The discharge of improperly treated wastewater can contaminate receiving waterbodies. Within these waterbodies, exposure to waters contaminated with human waste can endanger public health due to the potential presence of enteric viruses, which are a leading cause of waterborne disease outbreaks (Farkas et al., 2020, McMinn et al., 2017b). Historically, water quality assessments have relied on cultivating fecal indicator bacteria (FIB) enterococci and Escherichia coli. However, due to large morphological differences, FIB are poor predictors of the presence and persistence of other taxonomic groups such as enteric viruses (Korajkic et al., 2018, Korajkic et al., 2019). Enteric viruses (i.e., viruses that primarily infect the gastrointestinal tract) are commonly detected in wastewater, and are a predominant human health risk since viruses can persist in environments outside of their host, and ingesting only a few infectious virions can cause illness (Farkas et al., 2020, Korajkic et al., 2019).

Enteric viruses are typically found in low concentrations in environmental waters necessitating the use of filters to concentrate virus in order to acquire detectable densities for downstream assays such as culture and qPCR (Cashdollar et al., 2013a, Gibson, 2014, Korajkic et al., 2021, McMinn et al., 2017a, McMinn et al., 2017b). Historically the use of primary filters such as adsorption-elution (electropositive and electronegative), or secondary concentration procedures such as precipitation and centrifugation (polyethylene glycol, skimmed milk, centrifugal ultrafilters, etc.) have been used for viral concentration from liquids (Cashdollar and Wymer, 2013b). However, these approaches can be time consuming or are unable to process sufficient sample volumes (McMinn et al., 2012). Recently, hollow fiber ultrafilters (HFUF) used in medical dialysis have been successfully repurposed to concentrate viruses from environmental waters and wastewater (Hurst et al., 2023, Kelleher et al., 2025, Korajkic et al., 2022, Korajkic et al., 2021, McMinn et al., 2017b, McMinn et al., 2019, McMinn et al., 2021). HFUFs incorporate large filtration surface areas making them less prone to fouling in large-volume turbid matrices, improving our ability to reliably measure rarer viral targets (McMinn et al., 2017b). HFUF technology incorporates membranes that sieve water based on particle size, making this approach superior to others (adsorption or precipitation) that are dependent on source water chemistry, as well as viral capsid surface charge, to concentrate viruses present (Liu et al., 2012a). To date, HFUF have been used successfully for concentrating a wide range of microorganisms including viruses from a variety of challenging environmental matrices, including wastewater (Korajkic et al., 2022, Mull and Hill, 2012).

Recently, viral indicators such as somatic and F+ coliphage have been proposed as suitable proxies for the presence of enteric viruses in environmental waters and wastewater (McMinn et al., 2018, Singh et al., 2022). Coliphage are viruses that infect E. coli and are shed in waste through similar processes to those of enteric viral pathogens (King et al., 2011). A recent review suggested that coliphage have comparable fate and transport characteristics to enteric viral pathogens and can act as a more suitable indicator for pathogenic virus presence in recreational waters than FIB (Korajkic et al., 2019). Detection of coliphage rely on an easy-to-use culture-based assays using E. coli host monolayers for visualizing and enumerating lysogenic coliphage present. For these reasons, the United States Environmental Protection Agency (USEPA) has developed methods specific for coliphage detection and enumeration (United States Environmental Protection Agency, 2001a, United States Environmental Protection Agency, 2001b, United States Environmental Protection Agency, 2018b).

While coliphage are found at higher concentrations than pathogenic enteric viruses, they are typically found at lower concentrations than FIB, which potentially impacts their ability to be used during routine monitoring of fecal pollution in recreational waters (McMinn et al., 2017b). To increase the ability to detect endogenous somatic and F+ coliphage from ambient waters and wastewater, the USEPA created Method 1642, which details the use of the 15S Asahi Kasei Rexeed HFUF filter in a dead-end setup (D-HFUF) for concentrating larger volume of samples (2 L) of ambient waters and disinfected wastewater effluents. Due to manufacturing changes, the Asahi Kasei Rexeed filter is no longer available for purchase in US markets. This created an issue for laboratories when implementing US EPA Method 1642 for coliphage monitoring, so a suitable HFUF replacement filter is needed. In addition to the unavailability of a suitable replacement filter, coliphage specific methods (US EPA Methods 1602, 1642, and 1643) do not specify recommended sample holding times (United States Environmental Protection Agency, 2001b, United States Environmental Protection Agency, 2018a, United States Environmental Protection Agency, 2018b). While re-growth of coliphage in samples is less of a concern than for bacterial targets (United States Environmental Protection Agency, 2009, United States Environmental Protection Agency, 2023), assessments to establish holding times are still critical as viral viability can degrade overtime due to the impacts of biological and chemical residuals common to environmental waters and wastewaters (Jofre et al., 2016). To accurately assess recreational water quality, the establishment of acceptable sample holding times is needed to assure data collected is representative of the contamination levels present at the time of sampling

For this study, we identified HFUF options currently available for purchase in the US that were of similar price, function, and construction materials to that of the Asahi Kasei Rexeed filters. Through this process, two similar HFUF filters were identified for testing; the Fresenius F160NRE and the Elisio-15H. The goals of this study were to 1) compare the various filter options for their ability to recover (endogenous and spiked) somatic and F+ coliphages from freshwater, marine water, and wastewater matrices, and 2) identify acceptable sample holding times in the different water matrices by measuring endogenous coliphage levels over time.