Use of vancomycin in the culture of Mycobacterium tuberculosis from gastric lavage

Background & objectives: Earlier studies from the Tuberculosis Research Centre, Chennai, on culture of Mymbocterium (tuberculosis from gastric lavage (GL) specimens in selective Kirchner's medium (SK) resulted in a loss of 60 per cent culture results due to contamination with aerobic spore bearers (ASB). Addition of vancomycin to SK (SKV) effectively reduced the contamination rate to 20 per cent. The objective of the present study was to further reduce the contamination by collecting the specimens in bottles containing vancomycin, thus providing continuous exposure of the sample to the drug, which is bactericidal to ASB. Methods: One thousand GL specimens coIIected from children in vancomycin containing bottles were decontaminated and cultured in SK medium, with and without vancomycin, subcultured on Lowenstein Jensen (W) medium and the culture results compared. Results: The contamination of cultures in SK and SKV was 15 and 4 per cent respectively when the specimens were collected in bottles containing vancomycin compared to 60 and 20 per cent contamination reported in the earlier studies. interpretation & conclusion: The reduced contamination in SK and SKV is most likely due to the collection of sample in vancomycin containing bottles. Although a concurrent comparison of samples processed in vancomycin free conditions would have been ideal, it could not be done due to practical difficulties. The study thus confirms the value of vancomycin as a major deterrent for contamination due to aerobic spores and better results can be obtained if vancomycin is used in sample collection bottles, transport media and liquid culture media used in mycobacteriology laboratories particularly in humid and tropical environment

Evaluation of various methods of susceptibility to ofloxacin in strains of Mycobacterium tuberculosis

A comparison of three methods of susceptibility testing was undertaken on 30 susceptible and 25 resistant strains of Mycobacterium tuberculosis to determine an acceptable in vitro definition of resistance of ofloxacin. The strains were tested by the proportion method on Lowenstein Jensen (L-J) and 7H11 media and also by the BACTEC radiometric method. Using a criterion of 1 per cent or more growth at a concentration of 2 mg/1, there was a 100 per cent agreement with the conventional MIC method by the proportion tests on L-J as well as on 7H11 media. The BACTEC radiometric method, at the same concentration, yielded 98 per cent agreement. Thus, any of these methods could be used depending upon the infrastructure available

Nocardia bacteraemia in an HIV-positive patient - a case report

Nocardiosis has been recognized in recent times as an unusual opportunistic infection associated with HIV. Bacteraemia due to this pathogen is even rarer and only few cases have been reported in the literature. We report here a case of pulmonary nocardiosis with bactcracmia, which was initially diagnosed as pulmonary tuberculosis. A high index of suspicion is required to diagnose this infection as the clinical presentation and radiographic features mimic pulmonary tuberculosis

Early results from indirect drug susceptibility test for tubercle bacilli

The indirect susceptibility test results on L-J medium for tubercle bacilli against streptomycin, isoniazid and rifampicin were read at the end of 2 wk and compared with the results at 4 wk. It was found that drug resistance could be correctly predicted in over 70 per cent of cultures including multi-drug resistant tuberculosis (MDR TB) strains at the end of 2 wk. The susceptibility to para-nitrobenzoic acid (PNB) read at 2 wk was able to distinguish non-tuberculous mycobacteria from Mycobacterium tuberculosis cultures. The early detection of resistance by this procedure requires only minimum inputs, and can benefit the majority of patients harbouring drug resistant tubercle bacilli

Secondary bacterial flora in patients with pulmonary tuberculosis - a preliminary report

Sputum samples from 100 smear positive or skiagram positive pulmonary tuberculosis patients were cultured for superinfecting or co-injecting bacteria. These patients were equally divided into five groups. This included Croup-I who are not treated; Group-II who are treated up to three months; Group-III who are treated for more than three but less than six months; Group-IV treated more than six months and lastly Group-V who have completed the prescribed treatment schedule of varying durations. Neisseria catarrhalis and Strep. viridans predominated in all patients irrespective of group, other organisms isolated, were Micrococci, E.Coli, Serratia, Proteus and Pseudomonas. There was no significant difference in the pattern of organisms isolated from different group of patients. The antibiogram showed the usual susceptibility pattern

Vancomycin for controlling contamination of selective Kirchner’s liquid medium in the culture of gastric lavage for tubercle bacilli

Gastric lavage (GL) was collected for culture of tubercle bacilli from children too young to expectorate sputum. The selective Kirchner’s liquid medium (SKLM), routinely used as one of the media for culture of all extrapulmonary specimens, was found to often get contaminated when cultured with GL. We have shown that vancomycin at a concentration of 10 mg/l successfully reduced the contamination from about 60 to 20 per cent, and enhanced the Isolation rate of tubercle bacilli from 3 to 6 per cent. Decontamination of the liquid culture before subculture on solid medium also helped to reduce the contamination rate. Vancomycin was found to be an effective selective drug for use In the Kirchner’s liquid medium for culture of tubercle bacilli

Identification & differentiation of Mycobacterium avium & M. intracellulare by PCR- RFLP assay using the groES gene

Background & objectives: We report a new polymerase chain reaction (PCR) – restriction fragment length polymorphism (RFLP) assay using mycobacterial groES as a target to identify Mycobacterium avium and M. intracellulare in clinical samples. Methods: The assay was standardized using M. avium and M. intracellulare standard strains obtained from ATCC and was tested with 45 M. avium-M. intracellulare complex (MAC) clinical isolates (Of which 31 were from HIV+ individuals). The standard and clinical strains were typed with HPLC based mycolic acid fingerprinting. Results: Three polymorphisms (BamHI, BstNI and HgaI) were identified for inter-species differentiation among standard strains; of which, only HgaI was found to be useful in clinical isolates. Of the 45 isolates, 25 were M. avium and 20 were M. intracelluare. MAC isolates, which could not be differentiated by HPLC analysis, were also typed by this method. Interpretation & conclusions: The use of mycobacterial groES as a PCR-RFLP target for M. avium and M. intracellulare is a simple and rapid method that can complement HPLC in their differentiation

Seroprevalence of human immunodeficiency virus (HIV) infection among tuberculosis patients in Tamil Nadu

Background & objectives: The dual epidemic of HIV and tuberculosis is a cause for concern in those countries where these two infections are prevalent in epidemic proportions. We undertook a survey at two sites in North Arcot district of Tamil Nadu in 1992-1993, to know the seroprevalence of HIV infection among tuberculosis patients. The objective of this study was to re-examine the prevalence of HIV infection among tuberculosis patients in a repeat survey. Methods: The study was undertaken in four centres: District Tuberculosis Centre (DTC), Vellore, Tuberculosis Sanatorium, Pennathur (Vellore), District TB Centre (DTC), Kancheepuram and the Government Thiruvotteswarar Tuberculosis Hospital (GTTH), Chennai in the northern part of Tamil Nadu during 1997-1998. A total of 2361 newly diagnosed TB patients were registered in this study. HIV serology after pre-test counseling was done along with sputum examination for acid-fast bacillus by smear and culture for mycobacteria for all patients. Results: The overall HIV seroprevalence among TB patients was 4.7 per cent. The highest HIV seropositivity rate was found among patients aged 30-39 yr (10.6%). HIV seroprevalence showed a wide variation among the different centres ranging from 0.6. per cent in DTC, Kancheepuram to 9.4 per cent in Pennathur Sanatorium, Vellore. Sputum smear positivity was 88 per cent among the HIV-negative and 83 per cent among HIV-positive tuberculosis patients. Interpretation & conclusion: HIV infection is on the rise among TB patients in Tamil Nadu. Acid-fast smear microscopy is adequate for the diagnosis of pulmonary tuberculosis, and drug resistance among HIV positive patients is not a major problem at this point of time; hence antituberculosis regimens recommended by the Revised National Tuberculosis Control Program (RNTCP) can be used to treat HIV positive patients with tuberculosis

Simple Direct Drug Susceptibility Tests on Sputum Samples for Early Detection of Resistance in Tubercle Bacilli

Background: Direct sensitivity test either by sputum concentrate (DS) or swab method (DSM) set up along with the primary culture would avoid the delay of four or more weeks required for the indirect test. A comparison of these two methods against the standard indirect sensitivity method under routine laboratory conditions is necessary to prove their merit. Method: Smear positive sputum samples were aliquoted and sensitivity tests were set up by both the direct methods as also an indirect test set up from the primary culture of the same sample. Results: The agreement with the indirect test results for isoniazid (INH) ranged from 97-98% for the DS method and 93- 97% for the DSM method. The corresponding figures were 96-98% by the DS and 94-99% by the DSM method for rifampicin (R). The agreement was less satisfactory for ethambutol (Emb). Conclusion: This study showed that direct sensitivity tests such as DS and DSM methods can detect most of the cultures resistant to INH and R (MDR) from the time growth appears on the primary culture , even as early as the second week of setting up the tests