Effects of different methods of decontamination for successful cultivation of Mycobacterium tuberculosis

There has been an extensive invasion of tuberculosis at the global level by multidrug resistant as well as extensively drug resistant organisms. Attempts to recover the pathogen in pure culture have frequently failed since the specimens are often highly contaminated and also due to use of insufficient or over-active decontamination procedures. Hence in the present study different methods of decontamination were tested to evaluate their independent efficacies for culture of Mycobacterium tuberculosis.

A total of 359 samples (241 sputum, 59 urine, 50 endometrium biopsy, 9 pus samples) from clinically suspected cases of tuberculosis were subjected to four different methods of decontamination followed by inoculation in Lowenstein-Jensen medium (LJM), and bilayered medium (BLM) and Kirchner's liquid medium (KLM) to determine the influence of differential decontamination processes. Sputum scanty and positive specimens were graded and each sample was subjected to decontamination by four different techniques.

Treatment of specimens with 4 per cent NaOH yielded minimum recovery of pure cultures, while use of 2 per cent NaOH produced higher number of contaminants compared to other methods of decontamination. Addition of N-acetyl L-cystein (NALC) coupled with 2 per cent NaOH to the samples for decontamination provided fairly reasonable recovery, but the highest number of M. tuberculosis cultures could be obtained when the specimens were treated with tri-sodium phosphate and benzalkonium (TSPB). Among the sputum positive cases recovery of growth of M. tuberculosis was higher with greater number of bacilli present in the specimens. Regarding the influence of culture media, BLM produced not only rapid growth, but reasonably higher rate of isolation of M. tuberculosis.

Although use of TSPB was found to be an efficient method of decontamination for successful isolation of M. tuberculosis from contaminated samples, both NALC+ 2 per cent NaOH and TSPB also showed significant recovery of M. tuberculosis cultures in BLM that can facilitate early diagnosis and initiation of treatment.