Exposure to cetyl pyridinium chloride and loss of integrity of cell wall of mycobacteria.

Background: Cetyl pyridinium chloride (CPC) liquefied sputum was shown to reduce AFB smear positivity presumably damaging cell wall of M. tuberculosis. Settings: National Institute for Research in Tuberculosis, Chennai, (Tamil Nadu). Objective: To assess the cell wall damage of mycobacteria in CPC liquefied sputum, by Transmission Electron Microscopy (TEM) and mycobacteriophage adsorption studies. Methods: Pooled sputum sample from smear positive pulmonary TB patients was homogenized and liquefied with CPC. It was examined in TEM daily for four days, to assess cell wall damage of M. tuberculosis, and photomicrographs were taken. M. smegmatis mc2155, treated with CPC, was infected with mycobacteriophage (phAE129) to study phage adsorption on cell wall and plaque formation. CPC untreated sputum and M. smegmatis formed controls. Results: Photomicrographs showed that cell wall of M. tuberculosis was intact in controls and damaged in CPC preserved sputum for 96 hours. Plaque formation was seen and absent respectively in CPC untreated and treated M. smegmatis cells. Conclusion: Exposure to CPC damaged the cell wall of M. tuberculosis within 96 hours. Mycobacteriophage failed to form plaques after M. smegmatis mc2155 was treated with CPC implying inhibition of phage adsorption on damaged cell wall and thus providing a clue for poor staining and smear positivity in microscopy.

Rapid detection of rifampicin susceptibility of mycobacterium tuberculosis in sputum specimens by mycobacteriophage assay

To evaluate the performance of FASTPlaqueTB-RIF TM, a newly introduced bacteriophage assay for rapid detection of rifampicin susceptibility of Mycobacterium tuberculosis in sputum specimens. A comparative study of 40 sputum specimens from patients of pulmonary tuberculosis, using FASTPlaqueTB-RIF TM and Bactec 460 TB system carried out at the Armed Forces Institute of Pathology, Rawalpindi between September and November 2001. Of the 40 clinical isolates of Mycobacterium tuberculosis tested for rifampicin [RIF] susceptibility using the Bactec 460 TB system, 28 isolates were resistant to RIF and 12 isolates were susceptible. FASTPlaqueTB-RIF TM identified 24 specimens as resistant to RIF. Three specimens that revealed susceptible isolates on Bactec 460, were resistant by FASTPlaqueTB-RIF' while four specimens which revealed resistant isolates on Bactec 460, demonstrated susceptibility to RIF by FASTPlaqueTB-RIF TM. The sensitivity and specificity of FASTPlaqueTB-RIF TM were 86% and 73% respectively. The predictive values of positive and negative tests were 0.89 and 0.67 respectively. The overall accuracy of the technique was 82%. The phage assay took 48 hours to perform. Early detection of rifampicin resistance by the mycobacteriophage technique direct from sputum specimens is a potentially useful new test which would allow decision regarding appropriate therapy to be made early thus having a positive impact on patient care and on prevention of spread of MDR TB