Evaluation of thin-layered agar for Mycobacterium tuberculosis isolation and drug susceptibility testing

Abstract

Background: Tuberculosis (TB) is India s major public health problem. According to the WHO, India harbors the largest number of cases, and TB control remains a challenge in diagnosis, drug resistance, and treatment. We undertook this study to compare the isolation rates of Mycobacterium tuberculosis (MTB) in agar, egg-based media, incidence of multidrug-resistant (MDR), and extended drug resistance (XDR) in MTB. This study aimed to compare and evaluate thin-layered agar (TLA) for the cultivation and drug susceptibility pattern of MTB with the conventional egg-based Lowenstein–Jensen s (LJ) medium and to differentiate atypical Mycobacterium by incorporating para-nitrobenzoic acid (PNB) in the TLA medium. This cross-sectional study was conducted in Sri Ramachandra Medical College and Research Institute, Porur, Chennai. Methods: A total of 68 smear-positive samples were inoculated into TLA (Middle Brook 7H 11) and LJ media with and without antibiotics (rifampicin, isoniazid, and ofloxacin) simultaneously after decontamination by the modified Petroff s method. TLA with PNB was also used to differentiate the growth of nontuberculous mycobacterium (NTM). Incubation was done at 37°C, and reading was taken every 3rd day for 6 weeks in case of TLA and for 8 weeks in case of LJ medium. Results: Out of the 68 samples, 64 (94.1%) grew in LJ, and the growth observed at the end of the 1st, 2nd, 3rd, 4th, 5th, and 6–10th weeks was 0, 12 (18.8%), 10 (15.6%), 14 (21.9%), 15 (23.4%), and 13 (20.3%), respectively. Similarly, in TLA, 65 (95.5%) samples were grown, among which 22 (33.8%) grew in the 1st week and the rest (43 [66.2%]) in the 2nd week. MDR and XDR were observed in 4 (5.8%) and 3 (4.4%) samples, respectively. Seven of them were NTM. Conclusions: TLA is a better medium, with time to positivity ranging from 1 to 2 weeks with drug susceptibility and the pattern is also comparable with LJ medium. Incorporation of PNB in TLA helps in differentiating NTM.