Influence of environmental factors on virus detection by RT-PCR and cell culture.
The effects of clay, humic acid, u.v. light and shellfish tissue residues on the detection of poliovirus type 2 from environmental samples by culture and RT-PCR were investigated. RT-PCR showed 10-100 times greater sensitivity for PV2 detection in the absence of sample contaminants than did culture by plaque assay in BGM cell monolayers. Bentonite clay (100-1000 mg l-1) and shellfish tissue residues reduced virus detection by plaque assay, but the effect of bentonite was mitigated by simple elution procedures. Bentonite clay, humic acid (5-150 mg l-1) and mussel tissue reduced virus detection by RT-PCR by between 1 and 8 logs, although this was mitigated in part by elution and Sephadex filtration of extracts. Sephadex filtration of samples reduced culturable PV2 by 32-50%. Exposure of PV2 in water to u.v. light reduced culturability of PV2 but not detection by RT-PCR. This study demonstrates that virus detection in environmental samples is strongly influenced by naturally occurring substances and disinfection approaches. The accuracy of results of viral analyses of this nature should be carefully scrutinized with respect to sample constituents.
Lewis
G D
GD
School of Biological Sciences, University of Auckland, and Centre for Disease Control, Environmental Science and Research, Porirua, New Zealand. gd.lewis@auckland.ac.nz
Molloy
S L
SL
Greening
G E
GE
Dawson
J
J
D000818:Animals; D001546:Bentonite; D049872:Bivalvia; D018929:Cell Culture Techniques; D004279:DNA, Viral / Q000032:analysis; D006801:Humans; D006812:Humic Substances; D017955:Poliovirus / Q000235:genetics / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D014466:Ultraviolet Rays
D004279:DNA, Viral; D006812:Humic Substances; D001546:Bentonite