Detection of mRNA by reverse transcription-PCR as an indicator of viability in Escherichia coli cells.
The relationship between the detection of mRNA and cellular viability in Escherichia coli was investigated in cells killed by heat or ethanol. Reverse transcription-PCR (RT-PCR) methods were developed for detecting mRNA from rpoH, groEL, and tufA genes. mRNA from all three genes was detected immediately after the cells had been killed by heat or ethanol but gradually disappeared with time when dead cells were held at room temperature. In heat-killed cells, some mRNA targets became undetectable after 2 to 16 h, whereas after ethanol treatment, mRNA was still detected after 16 h. In contrast, 16S rRNA was detected by RT-PCR in all samples containing dead cells and did not disappear during a subsequent incubation of 16 h at room temperature. Of the different types of nucleic acid, mRNA is the most promising candidate for an indicator of viability in bacteria, but its persistence in dead cells depends on the inactivating treatment and subsequent holding conditions.
Sheridan
G E
GE
Institute of Food Research, Reading, United Kingdom.
Masters
C I
CI
Shallcross
J A
JA
MacKey
B M
BM
D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D018834:Chaperonin 60 / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004926:Escherichia coli / Q000166:cytology / Q000235:genetics* / Q000378:metabolism*; D000431:Ethanol / Q000494:pharmacology; D005798:Genes, Bacterial; D006360:Heat-Shock Proteins / Q000235:genetics; D006358:Hot Temperature; D016133:Polymerase Chain Reaction / Q000379:methods*; D012329:RNA, Bacterial / Q000235:genetics* / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism*; D012336:RNA, Ribosomal, 16S / Q000235:genetics / Q000378:metabolism; D012808:Sigma Factor / Q000235:genetics; D014157:Transcription Factors
D001426:Bacterial Proteins; D018834:Chaperonin 60; D017931:DNA Primers; D006360:Heat-Shock Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; D012336:RNA, Ribosomal, 16S; D012808:Sigma Factor; D014157:Transcription Factors; C069774:heat-shock sigma factor 32; D000431:Ethanol