Evaluation of trisodium phosphate as a transport medium and its utility in a single-step decontamination technique for the culture of M. tuberculosis.

ABSTRACT

Background Sputum specimens were received from various out-patient departments of a tertiary care referral hospital to acid-fast staining and mycobacterial culture. Material & Methods A simple, one-step decontamination and concentration method was adopted before subjecting the samples to acid-fast staining and mycobacterial culture. Trisodium phosphate, a cheap, “soft” decontaminant-cum-liquefying agent was used to treat the sputum specimens before Ziehl-Neelsen’s (ZN) acid-fast staining and Lowenstein-Jensen’s (LJ) medium culture. The sputum samples were collected directly into trisodium phosphate containing screw capped McCartney bottles (Day-0). The bottles were vortexed and left overnight at room temperature. On the subsequent morning (Day-1), the supernatants were discarded and smears made from deposits were stained by ZN stain. Results A total of 30 consecutive samples, which showed smear positivity by ZN technique, were selected for the present study. Deposits from these smear positive cases were cultured onto duplicate slants of LJ medium and incubated at 370C (Day-1). Duplicate slants of LJ media were inoculated from each of these preserved deposits on 2nd, 3rd , 4th, 6th and 8th days. Culture bottles were incubated at 370C for 8 weeks and positive growths were recorded. Culture retrieval was possible from 29 (96.7%) samples from deposits of Day –1 to Day-3. The culture positivity , however, had dropped to 26 (86.7%),18 (60%) and 6 (20%) from deposits of Day-4, Day-6 and Day-8, respectively. All the isolates were identified as M. tuberculosis and there was minimal contamination (0.83%). The culture retrieval dropped significantly only after Day-3. Conclusion This method is, therefore, suitable for transportation, preservation and decontamination of sputum samples before staining and culture, up to 3 days after collection. This will be helpful especially for collection of sputum samples from distant places and their transportation to nearest mycobacteriology laboratory as also for sputum samples arriving late in a working day’s schedule.