Elimination of viruses and indicator bacteria at each step of treatment during preparation of drinking water at seven water treatment plants.

Seven drinking water treatment plants were sampled twice a month for 12 months to evaluate the removal of indicator bacteria and cytopathogenic enteric viruses. Samples were obtained at each level of treatment: raw water, postchlorination, postsedimentation, postfiltration, postozonation, and finished (tap) water. Raw water quality was usually poor, with total coliform counts exceeding 105 to 106 CFU/liter and the average virus count in raw water of 3.3 most probable number of cytopathogenic units (MPNCU)/liter; several samples contained more than 100 MPNCU/liter. All plants distributed finished water that was essentially free of indicator bacteria as judged by analysis of 1 liter for total coliforms, fecal coliforms, fecal streptococci, coagulase-positive staphylococci, and Pseudomonas aeruginosa. The total plate counts at 20 and 35 degrees C were also evaluated as a measure of the total microbial population and were usually very low. Viruses were detected in 7% (11 of 155) of the finished water samples (1,000 liters) at an average density of 0.0006 MPNCU/liter the highest virus density measured being 0.2 MPNCU/liter. The average cumulative virus reduction was 95.15% after sedimentation and 99.97% after filtration and did not significantly decrease after ozonation or final chlorination. The viruses isolated from treated waters were all enteroviruses: poliovirus types 1, 2, and 3, coxsackievirus types B3, B4, and B5, echovirus type 7, and untyped picornaviruses.     

Elimination of viruses and indicator bacteria at each step of treatment during preparation of drinking water at seven water treatment plants

Seven drinking water treatment plants were sampled twice a month for 12 months to evaluate the removal of indicator bacteria and cytopathogenic enteric viruses. Samples were obtained at each level of treatment: raw water, postchlorination, postsedimentation, postfiltration, postozonation, and finished (tap) water. Raw water quality was usually poor, with total coliform counts exceeding 105 to 106 CFU/liter and the average virus count in raw water of 3.3 most probable number of cytopathogenic units (MPNCU)/liter; several samples contained more than 100 MPNCU/liter. All plants distributed finished water that was essentially free of indicator bacteria as judged by analysis of 1 liter for total coliforms, fecal coliforms, fecal streptococci, coagulase-positive staphylococci, and Pseudomonas aeruginosa. The total plate counts at 20 and 35 degrees C were also evaluated as a measure of the total microbial population and were usually very low. Viruses were detected in 7% (11 of 155) of the finished water samples (1,000 liters) at an average density of 0.0006 MPNCU/liter the highest virus density measured being 0.2 MPNCU/liter. The average cumulative virus reduction was 95.15% after sedimentation and 99.97% after filtration and did not significantly decrease after ozonation or final chlorination. The viruses isolated from treated waters were all enteroviruses: poliovirus types 1, 2, and 3, coxsackievirus types B3, B4, and B5, echovirus type 7, and untyped picornaviruses.     

Isolation of viruses from drinking water at the Point-Viau water treatment plant

Viruses were isolated from every sample of raw (100 L) and treated (1000 L) water collected at a water treatment plant drawing sewage-contaminated river water. Few plaque-forming isolates were formed but cytopathogenic viruses were isolated as frequently in drinking water as in raw water. In drinking water some samples contained more than 1 cytopathogenic unit per litre, but most contained 1-10/100 L. These viruses had not been inactivated or removed by prechlorination, flocculation, filtration, ozonation, and postchlorination. There were no coliforms present and a residual chlorine level had been maintained. Poliovirus type 1 was a frequent isolate but many isolates were nonpoliovirus. The presence of these viruses in drinking water raises questions about the efficacy of some water treatment processes to remove viruses from polluted water.     

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.     

Detection and quantitation of human enteric viruses in waste waters: increased sensitivity using a human immune serum globulin--immunoperoxidase assay on MA-104 cells

This study demonstrates that the most sensitive method for the detection and quantitation of cultivable human enteric viruses in water samples after repassage in the MA-104 cell line is the detection of infected cells by the human immune serum globulin--immunoperoxidase (HISG-IP) method recently described by the authors. This immunoperoxidase method is up to 50 times more sensitive than a liquid overlay assay by cytopathic effect in BGM cells. The viral content of waste waters was evaluated with this new methodology. By this method the average viral content of raw sewage (RS) was 900 mpniu/L (most probable number of infectious units per litre), 1056 mpniu/L in primary effluent (PE), and 106 mpniu/L in secondary effluent (SE). With a cytopathic effect assay on BGM cells, values of 85 (RS), 56 (PE), and 2 (SE) mpniu/L were observed, a striking underestimation of the viral content of secondary effluents.     

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.     

Viral pollution of surface waters due to chlorinated primary effluents.

The role of chlorinated primary effluents in viral pollution of the Ottawa River (Ontario) was assessed by examining 282 field samples of wastewaters from two different sewage treatment plants over a 2-year period. The talc-Celite technique was used for sample concentration, and BS-C-1 cells were employed for virus detection. Viruses were detected in 80% (75/94) of raw sewage, 72% (68/94) of primary effluent, and 56% (53/94) of chlorinated effluent samples. Both raw sewage and primary effluent samples contained about 100 viral infective units (VIU) per 100 ml. Chlorination produced a 10- to 50-fold reduction in VIU and gave nearly 2.7 VIU/100 ml of chlorinated primary effluent. With a combined daily chlorinated primary effluent output of approximately 3.7 x 10(8) liters, these two plants were discharging 1.0 x 10(10) VIU per day. Because the river has a mean annual flow of 8.0 x 10(10) liters per day, these two sources alone produced a virus loading of 1.0 VIU/8 liters of the river water. This river also receives at least 9.0 x 10(7) liters of raw sewage per day and undetermined but substantial amounts of storm waters and agricultural wastes. It is used for recreation and acts as a source of potable water for some 6.0 x 10(5) people. In view of the potential of water for disease transmission, discharge of such wastes into the water environment needs to be minimized.     

Elimination of human enteric viruses during conventional waste water treatment by activated sludge

The present study was undertaken to determine if viruses were selectively eliminated during waste water treatment. Human enteric viruses were detected at all steps of treatment in a conventional activated sludge waste water treatment plant. Liquid overlays and large volume sampling with multiple passages on BGM cells permitted the detection of poliovirus (serotypes 1, 2, and 3), coxsackievirus B (serotypes 1, 2, 3, 4, and 5), and echovirus (serotypes 3, 14, and 22), as well as reoviruses. The mean virus concentration was 95.1 most probable number of infectious units per litre (mpniu/L) in raw sewage, 23.3 in settled water, 1.4 in effluent after activated sludge treatment, and 40.3 mpniu/L in sludge samples. All samples of raw sewage and settled water, 79% of effluent water, and 94% of sludge samples contained viruses. The mean reduction was 75% after settling and 98% after activated sludge treatment. Poliovirus type 3 was rarely isolated after the activated sludge treatment, but was still detected in about one-third of the sludge samples. Reoviruses and coxsackieviruses were detected at similar rates from all samples and appear to be more resistant to the activated sludge treatment than poliovirus type 3. Poliovirus types 1 and 2 were present in almost every sample of raw sewage and settled water and still found in about half of the effluent and sludge samples, indicating a level of resistance similar to that of reoviruses and coxsackieviruses.     

Viral Pollution of Surface Waters Due to Chlorinated Primary Effluents

The role of chlorinated primary effluents in viral pollution of the Ottawa River (Ontario) was assessed by examining 282 field samples of wastewaters from two different sewage treatment plants over a 2-year period. The talc-Celite technique was used for sample concentration, and BS-C-1 cells were employed for virus detection. Viruses were detected in 80% (75/94) of raw sewage, 72% (68/94) of primary effluent, and 56% (53/94) of chlorinated effluent samples. Both raw sewage and primary effluent samples contained about 100 viral infective units (VIU) per 100 ml. Chlorination produced a 10- to 50-fold reduction in VIU and gave nearly 2.7 VIU/100 ml of chlorinated primary effluent. With a combined daily chlorinated primary effluent output of approximately 3.7 × 10⁸ liters, these two plants were discharging 1.0 × 10¹⁰ VIU per day. Because the river has a mean annual flow of 8.0 × 10¹⁰ liters per day, these two sources alone produced a virus loading of 1.0 VIU/8 liters of the river water. This river also receives at least 9.0 × 10⁷ liters of raw sewage per day and undetermined but substantial amounts of storm waters and agricultural wastes. It is used for recreation and acts as a source of potable water for some 6.0 × 10⁵ people. In view of the potential of water for disease transmission, discharge of such wastes into the water environment needs to be minimized.     

Fate of human enteric viruses, coliphages, and Clostridium perfringens during drinking-water treatment.

The elimination of human enteric viruses, coliphages, and Clostridium perfringens was studied during a conventional complete drinking-water treatment process. The respective concentrations (geometric mean) of these microorganisms in 100-L samples of river water were, respectively, as follows: viruses, 79 mpniu (most probable number of infectious units) per 100 L, coliphages, 6565 pfu (plaque-forming units) per 100 L. and clostridia, 11,349 cfu (colony-forming units) per 100 L. After predisinfection, flocculation with alum, and settling, human enteric viruses were not detected in any of the 100-L samples (less than 4 mpniu/100 L), but coliphages were detected in 7 of 14 samples and clostridia in 15 of 16 samples. In filtered water samples, human enteric viruses were detected in 2 of 31 samples, coliphages in 10 of 33, and clostridia in 17 of 33. Finished water was free of human enteric viruses (0/162 samples), but coliphages were detected in one sample (1.5 pfu/100 L) and clostridia in three, at 1.0, 4.1, and 7.0 cfu/100 L. It thus appears that coliphages and clostridia, which are present in larger numbers than viruses in river water and which may have similar resistance to drinking-water treatments, may be useful for estimating the level of treatment attained when large volumes of water (1000 L or greater) are sampled.     

Long-term survival of human rotavirus in raw and treated river water

This study was aimed at assessing the role of water as a vehicle for rotavirus spread by determining how well these viruses survive in the water environment. A cell culture adapted strain of human rotavirus subgroup 2, grown in MA-104 cells, was used as a model. Virus survival was tested in the following types of water samples, derived from the Ottawa River, at two different times of the year: (i) raw water (RW), (ii) muncipally treated tap water (TW), and (iii) raw water that had been filtered (FW) through a membrane (0.22 micron). The water samples, with approximately 5.0 X 10(4) plaque-forming units (PFU) of the virus, were held at either 4 or 20 degrees C and tested for infectious virus over a period of 64 days. The TW samples had a total and free chlorine content of 0.05 and less than 0.05 mg/L, respectively. The chlorine in these samples was not neutralized before virus contamination. Irrespective of the holding temperature, the virus titre in FW remained essentially unaltered throughout the test period. In TW held at 4 degrees C, there was no significant drop in the virus titre even after 64 days, whereas at 20 degrees C the titre in TW was reduced by about 2 log10 over the same period. Even though the loss of virus infectivity was most rapid in RW held at 20 degrees C, it took about 10 days for a 99.0% reduction in the plaque titre of the virus. These findings, therefore, indicate that rotaviruses can survive for several days in raw and treated river water thus making recreational and potable waters potential vehicles for the transmission of rotavirus infections.     

Detection of enteric viruses in treated drinking water

The occurrence of viruses in conventionally treated drinking water derived from a heavily polluted source was evaluated by collecting and analyzing 38 large-volume (65- to 756-liter) samples of water from a 9 m3/s (205 X 10(6) gallons [776 X 10(6) liters] per day) water treatment plant. Samples of raw, clarified, filtered, and chlorinated finished water were concentrated by using the filter adsorption-elution technique. Of 23 samples of finished water, 19 (83%) contained viruses. None of the nine finished water samples collected during the dry season contained detectable total coliform bacteria. Seven of nine finished water samples collected during the dry season met turbidity, total coliform bacteria, and total residual chlorine standards. Of these, four contained virus. During the dry season the percent removals were 25 to 93% for enteric viruses, 89 to 100% for bacteria, and 81% for turbidity. During the rainy season the percent removals were 0 to 43% for enteric viruses, 80 to 96% for bacteria, and 63% for turbidity. None of the 14 finished water samples collected during the rainy season met turbidity standards, and all contained rotaviruses or enteroviruses.     

Detection of enteric viruses in treated drinking water.

The occurrence of viruses in conventionally treated drinking water derived from a heavily polluted source was evaluated by collecting and analyzing 38 large-volume (65- to 756-liter) samples of water from a 9 m3/s (205 X 10(6) gallons [776 X 10(6) liters] per day) water treatment plant. Samples of raw, clarified, filtered, and chlorinated finished water were concentrated by using the filter adsorption-elution technique. Of 23 samples of finished water, 19 (83%) contained viruses. None of the nine finished water samples collected during the dry season contained detectable total coliform bacteria. Seven of nine finished water samples collected during the dry season met turbidity, total coliform bacteria, and total residual chlorine standards. Of these, four contained virus. During the dry season the percent removals were 25 to 93% for enteric viruses, 89 to 100% for bacteria, and 81% for turbidity. During the rainy season the percent removals were 0 to 43% for enteric viruses, 80 to 96% for bacteria, and 63% for turbidity. None of the 14 finished water samples collected during the rainy season met turbidity standards, and all contained rotaviruses or enteroviruses.     

Removal of indicator bacteria, human enteric viruses, Giardia cysts, and Cryptosporidium oocysts at a large wastewater primary treatment facility

Pathogens and fecal indicator bacteria occurrence and removal were studied for a period of 6 months at the Montreal Urban Community wastewater treatment facility. With a capacity of about 7.6 million cubic metres per day (two billion U.S. gallons per day), it is the largest primary physico-chemical treatment plant in America. The plant discharges a nondisinfected effluent containing about 20 mg/L of suspended matter and 0.5 mg/L of total phosphorus on the basis of average annual concentrations. BDO5 (annual mean) is 75 mg/L before treatment and 32 mg/L after treatment. Samples were collected for a period of 6 months, and they demonstrated that the plant was not efficient at removing indicator bacteria and the pathogens tested. Fecal coliforms were the most numerous of the indicator bacteria and their removal averaged 25%. Fecal streptococci removal was 29%, while Escherichia coli removal was 12%. In untreated sewage, fecal coliforms, E. coli, and human enteric viruses were more numerous in summer and early autumn. Fecal streptococci counts remained relatively similar throughout the period. Clostridium perfringens removal averaged 51%. Giardia cysts levels were not markedly different throughout the study period, and 76% of the cysts were removed by treatment. Cryptosporidium oocyst counts were erratic, probably due to the methods, and removal was 27%. Human enteric viruses were detected in all samples of raw and treated wastewater with no removal observed (0%). Overall, the plant did not perform well for the removal of fecal indicator bacteria, human enteric viruses, or parasite cysts. Supplementary treatment and disinfection were recommended to protect public health. Various alternatives are being evaluated.     

高碑店污水系统病毒污染情况的检测

本文报道用石英粉吸附——洗脱法浓集污水病毒,其回收率为51～90％。对1985年3～7月采集的不同水样进行病毒分离和蚀斑滴定,结果表明,高碑店污水系统中的病毒大部分为肠道病毒。春季以脊髓灰质炎病毒为主,夏季以柯赛奇B3及非肠道病毒为主。不同类型污水中的病毒浓度依次为:原污水58PFU/1;二级污水<19,约为9PFU/1;医用污水<16,约为1PFU/1,地下水<1.7PFU/1,未检出病毒。夏季水样内病毒浓度较春季低。     

Virus concentration from secondary wastewater: Comparative study between epoxy fiberglass and glass powder adsorbents

Summary We compare the efficiency of adsorption-elution methods using microfiberglass or glass powder adsorbents for concentrating indigenous viruses from secondary wastewater effluents sampled in two different cities in France.Analysis of 18 samples using both methods allowed recovery of viruses in all cases, with average concentrations of 2.7 to 29 MPNCU/1.Virological quantification gave better results with the glass powder method in 13 out of 18 cases. Using a new MPN technique which gives good 95% confidence limits, the results obtained with glass powder were statistically better than those obtained with microfiberglass only in 5 out of 9 experiments. From these data, it is very difficult to choose one or another method when only the average number of recovered viruses are considered. Cost and technical limitations (such as processing time, fragility of equipment and clogging) which are discussed here, must also be considered when making this choice.     

Detection of animal and human enteric viruses in water from the Assomption River and its tributaries

Animal enteroviruses, reoviruses, and human enteric viruses were detected in water samples (20 L) from a major river system, the Assomption River in the province of Quebec. Animal enteroviruses, probably of porcine origin (this region is a major producer of pork), were isolated on porcine cell cultures and were found in 29 to 60% of water samples from the different sites on the river and in 19 to 48% of the water samples from the tributaries. The average concentration of these animal enteroviruses in water from the Assomption River was 2 to 7 mpniu/L (most probable number of infectious units per litre), and that from the tributaries varied from 3 to 24 mpniu/L. Reoviruses were detected in infected cell cultures by an enzyme-linked immunosorbent assay. Their origin is probably avian (broiler chicken farms) or human (untreated domestic waste waters) and they were detected in 19 to 52% of the water samples from the Assomption River at an average concentration of 3 to 12 mpniu/L. In water samples from the tributaries, 5 to 71% of the samples were positive at an average concentration of 5 to 24 mpniu/L. Human enteric viruses were detected in MA-104 cells by an immunoperoxidase assay using human immune serum globulin. They were detected in 13 to 72% of water samples from the Assomption River and 14 to 71% of the water samples from the tributaries. The average concentration of these human enteric viruses in Assomption River water varied from 1 to 12 and from 2 to 145 mpniu/L in water samples from the tributaries.(ABSTRACT TRUNCATED AT 250 WORDS)     

Trypsin-treated Ma-104: a sensitive cell line for isolating enteric viruses from environmental samples

During a 1-year survey of enteroviruses in wastewater samples from the Lorraine area, three widely used continuous monkey kidney cell lines were tested: BGM, Vero, and trypsin-treated Ma-104. Decontaminated samples from secondary wastewater treatment plants (influent or effluent) were directly inoculated onto cells, and viruses were revealed after two passages with a liquid medium technique. Out of the total percentage of positive isolates with the three systems (32.7) 24.7% were found with Ma-104, 14.1% with BGM, and only 1.7% with Vero cells. Poliovirus was recovered more frequently with Ma-104 (12.3%) than with BGM (1.7%). Reovirus (3.5%) and echovirus (1.7%) were only found with Ma-104 cells; however, BGM cells allowed the isolation of a few group B coxsackieviruses (5.9%). It must be pointed out that 7.0% of samples with an unconfirmed cytopathic effect were found with BGM against 3.4% found with Ma-104, but they did not have significant differences. Because of its large spectrum of sensitivity, easy maintenance, and resistance to toxic effects, trypsin-treated Ma-104 may be recommended in conjunction with other cell lines for the detection of viruses from environmental samples, especially with the use of a liquid method.     

Trypsin-treated Ma-104: a sensitive cell line for isolating enteric viruses from environmental samples.

During a 1-year survey of enteroviruses in wastewater samples from the Lorraine area, three widely used continuous monkey kidney cell lines were tested: BGM, Vero, and trypsin-treated Ma-104. Decontaminated samples from secondary wastewater treatment plants (influent or effluent) were directly inoculated onto cells, and viruses were revealed after two passages with a liquid medium technique. Out of the total percentage of positive isolates with the three systems (32.7) 24.7% were found with Ma-104, 14.1% with BGM, and only 1.7% with Vero cells. Poliovirus was recovered more frequently with Ma-104 (12.3%) than with BGM (1.7%). Reovirus (3.5%) and echovirus (1.7%) were only found with Ma-104 cells; however, BGM cells allowed the isolation of a few group B coxsackieviruses (5.9%). It must be pointed out that 7.0% of samples with an unconfirmed cytopathic effect were found with BGM against 3.4% found with Ma-104, but they did not have significant differences. Because of its large spectrum of sensitivity, easy maintenance, and resistance to toxic effects, trypsin-treated Ma-104 may be recommended in conjunction with other cell lines for the detection of viruses from environmental samples, especially with the use of a liquid method.