Concentration of poliovirus from tap water using positively charged microporous filters.

Microporous filters that are more electropositive than the negatively charged filters currently used for virus concentrations from water by filter adsorption-elution methods were evaluated for poliovirus recovery from tap water. Zeta Plus filters composed of diatomaceous earth-cellulose-"charge-modified" resin mixtures and having a net positive charge of up to pH 5 to 6 efficiently adsorbed poliovirus from tap water at ambient pH levels 7.0 to 7.5 without added multivalent cation salts. The adsorbed virus were eluted with glycine-NaOH, pH 9.5 to 11.5. Electropositive asbestos-cellulose filters efficiently adsorbed poliovirus from tap water without added multivalent cation salts between pH 3.5 and 9.0, and the absorbed viruses could be eluted with 3% beef extract, pH 9, but not with pH 9.5 to 11.5 glycine-NaOH. Under water quality conditions in which poliovirus recoveries from large volumes of water were less than 5% with conventional negatively charged filters and standard methods, recoveries with Zeta Plus filters averaged 64 and 22.5% for one- and two-stage concentration procedures, respectively. Electropositive filters appear to offer distinct advantages over conventional negatively charged filters for concentrating enteric viruses from water, and their behavior tends to confirm the importance of electrostatic forces in virus recovery from water by microporous filter adsorption-elution methods.     

Concentration of poliovirus from tap water using positively charged microporous filters.

Microporous filters that are more electropositive than the negatively charged filters currently used for virus concentrations from water by filter adsorption-elution methods were evaluated for poliovirus recovery from tap water. Zeta Plus filters composed of diatomaceous earth-cellulose-"charge-modified" resin mixtures and having a net positive charge of up to pH 5 to 6 efficiently adsorbed poliovirus from tap water at ambient pH levels 7.0 to 7.5 without added multivalent cation salts. The adsorbed virus were eluted with glycine-NaOH, pH 9.5 to 11.5. Electropositive asbestos-cellulose filters efficiently adsorbed poliovirus from tap water without added multivalent cation salts between pH 3.5 and 9.0, and the absorbed viruses could be eluted with 3% beef extract, pH 9, but not with pH 9.5 to 11.5 glycine-NaOH. Under water quality conditions in which poliovirus recoveries from large volumes of water were less than 5% with conventional negatively charged filters and standard methods, recoveries with Zeta Plus filters averaged 64 and 22.5% for one- and two-stage concentration procedures, respectively. Electropositive filters appear to offer distinct advantages over conventional negatively charged filters for concentrating enteric viruses from water, and their behavior tends to confirm the importance of electrostatic forces in virus recovery from water by microporous filter adsorption-elution methods.     

Improved method for recovery of bacteriophage from large volumes of water using negatively charged microporous filters

Current virus-recovery procedures using negatively charged microporous filters provide an inexpensive, reliable method for the recovery and detection of enteroviruses from water and wastewater; however, adjustment of the test samples to pH 3.5 to promote enterovirus adsorption results in significant inactivation of bacteriophage and an inability to simultaneously recover them from large volumes of water using this procedure. Procedures specifically designed for the detection of bacteriophage are currently in use but generally are only effective for small volumes of water. Positively charged filters can be used to recover both enteroviruses and bacteriophage from large volumes of water at neutral pH; however, the filters are expensive. The addition of manganese chloride to test solutions at pH 3.5 prior to filtration through negatively charged Filterite filters allowed for sampling of larger volumes of water by reducing the inactivation of bacteriophage and increasing the recovery of PRD1, MS2, and naturally isolated bacteriophage by a factor of four or five when compared with recoveries from solutions without MnCl2. This method provides an inexpensive, reliable alternative to large-volume bacteriophage recovery procedures that use positively charged filters at neutral pH.     

Concentration of coliphages from large volumes of water and wastewater.

Membrane filter adsorption-elution technology has been extensively used for the concentration and detection of animal viruses from large volumes of water. This study describes the development of positively charged microporous filters (Zeta Plus) for the concentration of coliphages from large volumes of water and wastewater. Four different coliphages were studied: MS-2, phi X174, T2, and T4. Positively charged microporous filters were found to efficiently adsorb these coliphages from tap water, sewage, and lake water at neutral pH. Adsorbed viruses were eluted with a 1:1 mixture of 8% beef extract and 1 M sodium chloride at pH 9. Using this method, coliphages could be concentrated from 17-liter volumes of tap water with recoveries ranging from 34 to 100%. Coliphages occurring naturally in raw and secondarily treated sewage were recovered with average efficiencies of 56.5 and 55.0%, respectively. This method should be useful in isolation of rare phages, the ecology of phages in natural waters, and the evaluation of water quality.     

Evaluation of filters for recovery of Campylobacter jejuni from water.

Campylobacter jejuni has been incriminated in several large waterborne outbreaks, but it has rarely been isolated from water itself. Better methodology is needed for the isolation of C. jejuni from water. We evaluated three types of 0.45-micron microporous filters and three different pore sizes of positively charged depth filters for their ability to recover C. jejuni from seeded, sterile tap and surface water. The microporous filters tested were Millipore HA, Gelman GN6, and Zetapor. Three pore sizes of Zeta Plus depth filters (05S, 30S, and 50S) were evaluated by using an adsorption-elution technique. The overall percent recovery in both tap and surface water by microporous filters was: Zetapor, 66%; Millipore HA, 33%; and Gelman GN6, 33%. Adsorption-elution with Zeta Plus 50S allowed 89% recovery of C. jejuni. These data suggest that both the positively charged Zetapor microporous filter and the Zeta Plus 50S depth filter are effective filters for the recovery of C. jejuni from water.     

Evaluation of filters for recovery of Campylobacter jejuni from water

Campylobacter jejuni has been incriminated in several large waterborne outbreaks, but it has rarely been isolated from water itself. Better methodology is needed for the isolation of C. jejuni from water. We evaluated three types of 0.45-micron microporous filters and three different pore sizes of positively charged depth filters for their ability to recover C. jejuni from seeded, sterile tap and surface water. The microporous filters tested were Millipore HA, Gelman GN6, and Zetapor. Three pore sizes of Zeta Plus depth filters (05S, 30S, and 50S) were evaluated by using an adsorption-elution technique. The overall percent recovery in both tap and surface water by microporous filters was: Zetapor, 66%; Millipore HA, 33%; and Gelman GN6, 33%. Adsorption-elution with Zeta Plus 50S allowed 89% recovery of C. jejuni. These data suggest that both the positively charged Zetapor microporous filter and the Zeta Plus 50S depth filter are effective filters for the recovery of C. jejuni from water.     

Concentration of poliovirus from tap water onto membrane filters with aluminum chloride at ambient pH levels.

A method is described for the concentration of an enterovirus from large volumes of tap water by addition of small amounts of aluminum chloride to enhance virus removal by membrane filters. Tap water treated with 2 X 10(-5) M aluminum chloride showed a slight decrease in pH (less than 0.5), a slight increase in turbidity, and enhanced removal of poliovirus by membrane filters. Virus was quantitatively recovered by treating the filters with a basic buffer, and this eluate was reconcentrated to a small volume by adsorption to aluminum hydroxide flocs. Using these procedures, virus from 1,000 liters of water was reduced to a final eluate of 20 to 80 ml with a mean recovery of 70%.     

Concentration of poliovirus from tap water onto membrane filters with aluminum chloride at ambient pH levels.

A method is described for the concentration of an enterovirus from large volumes of tap water by addition of small amounts of aluminum chloride to enhance virus removal by membrane filters. Tap water treated with 2 X 10(-5) M aluminum chloride showed a slight decrease in pH (less than 0.5), a slight increase in turbidity, and enhanced removal of poliovirus by membrane filters. Virus was quantitatively recovered by treating the filters with a basic buffer, and this eluate was reconcentrated to a small volume by adsorption to aluminum hydroxide flocs. Using these procedures, virus from 1,000 liters of water was reduced to a final eluate of 20 to 80 ml with a mean recovery of 70%.     

Concentration of simian rotavirus SA-11 from tap water by membrane filtration and organic flocculation

Simian rotavirus SA-11 was concentrated from tap water by adsorption to and elution from microporous filters, followed by organic flocculation. Two types of filters were compared for their ability to concentrate the virus. Both Zeta Plus 60S and Cox AA type M-780 filters were efficient for virus adsorption, but the efficiency of virus elution was higher with Zeta Plus than with Cox filters. Optimum conditions for virus recovery from Zeta Plus filters included an input water pH of 6.5 to 7.5 and the use of 3% beef extract (pH 9.0) for elution. Under these conditions, an average of 62 to 100% of the virus was recovered in the concentrate. Organic flocculation was used as a second-step concentration method, with average recoveries of 47 to 69%. When the two methods were used to concentrate small numbers (7 to 75 PFU/liter) of input rotavirus, an average of 75 +/- 40% recovery was achieved. With large volumes of input water, however, recovery was reduced to 16 +/- 7%.     

Concentration of Simian Rotavirus SA-11 from Tap Water by Membrane Filtratiòn and Organic Flocculation

Simian rotavirus SA-11 was concentrated from tap water by adsorption to and elution from microporous filters, followed by organic flocculation. Two types of filters were compared for their ability to concentrate the virus. Both Zeta Plus 60S and Cox AA type M-780 filters were efficient for virus adsorption, but the efficiency of virus elution was higher with Zeta Plus than with Cox filters. Optimum conditions for virus recovery from Zeta Plus filters included an input water pH of 6.5 to 7.5 and the use of 3% beef extract (pH 9.0) for elution. Under these conditions, an average of 62 to 100% of the virus was recovered in the concentrate. Organic flocculation was used as a second-step concentration method, with average recoveries of 47 to 69%. When the two methods were used to concentrate small numbers (7 to 75 PFU/liter) of input rotavirus, an average of 75 ± 40% recovery was achieved. With large volumes of input water, however, recovery was reduced to 16 ± 7%.     

Evaluation of MK filters for recovery of enteroviruses from tap water.

The MK filter is an electropositively charged filter that can be used to concentrate enteroviruses from large volumes (400 to 1,000 liters) of water. This filter is less expensive than the commonly used 1MDS electropositive filter. In this study, we compared the recovery of poliovirus 1 (PV1) and that of coxsackievirus B3 (CB3) from 378 liters of tap water, using both the MK and the 1MDS filters. Viruses were eluted from the filters with 3% beef extract buffered with 0.05 M glycine (pH 9.5) and reconcentrated via organic flocculation. At high virus inputs (approximately 10(6) PFU), the overall recovery (after elution and reconcentration) of PV1 and CB3 from tap water with the MK filter was less than that achieved with the 1MDS filter (P < 0.05). The recoveries of PV1 from tap water with the MK and 1MDS filters were 73.2% +/- 26% (n = 5 trials) and 90.2% +/- 5.9% (n = 5 trials), respectively. The recoveries of CB3 from tap water with the MK and 1MDS filters were 32.8% +/- 34.5% (n = 4 trials) and 95.8% +/- 12.0% (n = 4 trials), respectively. This study indicated that the MK filter consistently provided lower recovery, with wider variability, of PV1 and CB3 from tap water than the 1MDS filter.     

Concentration of coliphage from water and sewage with charge-modified filter aid

Methods of detecting and concentrating animal viruses from large volumes of water and wastewater have experienced rapid development in recent years, but only a few methods are available for the concentration of bacteriophages. The present study describes the use of a charge-modified (Zeta Plus) filter aid (AMF Cuno, Meriden, Conn.) for the concentration of coliphages from large volumes of water and sewage. Coliphages MS-2 and f2 were efficiently adsorbed from water and sewage to the positively charged filter aid. Elution was accomplished with 4% beef extract--0.5 M NaCl adjusted to pH 9.5. The recovery of f2 from 10- to 20-liter volumes of tap water ranged between 11 and 70%, and the recovery of MS-2 ranged between 43 and 70%. The efficiency of recovery of naturally occurring coliphages from secondarily treated sewage ranged between 16 and 44%. This technique appears to be promising because it requires low-cost equipment (47-mm polypropylene filter housing), is easy to handle, and can filter large volumes of water (greater than or equal to 20 liters) with good recoveries. Filtrations can be conducted at the ambient pH of the water, and the unit cost per filtration (i.e., the cost of filter aid) comes to less than three cents per sampling. The technique could be useful in evaluation of viral water quality, study of ecology and occurrence of phages in natural waters, and isolation of rare phages from natural waters.     

Concentration of coliphage from water and sewage with charge-modified filter aid.

Methods of detecting and concentrating animal viruses from large volumes of water and wastewater have experienced rapid development in recent years, but only a few methods are available for the concentration of bacteriophages. The present study describes the use of a charge-modified (Zeta Plus) filter aid (AMF Cuno, Meriden, Conn.) for the concentration of coliphages from large volumes of water and sewage. Coliphages MS-2 and f2 were efficiently adsorbed from water and sewage to the positively charged filter aid. Elution was accomplished with 4% beef extract--0.5 M NaCl adjusted to pH 9.5. The recovery of f2 from 10- to 20-liter volumes of tap water ranged between 11 and 70%, and the recovery of MS-2 ranged between 43 and 70%. The efficiency of recovery of naturally occurring coliphages from secondarily treated sewage ranged between 16 and 44%. This technique appears to be promising because it requires low-cost equipment (47-mm polypropylene filter housing), is easy to handle, and can filter large volumes of water (greater than or equal to 20 liters) with good recoveries. Filtrations can be conducted at the ambient pH of the water, and the unit cost per filtration (i.e., the cost of filter aid) comes to less than three cents per sampling. The technique could be useful in evaluation of viral water quality, study of ecology and occurrence of phages in natural waters, and isolation of rare phages from natural waters.     

Method for Salmonella concentration from water at pH 3.5, using micro-fiber glass filters.

A method is described for the concentration of Salmonella from water. As is done with enterovirus, Salmonella bacteria were concentrated from water in two steps: by pH 3.5 adsorption on and pH 9.5 elution from 8-micron porosity micro-fiber glass filter tubes. This method worked in less than 30 min, and Salmonella typhimurium was inactivated only slightly in spite of rapid pH variations (pH 3.5 to 9.5). It was demonstrated that the retention by the filters stems from two phenomena: a low retention in the micro-fiber glass labyrinth for small filtered volumes, and a high retention by adsorption at pH 3.5 for any filtered volume (experiments done with 15- and 80-liter samples). Addition in tap water of trivalent ions like Al3+ did not increase Salmonella adsorption. In most of the trials, Salmonella recovery varied from 42 to 93%. Preliminary field investigations indicate that enterovirus and Salmonella may both be concentrated from the same water sample by this procedure.     

Method for Salmonella concentration from water at pH 3.5, using micro-fiber glass filters.

A method is described for the concentration of Salmonella from water. As is done with enterovirus, Salmonella bacteria were concentrated from water in two steps: by pH 3.5 adsorption on and pH 9.5 elution from 8-micron porosity micro-fiber glass filter tubes. This method worked in less than 30 min, and Salmonella typhimurium was inactivated only slightly in spite of rapid pH variations (pH 3.5 to 9.5). It was demonstrated that the retention by the filters stems from two phenomena: a low retention in the micro-fiber glass labyrinth for small filtered volumes, and a high retention by adsorption at pH 3.5 for any filtered volume (experiments done with 15- and 80-liter samples). Addition in tap water of trivalent ions like Al3+ did not increase Salmonella adsorption. In most of the trials, Salmonella recovery varied from 42 to 93%. Preliminary field investigations indicate that enterovirus and Salmonella may both be concentrated from the same water sample by this procedure.     

Concentration of viruses in beef extract by flocculation with ammonium sulfate

Bacteriophages and enteroviruses in water were adsorbed to positively charged filters (Virosorb 1MDS [AMF Cuno, Inc., Meriden, Conn.] or Seitz S [Republic Filters, Milldaler, Conn.]). Adsorbed viruses were eluted by treating the filters with 10% beef extract, pH 9. Organic flocculation of the beef extract at pH 3.5 permitted recovery of more than 40% of the enteroviruses tested but less than 15% of the bacteriophages present. A method was developed that uses salts at pH 7 to flocculate beef extract. Two volumes of saturated ammonium sulfate were added to beef extract, and both enteroviruses and bacteriophages were adsorbed to the flocs that formed. Greater than 70% of the enteroviruses and bacteriophages were recovered by centrifuging the sample and suspending the flocs in a small volume of distilled water.     

Recovery of Cryptosporidium oocysts from small and large volume water samples using a compressed foam filter system

A novel filter system comprising open cell reticulated foam rings compressed between retaining plates and fitted into a filtration housing was evaluated for the recovery of oocysts of Cryptosporidium from water. Mean recoveries of 90·2% from seeded small and large volume (100–2000 l) tap water samples, and 88·8% from 10–20 l river water samples, were achieved. Following a simple potassium citrate flotation concentrate clean-up procedure, mean recoveries were 56·7% for the tap water samples and 60·9% for river water samples. This represents a marked improvement in capture and recovery of Cryptosporidium oocysts from water compared with conventional polypropylene wound cartridge filters and membrane filters.     

An improved method for concentrating rotavirus from water samples

A modified adsorption-elution method for the concentration of seeded rotavirus from water samples was used to determine various factors which affected the virus recovery. An enzyme-linked immunosorbent assay was used to detect the rotavirus antigen after concentration. Of the various eluents compared, 0.05M glycine, pH 11.5 gave the highest rotavirus antigen recovery using negatively charged membrane filtration whereas 2.9% tryptose phosphate broth containing 6% glycine; pH 9.0 was found to give the greatest elution efficiency when a positively charged membrane was used. Reconcentration of water samples by a speedVac concentrator showed significantly higher rotavirus recovery than polyethylene glycol precipitation through both negatively and positively charged filters (p-value <0.001). In addition, speedVac concentration using negatively charged filtration resulted in greater rotavirus recovery than that using positively charged filtration (p-value = 0.004). Thirty eight environmental water samples were collected from river, domestic sewage, canals receiving raw sewage drains, and tap water collected in containers for domestic use, all from congested areas of Bangkok. In addition, several samples of commercial drinking water were analyzed. All samples were concentrated and examined for rotavirus antigen. Coliforms and fecal coliforms (0->1,800 MPN/100 ml) were observed but rotavirus was not detected in any sample. This study suggests that the speedVac reconcentration method gives the most efficient rotavirus recovery from water samples.     

Concentration of enteroviruses from large volumes of tap water, treated sewage, and seawater.

Methods are described for the efficient concentration of an enterovirus from large volumes of tap water, sewage, and seawater. Virus in acidified water (pH 3.5) in the presence of aluminum chloride was adsorbed to a 10-inch (ca. 25.4 cm) fiberglass depth cartridge and a 10-inch pleated epoxy-fiberglass filter in a series at flow rates of up to 37.8 liters (10 gallons) per min. Adsorbed viruses were eluted from the filters with glycine buffer (pH 10.5 to 11.5), and the eluate was reconcentrated by using a combination of aluminum flocculation followed by hydroextraction. With this procedure, poliovirus in large volumes of tap water, seawater, and sewage could be concentrated with an average efficiency of 52, 53, and 50%, respectively. It was demonstrated that this method is capable of detecting surface solid-associated viruses originating from sewage treatment plants. No difference in virus recovery between laboratory batch studies and a set-up with acid-salt injection was found. This unified scheme for the concentration of viruses has many advantages over previously described systems. These include: high operating flow rates, low weight and small size, effectiveness with a variety of waters with widely varying qualities, and filters with a high resistance to clogging.     

Concentration of enteroviruses from large volumes of tap water, treated sewage, and seawater.

Methods are described for the efficient concentration of an enterovirus from large volumes of tap water, sewage, and seawater. Virus in acidified water (pH 3.5) in the presence of aluminum chloride was adsorbed to a 10-inch (ca. 25.4 cm) fiberglass depth cartridge and a 10-inch pleated epoxy-fiberglass filter in a series at flow rates of up to 37.8 liters (10 gallons) per min. Adsorbed viruses were eluted from the filters with glycine buffer (pH 10.5 to 11.5), and the eluate was reconcentrated by using a combination of aluminum flocculation followed by hydroextraction. With this procedure, poliovirus in large volumes of tap water, seawater, and sewage could be concentrated with an average efficiency of 52, 53, and 50%, respectively. It was demonstrated that this method is capable of detecting surface solid-associated viruses originating from sewage treatment plants. No difference in virus recovery between laboratory batch studies and a set-up with acid-salt injection was found. This unified scheme for the concentration of viruses has many advantages over previously described systems. These include: high operating flow rates, low weight and small size, effectiveness with a variety of waters with widely varying qualities, and filters with a high resistance to clogging.