Comparative Inactivation of Enteroviruses and Adenovirus 2 by UV Light

The doses of UV irradiation necessary to inactivate selected enteric viruses on the U.S. Environmental Protection Agency Contaminant Candidate List were determined. Three-log reductions of echovirus 1, echovirus 11, coxsackievirus B3, coxsackievirus B5, poliovirus 1, and human adenovirus type 2 were effected by doses of 25, 20.5, 24.5, 27, 23, and 119 mW/cm², respectively. Human adenovirus type 2 is the most UV light-resistant enteric virus reported to date.     

Influence of pH, salinity, and organic matter on the adsorption of enteric viruses to estuarine sediment.

This study was designed to determine the degree of adsorption of enteric viruses to marine sediment and factors controlling this association. Adsorption and elution characteristics of several enteroviruses and one rotavirus to estuarine sediments were studied under varying conditions of pH, salinity, and presence of soluble organics. Greater than 99% of the added poliovirus type 1 (LSc), coxsackievirus type B3 (Nancy), echovirus type 7 (Wallace), and rotavirus (SA-11) adsorbed to sediment. Echovirus 1 (Farouk) and a recent isolate typed as coxsackievirus B4 adsorbed significantly less than poliovirus 1 under similar conditions of varying salinity and pH. The presence of soluble organic matter, in the form of secondary sewage effluent or humic acid, did not affect these patterns of adsorption. Only echovirus 1 (Farouk) desorbed when the pH or salinity was altered and then only to a small extent. Three recent isolates of echovirus 1 and echovirus 29 (strain JV-10) also demonstrated varying amounts of adsorption to sediment. These data indicate that enteric viruses can become readily associated with sediment in the estuarine environment and that this association may play a major role in their hydrotransportation and survival.     

Ultraviolet Devitalization of Eight Selected Enteric Viruses in Estuarine Water

The effect of ultraviolet (UV) radiation on the devitalization of eight selected enteric viruses suspended in estuarine water was determined. The surviving fractions of each virus were calculated and then plotted against the UV exposure time for purposes of comparison. Analytical assessment of the survival data for each virus consisted of least squares regression analysis for determination of intercepts and slope functions. All data were examined for statistical significance. When the slope function of each virus was compared against the slope function of poliovirus type 1, the analytical findings indicated that poliovirus types 2 and 3, echovirus types 1 and 11, and coxsackievirus A-9 exhibited similar devitalization characteristics in that no statistically significant difference was found (P > 0.05). Conversely, the devitalization characteristics of coxsackievirus B-1 and reovirus type 1 were dissimilar from those of poliovirus type 1 in that a statistically significant difference was found between the slope functions (P < 0.05). This observed difference in devitalization of coxsackievirus B-1 and reovirus type 1 was attributed primarily to the frequency distribution of single and aggregate virions, the geometric configuration, the size of the aggregates, and the severity of aggregation. The devitalization curve of coxsackievirus B-1 was characteristic of a retardant die-away curve. The devitalization curve of reovirus type 1 was characteristic of a multihittype curve. The calculated devitalization half-life values for poliovirus types 1, 2, and 3; echovirus types 1 and 11; coxsackievirus types A-9 and B-1; and reovirus type 1 were 2.8, 3.1, 2.7, 2.8, 3.2, 3.1, 4.0, 4.0 sec, respectively. These basic data should facilitate an operative extrapolation of the findings to the applied situation. It was concluded that UV can be highly effective and provide a reliable safety factor in treating estuarine water.     

Infections du système nerveux central à entérovirus: 223 cas vus à un h?pital pédiatrique entre 1973 et 1981.

Between 1973 and 1981, 223 patients were seen at hôpital Sainte-Justine in Montreal for enteroviral infection of the nervous system. In 161 the cause was documented by isolation of an enterovirus from the cerebrospinal fluid (CSF). The viruses most frequently isolated were echovirus 11 (36 isolates), echovirus 30 (24), coxsackievirus B2 (23), coxsackievirus B3 (19), echovirus 6 (18), coxsackievirus B5 (16), coxsackievirus A9 (15), echovirus 9 (13), echovirus 7 (12) and coxsackievirus B1 (11). Aseptic meningitis was diagnosed in 200 cases and encephalitis in 12. The remaining 11 patients presented with the features of septicemia or with convulsions. In 33 patients an enterovirus was isolated from the CSF in the absence of pleocytosis. Polymorphonuclear cell predominance was noted in the initial CSF sample in 95 cases; it was persistent in 11. There were five mixed infections; in three cases two viruses were isolated from the same CSF sample. Two patients died: one, a child with hypogammaglobulinemia, had fatal polioencephalitis; the other, a 6-month-old infant brought to the emergency room in unexplained cardiopulmonary arrest, had echovirus 6 meningitis. Of the 172 patients admitted to hospital 96 received parenteral antibiotic therapy. The impact of enteroviral infections of the central nervous system on hospital resources could be substantially reduced if a rapid, sensitive and specific laboratory method of diagnosing these infections were available.     

Influence of water quality on enteric virus concentration by microporous filter methods.

Four enteric viruses, poliovirus type 1, echovirus type 1, reovirus type 3, and simian adenovirus SV-11, were concentrated from seeded 1.3-liter volumes of raw, finished, and granular activated carbon-treated waters by adsorption to 47-mm-diameter (17 cm2), electropositive ( Virosorb 1MDS ) filters at pH 7.5 or electronegative ( Filterite ) filters at pH 3.5 with and without 5 mM added MgCl2, followed by elution with 0.3% beef extract in 50 mM glycine at pH 9.5. Removal of particulates from raw and finished waters by 0.2-micron prefiltration before virus addition and pH adjustment had little effect on virus concentration efficiencies. Soluble organic compounds reduced virus adsorption efficiencies from both raw and finished waters compared with granular activated carbon-treated water, but the extent of interference varied with virus type and adsorption conditions. For electropositive 1MDS filters, organic interference was similar with all virus types. For Filterite filters, organic interference was evident with poliovirus and echovirus, but could be overcome by adding MgCl2. Reovirus and SV-11 were not adversely affected by organics during adsorption to Filterite filters. Elution of reovirus and adenovirus was inefficient compared with that of poliovirus and echovirus. None of the three adsorption schemes ( 1MDS at pH 7.5 and Filterite with and without 5 mM MgCl2 at pH 3.5) could be judged superior for all viruses and water types tested.     

Influence of water quality on enteric virus concentration by microporous filter methods

Four enteric viruses, poliovirus type 1, echovirus type 1, reovirus type 3, and simian adenovirus SV-11, were concentrated from seeded 1.3-liter volumes of raw, finished, and granular activated carbon-treated waters by adsorption to 47-mm-diameter (17 cm2), electropositive ( Virosorb 1MDS ) filters at pH 7.5 or electronegative ( Filterite ) filters at pH 3.5 with and without 5 mM added MgCl2, followed by elution with 0.3% beef extract in 50 mM glycine at pH 9.5. Removal of particulates from raw and finished waters by 0.2-micron prefiltration before virus addition and pH adjustment had little effect on virus concentration efficiencies. Soluble organic compounds reduced virus adsorption efficiencies from both raw and finished waters compared with granular activated carbon-treated water, but the extent of interference varied with virus type and adsorption conditions. For electropositive 1MDS filters, organic interference was similar with all virus types. For Filterite filters, organic interference was evident with poliovirus and echovirus, but could be overcome by adding MgCl2. Reovirus and SV-11 were not adversely affected by organics during adsorption to Filterite filters. Elution of reovirus and adenovirus was inefficient compared with that of poliovirus and echovirus. None of the three adsorption schemes ( 1MDS at pH 7.5 and Filterite with and without 5 mM MgCl2 at pH 3.5) could be judged superior for all viruses and water types tested.     

Effect of Anticellular Serum on the Attachment of Enteroviruses to HeLa Cells

Anticellular serum (ACS), in the absence of an active complement system, was shown to inhibit the attachment of poliovirus types 1 and 2, echovirus type 6, and coxsackievirus types A13, B1, and B3 to viral receptors of live HeLa cells. This is the first report to provide evidence that ACS has an inhibitory effect on the interaction between host cells and coxsackieviruses of group B. The titer of inhibitory activity of ACS varied inversely with the cell concentration used, the reaction being virtually completed after an incubation period of 30 min at 37 C. The inhibitory activity of ACS persisted for more than 4 hr at 37 C, and was shown to be reversible at pH 2.0, revealing that although the receptors for attaching virus were inactivated by ACS the inactivation was not permanent. These findings are consistent with the concept that antibodies in the ACS combine with and blockade viral receptors located at the cell surface. An antiserum with a specificity for inhibiting attachment of coxsackievirus B1 was obtained by dual absorption of ACS with cells saturated with coxsackievirus type B3 and chymotrypsin-treated cells. These findings offer an approach whereby the antigenic relationship of viral receptors to other constituents of the cell surface can be studied.     

GIV noroviruses and other enteric viruses in bivalves: a preliminary study

We evaluated the presence of the enteric viruses: norovirus, adenovirus, enterovirus, astrovirus, hepatitis A virus, and hepatitis E virus in bivalves using nested PCR methods and cell culture assays. Noroviruses GII.4 and GIV.1, adenoviruses types 1 and 2, hepatitis A, and echovirus type 7 were detected in the shellfish tested, which were often co-infected. This is the first study to detect such a high level of viral contamination in Italian mussels (up to four different viral groups in a single sample), and the first to document the presence of GIV NoV in shellfish.     

Relationships between environmental factors, bacterial indicators, and the occurrence of enteric viruses in estuarine sediments

Current standards for evaluation of the public health safety of recreational and shellfish-harvesting waters are based upon bacteriological analysis, but do not include an evaluation of the number of viruses. The objective of this study was to determine the occurrence of enteric viruses in estuarine sediments and to find a relationship, if any, between the presence of viruses in seawater or sediment or both and various biological and physicochemical characteristics of the environment. Viruses were found in greater numbers in sediment than in overlying seawater on a volume basis. Several types of enteroviruses were isolated: coxsackievirus types A16, B1, and B5, echovirus type 1, and poliovirus type 2. On several occasions, viruses were isolated from sediments when overlying seawaters met bacteriological water quality standards for recreational use. Statistical analysis of the relationship between viruses in seawater or in sediment and other variables measured yielded only one significant association: the number of viruses in sediment was found to be positively correlated with the number of fecal coliforms in sediment. No other physical, chemical, or biological characteristic of seawater or sediment that was measured showed statistically significant association with viral numbers. No correlation was found between bacterial indicators and virus in the overlying waters. The data indicated that evaluation of the presence of bacteria and viruses in sediment may provide additional insight into long-term water quality conditions and that indicator bacteria in water are not reflective of the concentration of enteric viruses in marine waters.     

Relationships Between Environmental Factors, Bacterial Indicators, and the Occurrence of Enteric Viruses in Estuarine Sediments

Current standards for evaluation of the public health safety of recreational and shellfish-harvesting waters are based upon bacteriological analysis, but do not include an evaluation of the number of viruses. The objective of this study was to determine the occurrence of enteric viruses in estuarine sediments and to find a relationship, if any, between the presence of viruses in seawater or sediment or both and various biological and physicochemical characteristics of the environment. Viruses were found in greater numbers in sediment than in overlying seawater on a volume basis. Several types of enteroviruses were isolated: coxsackievirus types A16, B1, and B5, echovirus type 1, and poliovirus type 2. On several occasions, viruses were isolated from sediments when overlying seawaters met bacteriological water quality standards for recreational use. Statistical analysis of the relationship between viruses in seawater or in sediment and other variables measured yielded only one significant association: the number of viruses in sediment was found to be positively correlated with the number of fecal coliforms in sediment. No other physical, chemical, or biological characteristic of seawater or sediment that was measured showed statistically significant association with viral numbers. No correlation was found between bacterial indicators and virus in the overlying waters. The data indicated that evaluation of the presence of bacteria and viruses in sediment may provide additional insight into long-term water quality conditions and that indicator bacteria in water are not reflective of the concentration of enteric viruses in marine waters.     

Viral pollution of the rivers in Toyama City

Viral pollution of the river water in Toyama City was surveyed during the two-year period from July 1979 to July 1981, and the ecology of viruses in the river water is discussed. Virus isolation from the river water samples, or from the water squeezed from cotton pads that were immersed in the stream for 3 days, was carried out by the "filter adsorption/elution" method. River waters were found to be contaminated with various species of enteric viruses, that is, poliovirus, echovirus, coxsackievirus, adenovirus, and reovirus. Poliovirus was isolated during the period immediately after the oral administration of polio vaccine, and coxsackie B virus was frequently isolated all year around. The enterovirus concentration in the river water was significantly high with a maximum of five plaque-forming units of coxsackie B2 virus per 250 ml. The species and type distribution of enteroviruses isolated from the river water coincided well with that of viruses isolated from inhabitants of Toyama Prefecture, with the exception of reovirus which was the largest population of virus species in the river water.     

Detection of enteroviruses and mammalian reoviruses in Korean environmental waters

Using the standard total culturable virus assay-most probable number (TCVA-MPN) method, we evaluated a total of 348 samples, including surface water, finished water, and tap water samples, collected from randomly selected water treatment plants in Korea from August 2001 through July 2005 according to the Information Collection Rule. All the TCVA-positive samples were also subjected to integrated cell culture-PCR (ICC-PCR) methods for the detection of enteroviruses, hepatitis A virus, adenoviruses, and reoviruses. The most probable number of infectious units per 100 liters for the environmental water samples ranged from 0.5 to 47.3. Nine of the 13 TCVA-positive samples (69.2%) were found by ICC-PCR to be positive for human enteroviruses, which were confirmed to be coxsackievirus type B3, coxsackievirus type B4, coxsackievirus type B6, echovirus type 30, and vaccine strain poliovirus type 3 by direct sequencing. Eleven of the 13 TCVA-positive samples (84.6%) were found by ICC-PCR assay to be positive for reoviruses. The serotype of all the reoviruses was the same as reovirus type 1 by direct sequencing. Both enteroviruses and reoviruses were concurrently detected in seven TCVA-positive samples (53.8%).     

Inactivation of feline calicivirus and adenovirus type 40 by UV radiation

Little information regarding the effectiveness of UV radiation on the inactivation of caliciviruses and enteric adenoviruses is available. Analysis of human calicivirus resistance to disinfectants is hampered by the lack of animal or cell culture methods that can determine the viruses' infectivity. The inactivation kinetics of enteric adenovirus type 40 (AD40), coliphage MS-2, and feline calicivirus (FCV), closely related to the human caliciviruses based on nucleic acid organization and capsid architecture, were determined after exposure to low-pressure UV radiation in buffered demand-free (BDF) water at room temperature. In addition, UV disinfection experiments were also carried out in treated groundwater with FCV and AD40. AD40 was more resistant than either FCV or coliphage MS-2 in both BDF water and groundwater. The doses of UV required to achieve 99% inactivation of AD40, coliphage MS-2, and FCV in BDF water were 109, 55, and 16 mJ/cm(2), respectively. The doses of UV required to achieve 99% inactivation of AD40, coliphage MS-2, and FCV in groundwater were slightly lower than those in BDF water. FCV was inactivated by 99% by 13 mJ/cm(2) in treated groundwater. A dose of 103 mJ/cm(2) was required for 99% inactivation of AD40 in treated groundwater. The results of this study indicate that if FCV is an adequate surrogate for human caliciviruses, then their inactivation by UV radiation is similar to those of other single-stranded RNA enteric viruses, such as poliovirus. In addition, AD40 appears to be more resistant to UV disinfection than previously reported.     

Enhancement of enterovirus infectivity in vitro by pretreating host cell monolayers with the cationic polymer polyethyleneimine.

Laboratory strains of enteroviruses, as well as viruses isolated from raw wastewater, were found to exhibit enhanced infectivity in vitro when BGM cell monolayers were pretreated with the cationic polymer polyethyleneimine (PEI). Viruses were assayed by the cytopathic effect technique and as PFU under methylcellulose and agar overlays with monolayers treated with 0 to 5.0 x 10(-3)% (wt/vol) PEI in phosphate-buffered saline supplemented with 2% fetal bovine serum. Poliovirus type 1 cytopathic effect occurred at an enhanced rate in cells treated with 5.0 x 10(-3)% PEI compared with untreated cells. PEI-treated cells were found to adsorb viruses much more effectively than untreated cells did. When the methylcellulose overlay procedure was used, rates of infectivity were enhanced as follows: poliovirus type 1, 5.5-fold; echovirus type 1, 1.2-fold; echovirus type 5, 5.2-fold; and coxsackievirus type B5, 4.9-fold. Viruses concentrated from raw wastewater showed a 3.8-fold increase in titer when quantitated by the most-probable-number method and a 3.3-fold increase when quantitated as PFU under an agar overlay.     

Enhancement of enterovirus infectivity in vitro by pretreating host cell monolayers with the cationic polymer polyethyleneimine

Laboratory strains of enteroviruses, as well as viruses isolated from raw wastewater, were found to exhibit enhanced infectivity in vitro when BGM cell monolayers were pretreated with the cationic polymer polyethyleneimine (PEI). Viruses were assayed by the cytopathic effect technique and as PFU under methylcellulose and agar overlays with monolayers treated with 0 to 5.0 x 10(-3)% (wt/vol) PEI in phosphate-buffered saline supplemented with 2% fetal bovine serum. Poliovirus type 1 cytopathic effect occurred at an enhanced rate in cells treated with 5.0 x 10(-3)% PEI compared with untreated cells. PEI-treated cells were found to adsorb viruses much more effectively than untreated cells did. When the methylcellulose overlay procedure was used, rates of infectivity were enhanced as follows: poliovirus type 1, 5.5-fold; echovirus type 1, 1.2-fold; echovirus type 5, 5.2-fold; and coxsackievirus type B5, 4.9-fold. Viruses concentrated from raw wastewater showed a 3.8-fold increase in titer when quantitated by the most-probable-number method and a 3.3-fold increase when quantitated as PFU under an agar overlay.     

New method using a positively charged microporous filter and ultrafiltration for concentration of viruses from tap water

The methods used to concentrate enteric viruses from water have remained largely unchanged for nearly 30 years, with the most common technique being the use of 1MDS Virozorb filters followed by organic flocculation for secondary concentration. Recently, a few studies have investigated alternatives; however, many of these methods are impractical for use in the field or share some of the limitations of this traditional method. In the present study, the NanoCeram virus sampler, an electropositive pleated microporous filter composed of microglass filaments coated with nanoalumina fibers, was evaluated. Test viruses were first concentrated by passage of 20 liters of seeded water through the filter (average filter retention efficiency was ≥ 99.8%), and then the viruses were recovered using various salt-based or proteinaceous eluting solutions. A 1.0% sodium polyphosphate solution with 0.05 M glycine was determined to be the most effective. The recovered viruses were then further concentrated using Centricon Plus-70 centrifugal ultrafilters to a final volume of 3.3 (±0.3 [standard deviation]) ml; this volume compares quite favorably to that of previously described methods, such as organic flocculation (~15 to 40 ml). The overall virus recovery efficiencies were 66% for poliovirus 1, 83% for echovirus 1, 77% for coxsackievirus B5, 14% for adenovirus 2, and 56% for MS2 coliphage. In addition, this method appears to be compatible with both cell culture and PCR assays. This new approach for the recovery of viruses from water is therefore a viable alternative to currently used methods when small volumes of final concentrate are an advantage.     

Inactivation of Feline Calicivirus and Adenovirus Type 40 by UV Radiation

Little information regarding the effectiveness of UV radiation on the inactivation of caliciviruses and enteric adenoviruses is available. Analysis of human calicivirus resistance to disinfectants is hampered by the lack of animal or cell culture methods that can determine the viruses' infectivity. The inactivation kinetics of enteric adenovirus type 40 (AD40), coliphage MS-2, and feline calicivirus (FCV), closely related to the human caliciviruses based on nucleic acid organization and capsid architecture, were determined after exposure to low-pressure UV radiation in buffered demand-free (BDF) water at room temperature. In addition, UV disinfection experiments were also carried out in treated groundwater with FCV and AD40. AD40 was more resistant than either FCV or coliphage MS-2 in both BDF water and groundwater. The doses of UV required to achieve 99% inactivation of AD40, coliphage MS-2, and FCV in BDF water were 109, 55, and 16 mJ/cm², respectively. The doses of UV required to achieve 99% inactivation of AD40, coliphage MS-2, and FCV in groundwater were slightly lower than those in BDF water. FCV was inactivated by 99% by 13 mJ/cm² in treated groundwater. A dose of 103 mJ/cm² was required for 99% inactivation of AD40 in treated groundwater. The results of this study indicate that if FCV is an adequate surrogate for human caliciviruses, then their inactivation by UV radiation is similar to those of other single-stranded RNA enteric viruses, such as poliovirus. In addition, AD40 appears to be more resistant to UV disinfection than previously reported.     

Aseptic meningitis,Montreal, 1969: A Clinical and Laboratory Study

During the summer and fall of 1969 an outbreak of aseptic meningitis occurred in Montreal and its environs. Forty-four patients were admitted to the infectious disease ward of The Montreal Children''s Hospital in August, September and October. Half of the patients were in the age group 6 to 10 years. The ratio of males to females was two to one. Patients showed the typical signs and symptoms of aseptic meningitis, namely fever, vomiting, headache, neck stiffness and pleocytosis of the cerebrospinal fluid.Viruses were recovered from 19 (43%) of the 44 cases. The predominant virus in the outbreak was a non-hemagglutinating strain of echovirus 6. Other virus types isolated in the outbreak were echovirus 7, coxsackievirus A9 and coxsackievirus B4. Serological investigations confirmed the validity of the echovirus isolations and identified additional cases.     

Removal of Enteroviruses from Sewage by Bench-Scale Rotary-Tube Trickling Filters

The efficacy of a rotary-tube type of trickling filter for removing coxsackievirus A9, poliovirus 1, and echovirus 12 suspended in raw settled sewage was investigated. At filtration rates equivalent to about 10 MGD (million gallons per day)/acre (ca. 3,785 m3/day per acre), the filters removed 95% of the poliovirus, 83% of echovirus 12, and 94% of coxsackievirus A9. Coliform, fecal streptococci, biochemical oxygen demand, and chemical oxygen demand removals were remarkably similar, averaging 94, 92, 93, and 95%, respectively. At filtration rates equivalent to about 23 MGD/acre, 59% of the poliovirus, 63% of the echovirus 23, and 81% of the coxsackievirus A9 were removed. Coliform, fecal streptococci, biochemical oxygen demand, and chemical oxygen demand removals at this filtration rate were 68, 75, 72, and 56%, respectively. Viruses were assumed to be adsorbed to the biological slime growing in the filters, but attempts to disassociate the viruses from the slime were unsuccessful, indicating that the slime-virus complex is very stable or that the viruses were somehow inactivated. The data indicate that coliform and fecal streptococci reductions in this type sewage treatment process can be used as an index of virus reduction. Disinfection, however, must be used to ensure a virus-free final effluent.