rRNA stability in heat-killed and UV-irradiated enterotoxigenic Staphylococcus aureus and Escherichia coli O157:H7.
Differentiation of viable cells from nonviable cells is of considerable importance in the development of methods to detect foodborne pathogens. To study the suitability of 16S rRNA as an indicator of cell viability in nucleic acid-based detection assays, we examined rRNA stability in two representative foodborne pathogens, Escherichia coli O157:H7 and enterotoxigenic Staphylococcus aureus, which were inactivated by extreme heat, moderate heat, and UV irradiation. Cell death under all conditions was confirmed by a failure to grow in brain heart infusion broth after incubation for 48 h at 37 degrees C. rRNA stability was monitored by a Northern blot analysis, and detection was evaluated by using reverse transcription (RT)-PCR performed with two primer sets (which produced 325- and 1, 400-bp amplicons). rRNA of neither pathogen was detected by Northern blot analysis and RT-PCR after cells were killed by autoclaving at 121 degrees C for 15 min. In contrast, intact rRNA of both pathogens were detected by Northern blotting and could be amplified by RT-PCR up to 48 h after cells were killed by heat treatment at 80 degrees C and UV irradiation at 254 nm. rRNA was a suitable target molecule for monitoring bacterial viability under extreme heat conditions, but the presence of rRNA was not correlated with viability following moderate heat inactivation or UV irradiation of cells.
McKillip
J L
JL
Department of Food Science and Technology, Southeast Dairy Foods Research Center, Mississippi State University, Mississippi State, Mississippi 39762-9805, USA.
Jaykus
L A
LA
Drake
M
M
D015152:Blotting, Northern; D017931:DNA Primers; D019453:Escherichia coli O157 / Q000235:genetics* / Q000472:pathogenicity / Q000528:radiation effects; D005514:Food Irradiation; D005516:Food Microbiology; D006358:Hot Temperature; D012329:RNA, Bacterial / Q000235:genetics / Q000528:radiation effects*; D012335:RNA, Ribosomal / Q000235:genetics / Q000528:radiation effects*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013211:Staphylococcus aureus / Q000235:genetics* / Q000472:pathogenicity / Q000528:radiation effects; D013997:Time Factors; D014466:Ultraviolet Rays
D017931:DNA Primers; D012329:RNA, Bacterial; D012335:RNA, Ribosomal