Polymerase chain reaction detection of nonviable bacterial pathogens.
Polymerase chain reaction (PCR) methodologies for detection of pathogens in environmental samples are currently available. However, positive amplification products for any set of primers only signal that the appropriate target nucleic acid sequences were present in the sample. The presence of the amplification products does not imply that the target organisms were viable. Here we show that PCR will detect nonviable cells, as long as intact target nucleic acid sequences are available. In an environmental water sample, nucleic acids degraded quickly and were not detectable by PCR after 3 weeks even when stored at 4 degrees C. However, these data show that there is a window of opportunity for PCR analyses to result in false positives with respect to viable cells. We further show that care must be taken in the way samples are stored for future PCR amplifications and that filter sterilization of media is not acceptable for long-term preservation of samples for PCR.
Josephson
K L
KL
Department of Soil and Water Science, University of Arizona, Tucson 85721.
Gerba
C P
CP
Pepper
I L
IL
D001419:Bacteria / Q000235:genetics* / Q000302:isolation & purification* / Q000472:pathogenicity; D015169:Colony Count, Microbial; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D004783:Environmental Microbiology; D004926:Escherichia coli / Q000235:genetics / Q000302:isolation & purification; D005069:Evaluation Studies as Topic; D005189:False Positive Reactions; D016133:Polymerase Chain Reaction / Q000379:methods*; D012485:Salmonella typhi / Q000235:genetics / Q000302:isolation & purification; D012764:Shigella sonnei / Q000235:genetics / Q000302:isolation & purification
D004269:DNA, Bacterial