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
Kj. Schwab et al., 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, Applied and environmental microbiology, 61(2), 1995, pp. 531-537
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.