is not synonymous with the presence of an infectious unit (IU). Dose-response studies were conducted using tissue culture assays for quantification of IU. There is limited quantitative information on the side-by-side data for IUs and genome copies although it is generally known that infectivity decays more rapidly than does the density of genome copies (R.A. Rodriguez et al., 2009). Based on a single report (He and Jiang, 2005), where three side-by-side polymerase chain reaction (PCR) and tissue culture assays were performed on adenovirus isolated from secondary effluent, it is estimated that the ratio between genome copies and infectious units is approximately 1,000:1. Thus, genome count densities estimated for adenovirus for each scenario were reduced by three orders of magnitude to convert to IUs during the risk estimation process.
Norovirus is one of the most important enteric viruses for both waterborne and foodborne outbreaks in the United States. Several recent studies have focused on occurrence of this virus in water and wastewater (Pusch et al., 2005; Haramoto et al., 2006; Katayama et al., 2008; Nordgren et al., 2009; Victoria et al., 2010). In these studies, the density of the norovirus genome varies over a wide range with densities as high as 107 gc/L reported in raw sewage. Based on the published literature, a density of 104 gc/L is estimated to be the median occurrence in secondary effluent. Once again, although the genome-based method is sensitive at detecting the presence of copies of the genome of the virus, it does not provide information on viral infectivity. Norovirus has not been successfully cultivated using conventional tissue culture methods, and so no work is available to establish the ratio between genome density and IU density.
A dose-response model for norovirus was used based on the study by Teunis et al. (2008), using the estimate for single unaggregated virus. Because norovirus has not been successfully cultivated in vitro, these studies were conducted using fresh virus and the genome count quantified by PCR. Published work has shown that the fraction of genome copies that are infectious drops rapidly in the environment (R.A. Rodriguez et al., 2009). Thus, for the purposes of this exemplar, the same 1,000:1 was applied before risk estimation.
Salmonella has long been a well-studied waterborne enteric pathogen. The concentration of this microorganism in raw sewage ranges between 102 and 104 cfu/100 mL (Asano et al., 2007). Taking the average of these two and assuming the same 2-log reduction during primary and secondary treatment that normally occurs for Escherichia coli produces an estimate of 5 × 102 cfu/L in secondary effluent for the exemplar. Again, the dose-response model for this organism has been developed previously based on epidemiological studies (Haas et al., 1999).
Cryptosporidium is associated with both drinking water and recreational water outbreaks in the United States. The health significance of this organism has motivated a number of studies to understand its occurrence and persistence in the water environment (Rose et al., 1996; Gennaccaro et al., 2003; Robertson et al., 2006; Lim et al., 2007; Castro-Hermida et al., 2008; Chalmers et al., 2010; Fu et al., 2010). The peer reviewed literature reports a range of Cryptosporidium densities in secondary treated effluents varying with season and geographical location. Studying this literature, a density of 50 oocysts/L is estimated as typical for secondary effluents. However, most of the data on oocyst concentration were determined using the indirect fluorescent-antibody assay (IFA), which also does not directly measure IUs. A study comparing oocyst densities as determined by IFA with IU densities as determined by a focus-detection-method most-probable-number technique in cell culture (Slifko et al., 1999) found a ratio of approximately 3:1 in 18 samples of secondary effluent (Gennaccaro et al., 2003). Using this ratio, a density of 50 oocysts/L produces an estimate of 17 IUs/L in secondary effluent for the exemplar. More than one dose-response model has been developed for this organism (Haas et al., 1999).
Assumptions Concerning Fate, Transport, and Removal
The following is a brief discussion of assumptions made regarding fate, transport, and removal for the pathogens in the exemplar.