ASSESSMENT OF MYCOBACTERIAL VIABILITY BY RNA AMPLIFICATION

We investigated whether the presence of intact RNA is a valuable indic ator of viability of mycobacteria with Mycobacterium smegmatis. M. sme gmatis was exposed to various concentrations of rifampin and ofloxacin suspended in broth for different periods of time. The NASBA nucleic a cid amplification system was used because of its rapid, sensitive, and specific detection of 168 rRNA. During drug exposure, the viability o f the mycobacteria, expressed by the number of CFU, was compared with the presence of 16S rRNA as determined by NASBA and,vith the presence of DNA coding for 16S rRNA as determined by PCR. Both NASBA and PCR we re shown to have a detection limit of approximately 5 x 10(2) CFU/ml. The intensity of the NASBA signal corresponded well with the number of CFU, and the lack of NASBA signal coincided with a loss of viability, which was reached after 3 days of exposure to bactericidal concentrat ions of both drugs. The presence of mycobacterial DNA, as determined b y the intensity of the PCR signal, and the viability of M. smegmatis w ere not related, but an increase in the number of cells and intensity of PCR signal correlated well. Bacterial viability may thus be assesse d by a rapid, sensitive, and specific, and semiquantitative technique by using NASBA. This system of viability testing provides the potentia l for rapid evaluation of drug susceptibility testing.