Electropositive Filter Research Articles

Evaluation of Sampling and Concentration Methods for Salmonella enterica Serovar Typhi Detection from Wastewater.

Salmonella enterica serovar (Salmonella Typhi) is the causative bacterial agent of typhoid fever. Environmental surveillance of wastewater and wastewater-impacted surface waters has proven effective in monitoring various pathogens and has recently been applied to Salmonella Typhi. This study evaluated eight sample collection and concentration methods with 12 variations currently being developed and used for Salmonella Typhi surveillance globally to better understand the performance of each method based on its ability to detect Salmonella Typhi and its feasibility. Salmonella Typhi strains Ty21a and Ty2 were seeded to influent wastewater at known concentrations to evaluate the following methods: grab sampling using electropositive filters, centrifugation, direct enrichment, or membrane filtration and trap sampling using Moore swabs. Concentrated samples underwent nucleic acid extraction and were detected and/or quantified via quantitative polymerase chain reaction (qPCR). Results suggest that all methods tested can be successful at concentrating Salmonella Typhi for subsequent detection by qPCR, although each method has its own strengths and weaknesses, including the Salmonella Typhi concentration it is best suited for, with a range of positive detections observed as low as 0.1-0.001 colony-forming units (CFU) Ty21a/mL and 0.01 CFU Ty2/mL. These factors should be considered when identifying a method for environmental surveillance and will greatly depend on the use case planned.

Comparison of nanodiamond coated quartz filter with commercial electropositive filters: Zeta potential and dye retention study

Diamond coated quartz(DQ) electropositive filters were compared with a few commercially available electropositive filters. The zeta potential of all the filters were measured and it was found that DQ had higher positive zeta potential over a wide pH range. Dye retention tests, using acid black II dye, performed on all the filters showed DQ to have higher retention. The study clearly showed that, under laboratory conditions, DQ has better performance compared to many commercial electropositive filters.

Method for detecting norovirus, hepatitis A and hepatitis E viruses in tap and bottled drinking water

Viruses are a leading cause of foodborne disease worldwide. Human norovirus and hepatitis viruses (hepatitis A (HAV) and hepatitis E (HEV)) are recognised to be the main viruses of importance to public health. The ISO 15216 procedure describes molecular methods for detecting HAV and norovirus in bottled water by using an electropositive filter to concentrate viruses. The aim of this study was to validate the Zeta Plus 1MDS membrane (1MDS) for detecting enteric viruses from tap and bottled water using the recent international standard ISO/DIS/16140-4:2018, which describes the protocol for validating methods for microbiology in the food chain. Method with direct lysis of viruses from the 1MDS filter, and RNA extraction was used for detecting noroviruses, HAV and HEV from different tap and bottled drinking water. By taking into account virus's inoculation levels above the LOD, the recovery rates of noroviruses and HAV obtained from pure RNA extracts ranged from 2.50% to 14.31% and for HEV from 27.87% to 53.54% according to the water samples analysed. The virus recovery rates did not differ according to the operator or drinking water analysed but did according to the virus inoculated. The LOD95 values were respectively 50 genome copies/mL for HAV and 2.8 genome copies/mL for HEV, 420 genome copies/mL for norovirus GI and 134 genome copies/mL of water sample for norovirus GII. LOQs were determined for HAV and HEV by the total error approach and were 15.8 genome copies/mL for HAV and 2.8 genome copies/mL of water sample for HEV. The described method could be used for detecting viruses from tap and bottled water for routine diagnosis needs.

Evaluation of reduction efficiencies of pepper mild mottle virus and human enteric viruses in full-scale drinking water treatment plants employing coagulation-sedimentation–rapid sand filtration or coagulation–microfiltration

Here, we evaluated the reduction efficiencies of indigenous pepper mild mottle virus (PMMoV, a potential surrogate for human enteric viruses to assess virus removal by coagulation-sedimentation–rapid sand filtration [CS–RSF] and coagulation–microfiltration [C–MF]) and representative human enteric viruses in four full-scale drinking water treatment plants that use CS–RSF (Plants A and B) or C–MF (Plants C and D). First, we developed a virus concentration method by using an electropositive filter and a tangential-flow ultrafiltration membrane to effectively concentrate and recover PMMoV from large volumes of water: the recovery rates of PMMoV were 100% when 100-L samples of PMMoV-spiked dechlorinated tap water were concentrated to 20 mL; even when spiked water volume was 2000 L, recovery rates of >30% were maintained. The concentrations of indigenous PMMoV in raw and treated water samples determined by using this method were always above the quantification limit of the real-time polymerase chain reaction assay. We therefore were able to determine its reduction ratios: 0.9–2.7-log10 in full-scale CS–RSF and 0.7–2.9-log10 in full-scale C–MF. The PMMoV reduction ratios in C–MF at Plant C (1.0 ± 0.3-log10) were lower than those in CS–RSF at Plants A (1.7 ± 0.5-log10) and B (1.4 ± 0.7-log10), despite the higher ability of MF for particle separation in comparison with RSF owing to the small pore size in MF. Lab-scale virus-spiking C–MF experiments that mimicked full-scale C–MF revealed that a low dosage of coagulant (polyaluminum chloride [PACl]) applied in C–MF, which is determined mainly from the viewpoint of preventing membrane fouling, probably led to the low reduction ratios of PMMoV in C–MF. This implies that high virus reduction ratios (>4-log10) achieved in previous lab-scale virus-spiking C–MF studies are not necessarily achieved in full-scale C–MF. The PMMoV reduction ratios in C–MF at Plant D (2.2 ± 0.6-log10) were higher than those at Plant C, despite similar coagulant dosages. In lab-scale C–MF, the PMMoV reduction ratios increased from 1-log10 (with PACl [basicity 1.5], as at Plant C) to 2–4-log10 (with high-basicity PACl [basicity 2.1], as at Plant D), suggesting that the use of high-basicity PACl probably resulted in higher reduction ratios of PMMoV at Plant D than at Plant C. Finally, we compared the reduction ratios of indigenous PMMoV and representative human enteric viruses in full-scale CS–RSF and C–MF. At Plant D, the concentrations of human norovirus genogroup II (HuNoV GII) in raw water were sometimes above the quantification limit; however, whether its reduction ratios in C–MF were higher than those of PMMoV could not be judged since reduction ratios were >1.4-log10 for HuNoV GII and 2.3–2.9-log10 for PMMoV. At Plant B, the concentrations of enteroviruses (EVs) and HuNoV GII in raw water were above the quantification limit on one occasion, and the reduction ratios of EVs (>1.2-log10) and HuNoV GII (>1.5-log10) in CS–RSF were higher than that of PMMoV (0.9-log10). This finding supports the usefulness of PMMoV as a potential surrogate for human enteric viruses to assess virus removal by CS–RSF.

Evaluation of three different filters and two methods for recovering viruses from drinking water

Among the enteric viruses implicated in waterborne outbreaks, human norovirus and hepatitis A virus (HAV) are a serious public health issue. Most foodborne viruses are difficult or currently unlikely to cultivate. Because of the lack of a cell culture method, real-time reverse transcriptase PCR is commonly used for the detection of norovirus in foodstuffs and environmental samples. Due to low infectious doses in humans and low virus concentration in water sample, filter adsorption methods were used for concentrating viruses from water. The ISO (Anonymous, ISO 15216-1, 2017) describes standardized molecular methods for detecting HAV and norovirus in bottled water. This method includes a two-step procedure: concentrating the virus using a microporous electropositive filter (47 mm diameter, 0.45 μm pore size) then molecular detection. The Zetapor filter, which had a charged membrane with a pore size of 0.45 μm, was commonly used in the past to concentrate viruses from water or from salad leaves following virus elution. But, unfortunately, the Zetapor filter is no longer marketed and it is therefore necessary to assess an alternative filter. The aim of this study was to compare the ability of two electropositive filters with a pore size of 0.45 μm or 0.22 μm and one uncharged filter (0.45 μm) to recover norovirus and HAV from two different types of drinking water (bottled water and tap water) with the adsorption-elution method proposed by ISO (Anonymous, ISO 15216-1, 2017) (method A) and with direct viral extraction using filters (method B). The mean extraction yields for norovirus and HAV calculated with RNA extracts ranged from 0.2 % – 4.81 % with method A and from 5.05 % – 53.58 % with method B, and did not differ significantly between the two types of drinking water tested. For method B, the mean extraction yields for HAV and norovirus were evaluated according to results from the three filters used. The recovery rate of HAV and norovirus ranged between 3.47 % and 62.41 % with the 0.45 μm electropositive filter and were higher than the other filters. The 0.45 μm electropositive filter could be used to concentrate viruses for routine viral monitoring of drinking water for researchers who want to adopt the method in their lab routine.

Development of an elution device for ViroCap virus filters

Environmental surveillance of waterborne pathogens is vital for monitoring the spread of diseases, and electropositive filters are frequently used for sampling wastewater and wastewater-impacted surface water. Viruses adsorbed to electropositive filters require elution prior to detection or quantification. Elution is typically facilitated by a peristaltic pump, although this requires a significant startup cost and does not include biosafety or cross-contamination considerations. These factors may pose a barrier for low-resource laboratories that aim to conduct environmental surveillance of viruses. The objective of this study was to develop a biologically enclosed, manually powered, low-cost device for effectively eluting from electropositive ViroCap™ virus filters. The elution device described here utilizes a non-electric bilge pump, instead of an electric peristaltic pump or a positive pressure vessel. The elution device also fully encloses liquids and aerosols that could contain biological organisms, thereby increasing biosafety. Moreover, all elution device components that are used in the biosafety cabinet are autoclavable, reducing cross-contamination potential. This device reduces costs of materials while maintaining convenience in terms of size and weight. With this new device, there is little sample volume loss due to device inefficiency, similar virus yields were demonstrated during seeded studies with poliovirus type 1, and the time to elute filters is similar to that required with the peristaltic pump. The efforts described here resulted in a novel, low-cost, manually powered elution device that can facilitate environmental surveillance of pathogens through effective virus recovery from ViroCap filters while maintaining the potential for adaptability to other cartridge filters.

EPA Method 1615. Measurement of Enterovirus and Norovirus Occurrence in Water by Culture and RT-qPCR. II. Total Culturable Virus Assay

A standardized method is required when national studies on virus occurrence in environmental and drinking waters utilize multiple analytical laboratories. The U.S Environmental Protection Agency’s (USEPA) Method 1615 was developed with the goal of providing such a standard for measuring Enterovirus and Norovirus in these waters. Virus is concentrated from water using an electropositive filter, eluted from the filter surface with beef extract, and then concentrated further using organic flocculation. Herein we present the protocol from Method 1615 for filter elution, secondary concentration, and measurement of total culturable viruses. A portion of the concentrated eluate from each sample is inoculated onto ten replicate flasks of Buffalo Green Monkey kidney cells. The number of flasks demonstrating cytopathic effects is used to quantify the most probable number (MPN) of infectious units per liter. The method uses a number of quality controls to increase data quality and to reduce interlaboratory and intralaboratory variation. Laboratories must meet defined performance standards. Method 1615 was evaluated by examining virus recovery from reagent-grade and ground waters seeded with Sabin poliovirus type 3. Mean poliovirus recoveries with the total culturable assay were 111% in reagent grade water and 58% in groundwaters.

EPA Method 1615. Measurement of Enterovirus and Norovirus Occurrence in Water by Culture and RT-qPCR. II. Total Culturable Virus Assay.

A standardized method is required when national studies on virus occurrence in environmental and drinking waters utilize multiple analytical laboratories. The U.S Environmental Protection Agency’s (USEPA) Method 1615 was developed with the goal of providing such a standard for measuring Enterovirus and Norovirus in these waters. Virus is concentrated from water using an electropositive filter, eluted from the filter surface with beef extract, and then concentrated further using organic flocculation. Herein we present the protocol from Method 1615 for filter elution, secondary concentration, and measurement of total culturable viruses. A portion of the concentrated eluate from each sample is inoculated onto ten replicate flasks of Buffalo Green Monkey kidney cells. The number of flasks demonstrating cytopathic effects is used to quantify the most probable number (MPN) of infectious units per liter. The method uses a number of quality controls to increase data quality and to reduce interlaboratory and intralaboratory variation. Laboratories must meet defined performance standards. Method 1615 was evaluated by examining virus recovery from reagent-grade and ground waters seeded with Sabin poliovirus type 3. Mean poliovirus recoveries with the total culturable assay were 111% in reagent grade water and 58% in groundwaters.

An acute gastroenteritis outbreak caused by GII.17 norovirus in Jiangsu Province, China.

Noroviruses are a common cause of acute gastroenteritis around the world; however, reports of outbreaks caused by GII.17 norovirus are rare. An outbreak caused by GII.17 norovirus in a senior high school in Wuxi, Jiangsu Province, China is reported here. An epidemiological investigation, pathogen detection, and case-control study were performed. Epidemiological data combined with the epidemic curve indicated that this outbreak was a point source type initially, followed by secondary transmission. The first case was identified as most likely the source of the outbreak. Risk analysis showed exposure to patients and sharing a communal water cooler to be associated with the spread of infection. Sequence analysis of GII-positive samples confirmed that the norovirus GII.17 variant was the etiological agent of this outbreak.

Solar water disinfection (SODIS): Impact on hepatitis A virus and on a human Norovirus surrogate under natural solar conditions.

This study evaluates the effectiveness of solar water disinfection (SODIS) in the reduction and inactivation of hepatitis A virus (HAV) and of the human Norovirus surrogate, murine Norovirus (MNV-1), under natural solar conditions. Experiments were performed in 330 ml polyethylene terephthalate (PET) bottles containing HAV or MNV-1 contaminated waters (10(3) PFU/ml) that were exposed to natural sunlight for 2 to 8 h. Parallel experiments under controlled temperature and/or in darkness conditions were also included. Samples were concentrated by electropositive charged filters and analysed by RT-real time PCR (RT-qPCR) and infectivity assays. Temperature reached in bottles throughout the exposure period ranged from 22 to 40ºC. After 8 h of solar exposure (cumulative UV dose of ~828 kJ/m2 and UV irradiance of ~20 kJ/l), the results showed significant (P<0.05) reductions from 4.0 (+/-0.56)x10(4) to 3.15 (+/-0.69)x10(3) RNA copies/100ml (92.1%, 1.1 log) for HAV and from 5.91 (+/-0.59)x10(4) to 9.24 (+/-3.91)x10(3) RNA copies/100 ml (84.4%, 0.81 log) for MNV-1. SODIS conditions induced a loss of infectivity between 33.4% and 83.4% after 4 to 8 h in HAV trials, and between 33.4% and 66.7% after 6 h to 8 h in MNV-1 trials. The results obtained indicated a greater importance of sunlight radiation over the temperature as the main factor for viral reduction.

A Small Volume Procedure for Viral Concentration from Water

Small-scale concentration of viruses (sample volumes 1-10 L, here simulated with spiked 100 ml water samples) is an efficient, cost-effective way to identify optimal parameters for virus concentration. Viruses can be concentrated from water using filtration (electropositive, electronegative, glass wool or size exclusion), followed by secondary concentration with beef extract to release viruses from filter surfaces, and finally tertiary concentration resulting in a 5-30 ml volume virus concentrate. In order to identify optimal concentration procedures, two different electropositive filters were evaluated (a glass/cellulose filter [1MDS] and a nano-alumina/glass filter [NanoCeram]), as well as different secondary concentration techniques; the celite technique where three different celite particle sizes were evaluated (fine, medium and large) followed by comparing this technique with that of the established organic flocculation method. Various elution additives were also evaluated for their ability to enhance the release of adenovirus (AdV) particles from filter surfaces. Fine particle celite recovered similar levels of AdV40 and 41 to that of the established organic flocculation method when viral spikes were added during secondary concentration. The glass/cellulose filter recovered higher levels of both, AdV40 and 41, compared to that of a nano-alumina/glass fiber filter. Although not statistically significant, the addition of 0.1% sodium polyphosphate amended beef extract eluant recovered 10% more AdV particles compared to unamended beef extract.

A Small Volume Procedure for Viral Concentration from Water

Small-scale concentration of viruses (sample volumes 1-10 L, here simulated with spiked 100 ml water samples) is an efficient, cost-effective way to identify optimal parameters for virus concentration. Viruses can be concentrated from water using filtration (electropositive, electronegative, glass wool or size exclusion), followed by secondary concentration with beef extract to release viruses from filter surfaces, and finally tertiary concentration resulting in a 5-30 ml volume virus concentrate. In order to identify optimal concentration procedures, two different electropositive filters were evaluated (a glass/cellulose filter [1MDS] and a nano-alumina/glass filter [NanoCeram]), as well as different secondary concentration techniques; the celite technique where three different celite particle sizes were evaluated (fine, medium and large) followed by comparing this technique with that of the established organic flocculation method. Various elution additives were also evaluated for their ability to enhance the release of adenovirus (AdV) particles from filter surfaces. Fine particle celite recovered similar levels of AdV40 and 41 to that of the established organic flocculation method when viral spikes were added during secondary concentration. The glass/cellulose filter recovered higher levels of both, AdV40 and 41, compared to that of a nano-alumina/glass fiber filter. Although not statistically significant, the addition of 0.1% sodium polyphosphate amended beef extract eluant recovered 10% more AdV particles compared to unamended beef extract.

A simple and efficient method for detecting avian influenza virus in water samples

Waterborne transmission plays an essential role in the transmission and spread of avian influenza viruses. The abundance of influenza viruses in environmental water is usually extremely low and viruses or viral genomes can hardly be detected by conventional reverse transcription (RT-) PCR without concentration. In the present study, an electropositive filter membrane was used to concentrate influenza viruses from water sample, in addition, a glass fiber filter has been used prior to positive charged membrane for the prefiltration. Unlike the traditional adsorption–elution method, Trizol-LS reagent was used to lyse the viruses attached directly to the electropositive filter membrane and the influenza virus genomic RNA was extracted, followed by RT-PCR analysis. The method established in this study could improve the efficiency of the conventional RT-PCR technique used to detect the M, NP, and HA genes of influenza virus in natural water samples. This method could also reduce the time taken for the traditional adsorption–elution concentration procedure.

Multiplex real-time RT-qPCR for the detection of Norovirus in bottled and tap water using murine norovirus as a process control

To provide a rapid and sensitive method for detecting NoV GI and NoV GII in water and to evaluate the use of the murine norovirus (MNV-1) as a process control. The method is based on viral concentration by filtration on electropositive filters and direct lysis of adsorbed viruses from filters before RNA extraction and RT-qPCR amplification. An one-step multiplex RT-qPCR assay was developed for the simultaneous detection of NoV GI, NoV GII and MNV-1. Then, water samples were artificially contaminated to determine mean virus recoveries and method sensitivity. The method showed a higher sensitivity for detecting NoV GII (10(3) genome copies per 0·5 l) than for NoV GI (10(4) genome copies per 0·5 l) in the presence of MNV-1 regardless of the type of water. The data also showed that MNV-1 is a robust option as process control. The method described provides a valuable tool for the monitoring of potential public health risks associated with NoV contamination in drinkable water. Given the increasing evidence for NoV involvement in food outbreaks, the one-step multiplex RT-qPCR assay we used in this study would be a very useful tool to investigate NoV contamination in other food products.

Quantitative characterization of the inhibitory effects of salt, humic acid, and heavy metals on the recovery of waterborne norovirus by electropositive filters

The virus adsorption-elution technique (VIRADEL) using electropositively charged filters is used frequently for recovering enteric viruses from water. The filter-absorbed virus is typically eluted, concentrated, and subsequently detected by culture or molecular methods. Human norovirus (HuNoV), one of the most important waterborne pathogens, cannot be cultivated by conventional culture methods and is typically detected using a reverse transcriptase-polymerase chain reaction (RT-PCR) assay. However, it is plausible that various inhibitors could be concentrated simultaneously during the VIRADEL process and affect RT-PCR assays. In this study, we evaluated the effect of typical inhibitors, including humic acid, heavy metals, and salt, on the recovery of norovirus by two different electropositive filters: 1MDS and Nanoceram. Known amounts of HuNoV and murine norovirus were inoculated in 1 L of surface water containing various concentrations of humic acid, heavy metals (cadmium and lead), or NaCl. Our results indicate that the presence of heavy metals or salt significantly reduced the recovery of virus from the electropositive filters. Thus, care should be taken when analyzing waterborne norovirus using electropositive filters in environments with high concentrations of heavy metal inhibitors or salts.

Development of a virus concentration method using lanthanum-based chemical flocculation coupled with modified membrane filtration procedures

Direct membrane filtration is often used to concentrate viruses in water but it may suffer from severe membrane fouling and clogging. Here, a lanthanum-based flocculation method coupled with modified membrane filtration procedures was developed and evaluated to detect viruses in large volume (40L) water samples. The lanthanum-based flocculation method could easily reduce the water sample volume by a factor of 40. Additional volume reduction was achieved by a two-step membrane filtration approach. First, selected membrane filters (including 1MDS electropositive filters and nitrocellulose electronegative filters-Millipore HATF filters) were used to reduce water sample volume further and compare their efficiencies in virus recovery. The Mg2+-modified HATF membrane performed better on MS2 retention with an average virus recovery of 83.4% (±4.5% [standard deviation]). After HATF membrane filtration and elution, centrifugal ultrafiltration through a 30kDa cut-off membrane resulted in an overall concentration factor of 20,000. Results from the infectivity assay showed that the MS2 recovery efficiencies from the NanoCeram- and 1MDS-based direct filtration and the lanthanum-based concentration coupled with the modified filtration procedure were 10.1% (±1.0%), 3.3% (±0.1%), and 17.5% (±1.1%), respectively. Results from the PCR analysis showed that the virus recoveries of the lanthanum-based method were 20.6% (±2.9%) and 19.5% (±3.4%) for MS2 and adenovirus, respectively, while no adenovirus could be detected through the NanoCeram- and 1MDS-based direct filtration. The lanthanum-based concentration method coupled with modified membrane filtration procedures is therefore a promising method for detecting waterborne viruses.

Methods for primary concentration of viruses from water samples: a review and meta-analysis of recent studies

Since the beginning of environmental virology in the mid-twentieth century, a key challenge to scientists in the environmental field has been how to collect, isolate and detect pathogenic viruses from water that is used for drinking and/or recreational purposes. Early studies investigated different types of membrane filters, with more sophisticated technologies being developed more recently. The purpose of this study was to look at the current state of the science of methods for the concentration of viruses from water. Several technologies were reviewed, and associated data were included in a meta-analysis which showed that electronegative filters, electropositive filters and ultrafilters are comparable in performance and that significant differences in recovery are due to virus type rather than filter type, water matrix or sample volume. This information is useful, as it will help to determine which method(s) should be used, particularly if there is a specific viral type being targeted for a particular study. In addition, it will be helpful when sampling different environmental water matrices and/or when budget allowance must be taken into consideration. Taken together, this will be useful in performing viral occurrence studies, which ultimately can help ensure safer water for both humans and the environment.

Effectiveness of Poliovirus Concentration and Recovery from Treated Wastewater by Two Electropositive Filter Methods

Enteric viruses are often present in low numbers in various water matrices. Virus sampling therefore involves multiple concentration steps to condense large samples down to small volumes for detection by cell culture or molecular assays. The NanoCeram® Virus Sampler has been demonstrated to be effective for the recovery of viruses from tap water, surface waters, and seawater. The goal of this study was to evaluate a new method using NanoCeram® filters for the recovery of poliovirus 1 (PV-1) from treated wastewater. Activated sludge effluent samples were spiked with PV-1 and concentrated in side-by-side tests by two methods: (1) NanoCeram® filtration, elution with sodium polyphosphate buffer, secondary concentration via centrifugal ultrafiltration; and (2) 1MDS filtration, elution with beef extract, secondary concentration via organic flocculation. The virus retention and elution efficiencies did not differ significantly between the two methods. In contrast, the secondary concentrate volume was smaller for the NanoCeram® method (8.4 vs. 30 mL) and the secondary concentration efficiencies were different between the two methods with 98 % for centrifugal ultrafiltration (NanoCeram® and 45 % for organic flocculation (1MDS). The overall method efficiencies were significantly different (P ≤ 0.05) with the NanoCeram® method yielding a 57 % and the 1MDS a 23 % virus recovery. In addition, there appeared to be less interference with viral detection via polymerase chain reaction with the NanoCeram® concentrates. This NanoCeram® method therefore is able to efficiently recover PV-1 from large volumes of wastewater and may serve as an inexpensive alternative to the standard 1MDS filter method for such applications.

Application of Acidic Elution to Virus Concentration Using Electropositive Filters

The effect of the type and pH of an elution solution on the recovery of poliovirus from water by a virus concentration method using an electropositive filter was evaluated. The experimental results obtained indicated the potential usefulness of H2SO4 (pH 1.5-3.5) as a novel solution for virus elution.

Concentration and Recovery of Viruses from Water: A Comprehensive Review

Enteric viruses are a cause of waterborne disease worldwide, and low numbers in drinking water can present a significant risk of infection. Because the numbers are often quite low, large volumes (100-1,000 L) of water are usually processed. The VIRADEL method using microporous filters is most commonly used today for this purpose. Negatively charged filters require the addition of multivalent salts and acidification of the water sample to effect virus adsorption, which can make large-volume sampling difficult. Positively charged filters require no preconditioning of samples, and are able to concentrate viruses from water over a greater pH range than electronegative filters. The most widely used electropositive filter is the Virosorb 1MDS; however, the Environmental Protection Agency has added the positively charged NanoCeram filters to their proposed Method 1615. Ultrafilters concentrate viruses based on size exclusion rather than electrokinetics, but are impractical for field sampling or processing of turbid water. Elution (recovery) of viruses from filters following concentration is performed with organic (e.g., beef extract) or inorganic solutions (e.g., sodium polyphosphates). Eluates are then reconcentrated to decrease the sample volume to enhance detection methods (e.g., cell culture infectivity assays and molecular detection techniques). While the majority of available filters have demonstrated high virus retention efficiencies, the methods to elute and reconcentrate viruses have met with varying degrees of success due to the biological variability of viruses present in water.