To develop simple, rapid, low-cost and robust assays for screening drugs against dormant and actively growing mycobacteria. Actively growing aerobic and hypoxia-adapted dormant cultures of Mycobacterium tuberculosis, Mycobacterium bovis BCG and Mycobacterium smegmatis were tested for susceptibility to standard antimicrobial drugs by resazurin reduction assay. The visual and fluorimetric MICs were compared with those obtained by the standard cfu assay. Drug MICs for M. tuberculosis and M. bovis BCG were determined by the aerobic resazurin microplate assay (REMA) and correlated well with those obtained by the cfu assay. Metronidazole and nitrofurans showed comparable bactericidal activity in the hypoxic resazurin reduction assay (HyRRA). The HyRRA assay was noted to be superior to the cfu assay in that it distinguished between metabolically active dormant bacteria and non-viable organisms, unlike the cfu assay that could not differentiate between these two populations. The HyRRA assay performed with good concordance in both fluorimetric and visual formats to distinguish between bactericidal and bacteriostatic effects of a drug. The REMA and HyRRA assays will be useful for anti-tubercular anti-dormancy compound screening and drug susceptibility testing in a safe, reliable, easy and cost-effective manner particularly in low resource countries. The application of the assays in M. smegmatis or M. bovis BCG offers the distinct advantage of rapidly and safely screening anti-tubercular compounds in a high-throughput format.

Tubercle bacilli are thought to persist in a dormant state during latent tuberculosis (TB) infection. Although little is known about the host factors that induce and maintain Mycobacterium tuberculosis (M. tb) within latent lesions, O₂ depletion, nutrient limitation and acidification are some of the stresses implicated in bacterial dormancy development/growth arrest. Adaptation to hypoxia and exposure to NO/CO is implemented through the DevRS/DosT two-component system which induces the dormancy regulon. Here we show that vitamin C (ascorbic acid/AA) can serve as an additional signal to induce the DevR regulon. Physiological levels of AA scavenge O₂ and rapidly induce the DevR regulon at an estimated O₂ saturation of <30%. The kinetics and magnitude of the response suggests an initial involvement of DosT and a sustained DevS-mediated response during bacterial adaptation to increasing hypoxia. In addition to inducing DevR regulon mechanisms, vitamin C induces the expression of selected genes previously shown to be responsive to low pH and oxidative stress, triggers bacterial growth arrest and promotes dormancy phenotype development in M. tb grown in axenic culture and intracellularly in THP-1 cells. Vitamin C mimics multiple intracellular stresses and has wide-ranging regulatory effects on gene expression and physiology of M. tb which leads to growth arrest and a ‘dormant’ drug-tolerant phenotype, but in a manner independent of the DevRS/DosT sytem. The ‘AA-dormancy infection model’ offers a potential alternative to other models of non-replicating persistence of M. tb and may be useful for investigating host-‘dormant’ M. tb interactions. Our findings offer a new perspective on the role of nutritional factors in TB and suggest a possible role for vitamin C in TB.

(2009). MMTV mouse models and the diagnostic values of MMTV-like sequences in human breast cancer. Expert Review of Molecular Diagnostics: Vol. 9, No. 5, pp. 423-440.

The present study concerns the isolation and characterization of potential probiotic bacteria isolated from indigenously fermented cereal-based products commonly produced by tribal people of the Aravali hills region of India and the documentation of their unexplored probiotic attributes. The isolated strains were evaluated for probiotic attributes, such as bile salt and acid tolerance, lysozyme and phenol tolerance, antagonistic and antifungal activity, cell autoaggregation, cell-surface hydrophobicity, simulated gastric and pancreatic digestion, antioxidative potential, bile salt hydrolase activity, and H₂O₂ production. The safety of isolates was assessed by antibiotic sensitivity, hemolytic activity, DNase activity, and biogenic amine production assays, while technological properties, such as fermenting ability, amylolytic activity, and EPS production, were also evaluated. A total of 70 LAB isolates were screened initially, and 6 strains showed good potential as probiotic candidates in in vitro assessments. The efficient strains were identified using phenotyping and biochemical characterization, which results were further confirmed and recognized at the strain level using phylogenetic analysis and 16S rDNA sequencing. The current study has shown that Lactiplantibacillus plantarum KMUDR7 isolated from “Makka ki Raab” has excellent probiotic attributes and could be a potential probiotic for product preparation. However, other strains, Lactobacillus delbrueckii subsp. bulgaricus KMUDR1 and Lacticaseibacillus rhamnosus KMUDR9, showed good properties, while KMUDR14, -17, and -20 also have comparable probiotic attributes.