CONCENTRATION AND PURIFICATION OF BEEF EXTRACT MOCK ELUATES FROM WATER SAMPLES FOR THE DETECTION OF ENTEROVIRUSES, HEPATITIS-A VIRUS, AND NORWALK VIRUS BY REVERSE TRANSCRIPTION-PCR

In this study we developed a concentration and purification procedure to facilitate reverse transcription (RT)-PCR detection of enteric viru ses in water sample concentrates obtained by conventional filter adsor ption-elution methods. One liter of beef extract-glycine eluate with o r without humic acid and seeded with poliovirus type 1, hepatitis A vi rus, and Norwalk virus was used as a model system, and the eluent was further processed for RT-PCR compatibility. The sample concentration a nd purification procedures which we used included polyethylene glycol precipitation, Pro-Cipitate precipitation, a second polyethylene glyco l precipitation, spin column chromatography, and ultrafiltration. The sample volumes were reduced from 1 liter to 20 to 50 mu l, and the sam ples were purified enough so that viruses could be detected by the RT- PCR The ability to detect low levels of enteric viruses by molecular t echniques was compared directly with the ability to detect enteric vir uses by cell culture infectivity procedures. As little as 3 PFU of pol iovirus type 1 in an initial 1 liter of mock eluate was detected by th e RT PCR.