RAPID METHOD FOR TESTING SUSCEPTIBILITY OF MYCOBACTERIUM-TUBERCULOSISBY USING DNA PROBES

The increasing number of multidrug-resistant Mycobacterium tuberculosi s strains has stimulated the interest of investigators in finding a ra pid method for susceptibility testing. We used commercially available rRNA DNA-bioluminescence-labelled probes (Accu-Probe, Gen Probe, Inc. San Diego, Calif.) for this purpose. The study was performed in three chronological steps. (i) We studied the correlation between the photom etric light units (PLUs) given by the hybridization method, the number s of CFU per milliliter, and turbidity as nephelometric units for six different inocula of an M. tuberculosis strain over 14 days. A good co rrelation (c > 0.9; P < 0.05) was found from the third day for all con centrations used. (ii) Over a period of 14 days we studied the evoluti on of the PLUs for 20 strains growing in medium with 0.2 mu l of isoni azid (H) per mi and 18 strains in medium with 1 mu l of rifampin (R) p er mi to standardize the method. Susceptible and resistant strains wer e used according to the reference proportions method in Middlebrook 7H 10, and the MICs were determined in solid and liquid media. The final inoculum of a 10(-2) dilution from a McFarland no. 1 standard and read ing at 3 and 5 days provided the best results. A quotient was establis hed to find a cutoff point between resistant and susceptible strains. (iii) We used the standardized parameters in 117 tests with H and R. O n day 3, the sensitivity, specificity, positive predictive value, and negative predictive value for detecting resistant strains were 86.8, 1 00, 100, and 90.1%, respectively, and on day 5 they were 96.2, 100, 10 0, and 94%, respectively. We concluded that the method is readily avai lable, is easy to perform, and could be useful for screening resistant M. tuberculosis strains.