fecal indicator bacteria, such as Escherichia coli and enterococci (3). However, these indicators have serious limitations, especially for the estimation of viral pollution. First, it has been largely demonstrated that viruses are more resistant than bacteria in the environment; therefore viral outbreaks may still be identified even in the absence of indicator bacteria (4, 5). Second, conventional indicators are present in human and animal feces, and they do not provide information about the origin of fecal contamination (6). Yet, this parameter is essential because the reservoir of many waterborne pathogens, especially viruses, is hu...
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