Methodology for recovery of indigenous viruses from primary sludge.

Freon extraction method for recovery of viruses from primary sludge has been studied with sonication and the beef extract eluation methods. Several variables within the freon extraction method were worked out and it was observed that equal volume of 10% buffered beef extract (pH 7.0), as that of the sludge sample, was required for optimum virus elution. Eluate decontamination with antibiotics, dilution and plaquing over BGM cell cultures using the overlay medium without phenol red, fortified with 1.7% milk powder resulted in best recoveries without any cytotoxicity to the cell cultures. A sample volume of 20 ml primary sludge was sufficient for quantification of viruses.

Detection of viruses in drinking water by concentration on magnetic iron oxide.

Discharge of raw domestic wastes containing human enteric viruses into water courses, consumption of untreated water from canals, streams, and shallow wells in villages, and cross-contamination of water in the distribution system because of intermittent water supply in urban areas continue to cause widespread outbreaks of infectious hepatitis in India. To detect a low number of viruses in 50- to 100-liter samples of water, a method was developed with magnetic iron oxide as the virus adsorbent. Poliovirus-seeded dechlorinated tap water, adjusted to pH 3.0 and 0.0005 M AlCl3, was filtered through a 10-g bed of iron oxide sandwiched between two AP20 prefilter pads held in a 142-mm-diameter, stainless-steel holder. Virus was eluted from iron oxide by recirculating three times a 100-ml volume of 3% beef extract, pH 9.0. The eluate was reconcentrated to 5 ml by adjusting to pH 3, adding 1 g of iron oxide, stirring for 30 min, and eluting the readsorbed virus with 5 ml of beef extract, pH 9.0. Virus recovery varied from 60 to 80%. Using the above method, we took a survey of drinking water at three locations in Nagpur during 1976 and found the presence of virus in 7 of 50 samples. The quantity of virus recovered ranged from 1 to 7 plaque-forming units per 30 to 60 liters. Virus was detected in some samples even with residual chlorine. No coliforms were detected in the virus-positive samples.