New Electropositive Filter for Concentrating Enteroviruses and Noroviruses from Large Volumes of Water

The U.S. Environmental Protection Agency''s information collection rule requires the use of 1MDS electropositive filters for concentrating enteric viruses from water, but unfortunately, these filters are not cost-effective for routine viral monitoring. In this study, an inexpensive electropositive cartridge filter, the NanoCeram filter, was evaluated for its ability to concentrate enteroviruses and noroviruses from large volumes of water. Seeded viruses were concentrated using the adsorption-elution procedure. The mean percent retention of seeded polioviruses by NanoCeram filters was 84%. To optimize the elution procedure, six protocols, each comprising two successive elutions with various lengths of filter immersion, were evaluated. The highest virus recovery (77%) was obtained by immersing the filters in beef extract for 1 minute during the first elution and for 15 min during the second elution. The recovery efficiencies of poliovirus, coxsackievirus B5, and echovirus 7 from 100-liter samples of seeded tap water were 54%, 27%, and 32%, respectively. There was no significant difference in virus recovery from tap water with a pH range of 6 to 9.5 and a water flow rate range of 5.5 liters/min to 20 liters/min. Finally, poliovirus and Norwalk virus recoveries by NanoCeram filters were compared to those by 1MDS filters, using tap water and Ohio River water. Poliovirus and Norwalk virus recoveries by NanoCeram filters from tap and river water were similar to or higher than those by the 1MDS filters. These data suggest that NanoCeram filters can be used as an inexpensive alternative to 1MDS filters for routine viral monitoring of water.Viruses that primarily infect and replicate in the gastrointestinal tract are known as enteric viruses. More than 140 different enteric viruses are known to infect humans. These include the enteroviruses, rotaviruses, hepatitis A virus, noroviruses, adenoviruses, and reoviruses, among others. Enteric viruses are capable of causing a wide range of illnesses, including gastroenteritis, paralysis, aseptic meningitis, herpangina, respiratory illness, fevers, myocarditis, etc. Given the potential public health impact of the enteric viruses, enteroviruses (echovirus and coxsackievirus), adenoviruses, and caliciviruses are on the U.S. Environmental Protection Agency''s contaminant candidate list 2 for regulatory consideration for drinking water (11). Within the Caliciviridae family, noroviruses are the primary viruses of concern for drinking water.Contaminated drinking water is considered to be a potential transmission route, and an infectious dose in humans may consist of only a small number of virus particles. Enteric viruses are introduced in aquatic environments through natural or human activities, such as leaking sewage and septic systems, urban runoff, landfills, injection of treated wastewater into aquifers, wastewater discharge, sewage outfall, etc. These viruses have been found in surface water, groundwater, and drinking water (1, 6, 13, 22, 26). Between 1971 and 2004, 789 drinking water outbreaks and 575,207 cases of illness were reported in the United States, and 8% of the reported outbreaks were due to enteric viruses (2, 5, 28, 29, 30, 46).The levels of enteric viruses in natural waters are often low, and as such, typical virus sampling involves a primary concentration of viruses from large volumes of water (hundreds to thousands of liters). Unlike other waterborne pathogens (such as bacteria and parasites), viruses are smaller, and thus, size exclusion filtration is often not practical, especially for turbid waters. In addition, viruses are negatively charged in natural environments and can be adsorbed onto a number of different matrices by electrostatic and hydrophobic interactions (16). Consequently, different types of matrices have been used to isolate enteric viruses from water. These include negatively and positively charged membranes or cartridge filters (10, 17, 32, 34, 35, 39), gauze pad (31), and glass powder or glass wool (14, 27). Of all of these methods, electronegative and electropositive filters are most commonly used. In the case of electronegative filters, the acidification of the water and addition of multivalent cations are required for optimal virus adsorption. Because of this need to condition the water to attain acceptable recoveries, it is difficult to use electronegative filters for field sampling. In contrast, electropositive filters do not require conditioning of the water. Among all the filters, 1MDS electropositive filters (Cuno, Meriden, CT) are the most commonly used filter for fresh and drinking water sampling; however, they are not cost-effective for routine viral monitoring of water and require pH adjustment for waters with pH values exceeding 8.0 (12).Viruses adsorbed on the filter are usually eluted and recovered using 1 to 1.6 liters of eluting solution (6, 12). Many different procedures are described in the literature to elute viruses from filters. These procedures include the use of different eluting solutions, such as 0.3%, 1.5% or 3% beef extract, urea-arginine phosphate buffer, glycine buffer, etc. (10, 12, 24, 37). There are also different elution processes, such as single elution, recirculation of eluents, or successive elution of filters (6, 8, 15, 43). Sobsey and Hickey (40) used only one elution with 0.3% beef extract in 50 mM glycine. Sobsey et al. (43) suggested that 1 liter of 1.5% beef extract be recirculated through the filters for 5 min. Dahling and Wright (8) reported that the highest virus recoveries were obtained by three elutions, each using 1.6 liters of 3% beef extract. Dahling (6) reported that the highest virus recoveries were obtained with two separate beef extract elutions, one being an overnight filter immersion in beef extract.Although methods for concentration of many enteric viruses have been developed, limited studies have been conducted for concentrating noroviruses from water. Huang et al. (21) described a norovirus concentration method using porcine calicivirus (Pan-1) as a surrogate. Pan-1 was sensitive to the high pH (9.5) of the eluting solution, which is commonly used. Myrmel et al. (33) described a method of norovirus concentration using feline calicivirus as a surrogate organism. The method used electronegative filters, and the recovery of virus was 5 to 10%. Many other studies reported detection of human noroviruses in environmental waters (18, 19, 25); however, none of these studies evaluated the recovery efficiencies of human noroviruses from large volumes of water.The objective of this study was to evaluate the NanoCeram (Argonide, Sanford, FL) cartridge filter for the concentration of enteroviruses and noroviruses from large volumes of water. NanoCeram filters have an active component of nano alumina (AlOOH) fibers, which give them a naturally occurring electropositive charge.