Comparison of the airborne survival of calf rotavirus and poliovirus type 1 (Sabin) aerosolized as a mixture.

A mixture of a cell culture-adapted strain (C-486) of calf rotavirus and poliovirus type 1 (Sabin) was prepared in tryptose phosphate broth containing 0.1% uranine (physical tracer) and antifoam at a final concentration of 0.001%. By using a six-jet Collison nebulizer, the mixture was aerosolized into a 300-liter stainless-steel rotating (4 rpm) drum. The temperature of the air inside the drum was kept at 20 +/- 1 degrees C, and the virus aerosols were held at the following three levels of relative humidity (RH): low (30 +/- 5%), medium (50 +/- 5%), and high (80 +/- 5%). An all-glass impinger, containing 10.0 ml of tryptose phosphate broth with antifoam, was used to collect samples of air from the drum. Both viruses were propagated and quantitated in MA-104 cells. The calf rotavirus was found to survive well at mid-range RH, where 60% of the infectious virus could be detected even after 24 h of virus aerosolization. At the low RH, the half-life of the infectious rotavirus was ca. 14 h. On the other hand, no infectious poliovirus could be recovered from the drum air at the low and medium RH. At the high RH, more than 50% of the infectious rotavirus became undetectable within 90 min of aerosolization. In contrast to this, the half-life of the poliovirus at the high RH was about 10 h. These data, based on the aerosolization of virus mixtures, therefore suggest that there is a pronounced difference in the way RH influences the airborne survival of these two types of viruses held under identical experimental conditions.     

Effect of relative humidity on the airborne survival of rotavirus SA11.

Rotavirus SA11, suspended in tryptose phosphate broth with 2.5 mg of rhodamine B per ml, was aerosolized (Collison nebulizer) into a rotating drum, and the aerosols were held at 20 +/- 1 degree C with the desired relative humidity (RH). An all-glass impinger with tryptose phosphate broth was used to collect 1-min (5.6-liter) samples of air from the drum. The virus was found to survive best at medium (50 +/- 5%) RH, where its half-life was nearly 40 h. The half-life of the virus at the low (25 +/- 5%) RH level was about 9 h. Even at 72 h of aerosol age, 45 and 21% of the infectious virus remained detectable in the air at the medium and low RH levels, respectively. The high (80 +/- 5%) RH level was found to be the least favorable to the survival of the virus, since 50% of the infectious virus became undetectable within 2 h of aerosolization. In a separate experiment at the midrange RH, 3% of the infectious virus was detectable in the drum air after 223 h (9 days) of aerosol age. Rotaviruses could, therefore, survive in air for prolonged periods, thus making air a possible vehicle for their dissemination.     

Effect of relative humidity on the airborne survival of rotavirus SA11

Rotavirus SA11, suspended in tryptose phosphate broth with 2.5 mg of rhodamine B per ml, was aerosolized (Collison nebulizer) into a rotating drum, and the aerosols were held at 20 +/- 1 degree C with the desired relative humidity (RH). An all-glass impinger with tryptose phosphate broth was used to collect 1-min (5.6-liter) samples of air from the drum. The virus was found to survive best at medium (50 +/- 5%) RH, where its half-life was nearly 40 h. The half-life of the virus at the low (25 +/- 5%) RH level was about 9 h. Even at 72 h of aerosol age, 45 and 21% of the infectious virus remained detectable in the air at the medium and low RH levels, respectively. The high (80 +/- 5%) RH level was found to be the least favorable to the survival of the virus, since 50% of the infectious virus became undetectable within 2 h of aerosolization. In a separate experiment at the midrange RH, 3% of the infectious virus was detectable in the drum air after 223 h (9 days) of aerosol age. Rotaviruses could, therefore, survive in air for prolonged periods, thus making air a possible vehicle for their dissemination.     

Effect of relative humidity on the airborne survival of rhinovirus-14

Rhinovirus-14, suspended in tryptose phosphate broth supplemented with uranine (physical tracer) and an antifoam, was aerosolized by use of a Collison nebulizer. The aerosols were held in a rotating drum with the relative humidity at either the low (30 +/- 5%), medium (50 +/- 5%), or high (80 +/- 5%) level at 20 +/- 1 degrees C. An all-glass impinger was used to recover the virus from the air in the drum, with the first air sample being collected after a 15-min period of aerosol stabilization. Subsequent air samples were withdrawn at 2, 4, 8, and 14 h after stabilization of the aerosol. At the low and medium relative humidity levels, the infectivity of the airborne virus was rapidly lost and less than 0.25% could be detected in the first air sample. At the high RH level, however, the airborne virus had a half-life of 13.7 +/- 1.91 h and nearly 30% of the input infectious virus could be detected in the drum air even after 24 h of aerosolization. These findings suggest that under certain environmental conditions, notably high relative humidity, air may act as a vehicle for the spread of rhinovirus infections.     

Effect of relative humidity, atmospheric temperature, and suspending medium on the airborne survival of human rotavirus

The Wa strain of human rotavirus, grown in MA-104 cells, was suspended either in tryptose phosphate broth or feces from a case of rotaviral diarrhea. It was then aerosolized into a rotating drum using a Collison nebulizer. The drum air was sampled using an all-glass impinger containing tryptose phosphate broth as collecting fluid. At 20 +/- 1 degree C, the virus aerosolized from tryptose phosphate broth was found to survive best at 50 +/- 5% relative humidity, where its half-life was 44.2 +/- 6.3 h. At 30 +/- 5% and 80 +/- 5% relative humidity, the half-life of the virus was 24.5 +/- 3.5 and 3.8 +/- 1.0 h, respectively. At 6 +/- 1 degree C, the airborne survival of the virus at the mid and low relative humidity levels was further enhanced, but at the high relative humidity it remained very similar to that seen at 20 +/- 1 degree C. When aerosols of fecally suspended human rotavirus were held at 20 +/- 1 degree C with 50 +/- 5% relative humidity, nearly 80% of the airborne virus particles remained infectious even at the aerosol age of 24 h. These findings may help in our understanding of the epidemiology of rotaviral infections.     

Survival of human rhinovirus type 14 dried onto nonporous inanimate surfaces: effect of relative humidity and suspending medium

To study the survival of human rhinovirus 14 on environmental surfaces, each stainless steel disk (1 cm in diameter) was contaminated with 10 microL (about 10(5) plaque-forming units) of the virus suspended in either 1 chi tryptose phosphate broth (TPB), 5 mg/mL of bovine mucin in normal saline, or undiluted human nasal discharge. The inoculum was dried in a laminar flow cabinet for 1 h under ambient conditions. The disks were then placed in a glass chamber (20 +/- 1 degree C) with the relative humidity at either low (20 +/- 5%), medium (50 +/- 5%), or high (80 +/- 5%) level. At appropriate intervals, the disk to be tested was placed in 1 mL of tryptose phosphate broth and the eluate titrated in A-5 HeLa cells. When the virus was suspended in either tryptose phosphate broth, mucin, or the nasal discharge and subjected to initial drying, there was a 3.0 +/- 1.0, 82.0 +/- 6.7, and 89.0 +/- 3.0% loss in virus infectivity, respectively. The half-life of the TPB-suspended virus was about 14 h at the high relative humidity as compared with less than 2 h at the other two relative humidity levels. The half-lives for the mucin-suspended virus at the high, medium, and low relative humidity were 1.42, 0.55, and 0.24 h, respectively; the corresponding values for the nasal discharge suspended virus being 0.17, 0.25, and 0.09 h.     

Effect of relative humidity and air temperature on survival of hepatitis A virus on environmental surfaces.

Stainless steel disks (diameter, 1 cm) were contaminated with fecally suspended hepatitis A virus (HAV; strain HM-175) and held at low (25% +/- 5%), medium (55% +/- 5%), high (80% +/- 5%), or ultrahigh (95% +/- 5%) relative humidity (RH) at an air temperature of 5,20, or 35 degrees C. HAV survival was inversely proportional to the level of RH and temperature, and the half-lives of the virus ranged from greater than 7 days at the low RH and 5 degrees C to about 2 h at the ultrahigh RH and 35 degrees C. In parallel tests with fecally suspended Sabin poliovirus (PV) type 1 at the low and ultrahigh RH, all PV activity was lost within 4 h at the low RH whereas at the ultrahigh RH it remained detectable up to 12 h. HAV could therefore survive much better than PV on nonporous environmental surfaces. Moreover, the ability of HAV to survive better at low levels of RH is in direct contrast to the behavior of other enteroviruses. These findings should help in understanding the genesis of HAV outbreaks more clearly and in designing better measures for their control and prevention.     

Effect of relative humidity and air temperature on survival of hepatitis A virus on environmental surfaces.

Stainless steel disks (diameter, 1 cm) were contaminated with fecally suspended hepatitis A virus (HAV; strain HM-175) and held at low (25% +/- 5%), medium (55% +/- 5%), high (80% +/- 5%), or ultrahigh (95% +/- 5%) relative humidity (RH) at an air temperature of 5,20, or 35 degrees C. HAV survival was inversely proportional to the level of RH and temperature, and the half-lives of the virus ranged from greater than 7 days at the low RH and 5 degrees C to about 2 h at the ultrahigh RH and 35 degrees C. In parallel tests with fecally suspended Sabin poliovirus (PV) type 1 at the low and ultrahigh RH, all PV activity was lost within 4 h at the low RH whereas at the ultrahigh RH it remained detectable up to 12 h. HAV could therefore survive much better than PV on nonporous environmental surfaces. Moreover, the ability of HAV to survive better at low levels of RH is in direct contrast to the behavior of other enteroviruses. These findings should help in understanding the genesis of HAV outbreaks more clearly and in designing better measures for their control and prevention.     

Comparison of bacteriophage and enteric virus removal in pilot scale activated sludge plants

AIMS: The aim of this experimental study was to determine comparatively the removal of two types of bacteriophages, a somatic coliphage and an F-specific RNA phage and of three types of enteric viruses, hepatitis A virus (HAV), poliovirus and rotavirus during sewage treatment by activated sludge using laboratory pilot plants. METHODS AND RESULTS: The cultivable simian rotavirus SA11, the HAV HM 175/18f cytopathic strain and poliovirus were quantified by cell culture. The bacteriophages were quantified by plaque formation on the host bacterium in agar medium. In each experiment, two pilots simulating full-scale activated sludge plants were inoculated with viruses at known concentrations, and mixed liquor and effluent samples were analysed regularly. In the mixed liquor, liquid and solid fractions were analysed separately. The viral behaviour in both the liquid and solid phases was similar between pilots of each experiment. Viral concentrations decreased rapidly following viral injection in the pilots. Ten minutes after the injections, viral concentrations in the liquid phase had decreased from 1.0 +/- 0.4 log to 2.2 +/- 0.3 log. Poliovirus and HAV were predominantly adsorbed on the solid matters of the mixed liquor while rotavirus was not detectable in the solid phase. In our model, the estimated mean log viral reductions after 3-day experiment were 9.2 +/- 0.4 for rotavirus, 6.6 +/- 2.4 for poliovirus, 5.9 +/- 3.5 for HAV, 3.2 +/- 1.2 for MS2 and 2.3 +/- 0.5 for PhiX174. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates that the pilots are useful models to assess the removal of infectious enteric viruses and bacteriophages by activated sludge treatment. Our results show the efficacy of the activated sludge treatment on the five viruses and suggest that coliphages could be an acceptable indicator of viral removal in this treatment system.     

Development of a method for detection of human rotavirus in water and sewage

The simian rotavirus SA11 was used to develop a simple, reliable, and efficient method to concentrate rotavirus from tap water, treated sewage, and raw sewage by absorption to and elution from Filterite fiberglass-epoxy filters. SA11 adsorbed optimally to Filterite filters from water containing 0.5 mM AlCl3 at pH 3.5. Filter-bound virus was eluted with 0.05 M glycine-NaOH supplemented with 10% tryptose phosphate broth at pH 10. SA11 was quantitated by plaque assay, whereas human rotavirus was detected by immunofluorescence. The method was applied to detect rotavirus in raw and treated sewage at two Houston, Tex., sewage treatment plants. The sewage isolates were identified as rotavirus, probably a human strain, based on several criteria. The sewage isolates were detectable by an immunofluorescence test, using anti-SA11 serum which would detect the simian, human bovine, and porcine rotaviruses. No reaction was noted by immunofluorescence with the reoviruses or several common enteroviruses. The sewage isolates were neutralized by convalescent sera from a human adult and infant who had been infected by rotavirus as well as by a hyperimmune serum prepared in guinea pigs against purified human rotavirus. Preimmune or preillness sera did not react with the isolates by neutralization or immunofluorescence. The natural isolates were sensitive to pH 11 and other inactivating agents, similar to SA11. The buoyant density of the sewage isolates in CsCl gradients was 1.36 g/cm3, which is the value usually reported for complete, infectious rotavirus particles. The double-shelled particle diameter was 67.1 +/- 2.4 nm. Finally, electron micrographs of cell lysates inoculated with the sewage isolate showed particles displaying characteristic rotavirus morphology.     

Development of a method for detection of human rotavirus in water and sewage.

The simian rotavirus SA11 was used to develop a simple, reliable, and efficient method to concentrate rotavirus from tap water, treated sewage, and raw sewage by absorption to and elution from Filterite fiberglass-epoxy filters. SA11 adsorbed optimally to Filterite filters from water containing 0.5 mM AlCl3 at pH 3.5. Filter-bound virus was eluted with 0.05 M glycine-NaOH supplemented with 10% tryptose phosphate broth at pH 10. SA11 was quantitated by plaque assay, whereas human rotavirus was detected by immunofluorescence. The method was applied to detect rotavirus in raw and treated sewage at two Houston, Tex., sewage treatment plants. The sewage isolates were identified as rotavirus, probably a human strain, based on several criteria. The sewage isolates were detectable by an immunofluorescence test, using anti-SA11 serum which would detect the simian, human bovine, and porcine rotaviruses. No reaction was noted by immunofluorescence with the reoviruses or several common enteroviruses. The sewage isolates were neutralized by convalescent sera from a human adult and infant who had been infected by rotavirus as well as by a hyperimmune serum prepared in guinea pigs against purified human rotavirus. Preimmune or preillness sera did not react with the isolates by neutralization or immunofluorescence. The natural isolates were sensitive to pH 11 and other inactivating agents, similar to SA11. The buoyant density of the sewage isolates in CsCl gradients was 1.36 g/cm3, which is the value usually reported for complete, infectious rotavirus particles. The double-shelled particle diameter was 67.1 +/- 2.4 nm. Finally, electron micrographs of cell lysates inoculated with the sewage isolate showed particles displaying characteristic rotavirus morphology.     

Effect of Temperature and Relative Humidity on Survival of Airborne Columbia SK Group Viruses

Three strains of the Columbia SK (Col-SK) group of viruses [Mengo, Maus Elberfeld (ME), and Col-SK viruses] have been studied in the airborne state. All three strains were found to give identical aerosol decay patterns at 16 or 26 C, when held at the same relative humidity (RH). During the first 5 min of aerosol storage time at 16 C, virus inactivation was RH-dependent, with survival maximal at either high (greater than 80%) or low (less than 5%) RH. After 5 min at 16 C, further inactivation, regardless of RH, was insignificant. At 26 C, the effect on survival of RH between 40 and 60% was even more pronounced than at 16 C, and continued after 5 min through 6 hr. Results of this study indicated that the inactivation of airborne Col-SK group viruses was similar to that of other ribonucleic acid (RNA) viruses, particularly poliovirus. Since members of the Col-SK group are picornaviruses, they may well serve as an aerosol model representative of small, ether-resistant, single-stranded RNA viruses.     

Effects of temperature, relative humidity, absolute humidity, and evaporation potential on survival of airborne Gumboro vaccine virus

Survival of airborne virus influences the extent of disease transmission via air. How environmental factors affect viral survival is not fully understood. We investigated the survival of a vaccine strain of Gumboro virus which was aerosolized at three temperatures (10°C, 20°C, and 30°C) and two relative humidities (RHs) (40% and 70%). The response of viral survival to four metrics (temperature, RH, absolute humidity [AH], and evaporation potential [EP]) was examined. The results show a biphasic viral survival at 10°C and 20°C, i.e., a rapid initial inactivation in a short period (2.3 min) during and after aerosolization, followed by a slow secondary inactivation during a 20-min period after aerosolization. The initial decays of aerosolized virus at 10°C (1.68 to 3.03 ln % min(-1)) and 20°C (3.05 to 3.62 ln % min(-1)) were significantly lower than those at 30°C (5.67 to 5.96 ln % min(-1)). The secondary decays at 10°C (0.03 to 0.09 ln % min(-1)) tended to be higher than those at 20°C (-0.01 to 0.01 ln % min(-1)). The initial viral survival responded to temperature and RH and potentially to EP; the secondary viral survival responded to temperature and potentially to RH. In both phases, survival of the virus was not significantly affected by AH. These findings suggest that long-distance transmission of airborne virus is more likely to occur at 20°C than at 10°C or 30°C and that current Gumboro vaccination by wet aerosolization in poultry industry is not very effective due to the fast initial decay.     

Rotavirus concentration from cell culture harvests: trypsin treatment followed by hydroextraction.

An 80- to 150-ml amount of calf or simian rotavirus-containing cell culture harvests of MA-104 cells were treated with 50 microgram of trypsin per ml and hydroextracted overnight (4 degrees C) with polyethylene glycol 6,000. The concentrate was resuspended in 8 to 10 ml of tryptose phosphate broth and plaque assayed. Between 85 and 97% of the input virus could be recovered with a concentration of up to 15-fold.     

Survival of enteric viruses on environmental fomites.

The survival of human enteric viruses on several porous (paper and cotton cloth) and nonporous (aluminum, china, glazed tile, latex, and polystyrene) environmental surfaces has been evaluated. Viruses persisted for extended periods on several types of materials commonly found in institutions and domestic environments. The stability of the viruses was generally influenced by environmental factors such as relative humidity (RH), temperature, and the type of surface contaminated. Overall, hepatitis A virus (HAV) and human rotavirus (HRV) were more resistant to inactivation than enteric adenovirus (ADV) and poliovirus (PV). The resistance to the desiccation step appears to be of major significance in determining the survival of a virus dried on fomites. ADV and PV showed a pronounced decrease in titer at this stage, whereas HAV and HRV displayed little decay at the desiccation step. HAV and HRV persistence was not affected by the presence of fecal material. On nonporous surfaces, PV and ADV persisted better in the presence of feces. However, on porous fomites the presence of fecal material had a negative influence on the survival of PV and ADV. Except for HRV, greater virus survival was observed at 4 degrees than at 20 degrees C. PV and HAV survival was enhanced at high RH; the survival of the latter was enhanced at least for nonporous materials. When dried on porous materials, HRV also exhibited greater persistence at high RH. The survival of ADV was not affected by RH. The validity of using bacteriophages of Bacteroides fragilis as indicators of human viruses dried on fomites was evaluated.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolation of enteroviruses from water, suspended solids, and sediments from Galveston Bay: survival of poliovirus and rotavirus adsorbed to sediments.

The distribution and quantitation of enteroviruses among water, suspended solids, and compact sediments in a polluted estuary are described. Samples were collected sequentially from water, suspended solids, fluffy sediments (uppermost layer of bottom sediments), and compact sediment. A total of 103 samples were examined of which 27 (26%) were positive for virus. Polioviruses were recovered most often, followed by coxsackie B viruses and echoviruses 7 and 29. Virus was found most often attached to suspended solids: 72% of these samples were positive, whereas only 14% of water samples without solids yielded virus. Fluffy sediments yielded virus in 47% of the samples, whereas only 5% of compact bottom-sediment samples were positive. When associated with solids, poliovirus and rotavirus retained their infectious quality for 19 days. The same viruses remained infectious for only 9 days when freely suspended in seawater. Collection of suspended solids at ambient water pH appears to be very useful for the detection of virus; it has advantages over collecting and processing large volumes of water, with accompanying pH adjustment and salt addition for processing.     

Isolation of enteroviruses from water, suspended solids, and sediments from Galveston Bay: survival of poliovirus and rotavirus adsorbed to sediments

The distribution and quantitation of enteroviruses among water, suspended solids, and compact sediments in a polluted estuary are described. Samples were collected sequentially from water, suspended solids, fluffy sediments (uppermost layer of bottom sediments), and compact sediment. A total of 103 samples were examined of which 27 (26%) were positive for virus. Polioviruses were recovered most often, followed by coxsackie B viruses and echoviruses 7 and 29. Virus was found most often attached to suspended solids: 72% of these samples were positive, whereas only 14% of water samples without solids yielded virus. Fluffy sediments yielded virus in 47% of the samples, whereas only 5% of compact bottom-sediment samples were positive. When associated with solids, poliovirus and rotavirus retained their infectious quality for 19 days. The same viruses remained infectious for only 9 days when freely suspended in seawater. Collection of suspended solids at ambient water pH appears to be very useful for the detection of virus; it has advantages over collecting and processing large volumes of water, with accompanying pH adjustment and salt addition for processing.     

Mechanism of Resistance to Enteroviruses of Some Primate Cells in Tissue Culture

Two cell lines, M10-45-2 and L-41, were studied, each of which possessed specific resistance either to poliovirus or to coxsackievirus. Infection of M10-45-2 cells with poliovirus ribonucleic acid (RNA) and L-41 cells with infectious coxsackievirus RNA was accompanied by production of complete viruses in each of the resistant cell lines. During incubation of the cells with the virus to which they were resistant, the amount of infectious virus did not decrease. Treatment with glycine-HCl buffer solution (pH 2.5) of resistant M10-45-2 cells after incubation with poliovirus at 0 C did not result in recovery of infectious virus, although such release did take place after treatment of sensitive M10 cells. Infection of resistant cells with virus containing poliovirus RNA and coxsackievirus proteins resulted in production of poliovirus in M10-45-2 cells but not in L-41 cells. The resistant cells are apparently unable to adsorb the virus to which they are resistant.     

Effect of poly(I).poly(C12U) (Ampligen) on enteric virus (rotavirus, poliovirus and Coxsackie B3 virus) infections

The effects of poly(I) poly(C₁₂U) (Ampligen) on infections with enteric viruses (rotavirus, poliovirus and Coxsackie B3 virus) were studied in vitro. Ampligen exhibited antiviral activity against rotavirus, especially when treatment was performed prior to inoculation of the virus. It was partially effective against Coxsackie B3 virus, but not against poliovirus. It is suggested that the observed effects may be due to the production of interferon induced by Ampligen.     

Rotavirus survival in conventionally treated drinking water

Samples of conventionally treated drinking water collected either as effluent (PE) at a treatment plant or out of a tap (TW) in our laboratory were seeded with simian rotavirus SA-11, which closely resembles rotavirus of human origin. The virus, grown in MA-104 cells, was suspended either in distilled water, Earle's balanced salt solution (EBSS), or tryptose phosphate broth (TPB), and added to the water samples to a final concentration of 5.7 X 10(3) plaque-forming units (PFU) per millilitre. After a contact time of 1 h at 22 degrees C, the samples were diluted and plaque assayed. There was no significant reduction in the virus titre in samples of TW (less than 0.05 mg/L free chlorine). The titre also remained almost the same in PE (0.75 mg/L free chlorine) when EBSS or TPB was used for virus suspension. There was, however, nearly a 1 log10 loss in the titre of the virus when it was suspended in distilled water before the contamination of PE. To study the long-term survival of the rotavirus in TW, the inoculated samples (5.0 X 10(4) PFU/mL) were held at either 4 or 20 degrees C in the dark and tested over a period of 64 days. At 20 degrees C it took 64 days to reduce the virus titre by 2 log10, whereas at 4 degrees C the virus titre dropped only 0.7 log10 during the same period. Rotaviruses could, therefore, survive well enough in conventionally treated drinking water to make it a possible vehicle for their transmission.