Vishnu Prasad Shenoy

Diabetes mellitus and HIV as co-morbidities in tuberculosis patients of rural south India.

OBJECTIVES Incidence of tuberculosis (TB) is greatest among patients with impaired immunity. India is experiencing a double epidemic of HIV and diabetes mellitus (DM), both of which are strongly associated with immuno-suppression. This study aimed to discover the prevalence of HIV and DM in both the pulmonary and extra-pulmonary TB patients of rural south India, retrospectively. METHODS Medical records of 192 microbiologically diagnosed pulmonary TB and 37 extra-pulmonary TB patients were thoroughly studied and data were extracted. The frequency distribution of HIV and DM was evaluated along with other demographic details such as age, sex and occupation in both groups. RESULTS The mean age of the pulmonary TB patients was 41.11±15.7 years, with significantly higher (p<0.0001) preponderance of DM (31.8%) over HIV (8.9%). 72.13% of the diabetic patients belonged to the age group of 41-60 years. Extra-pulmonary TB patients had a mean age of 34.62±12.9, years with a significantly higher (p<0.006) HIV prevalence of 32.43% over DM (5.4%). 75% of the HIV patients belonged to the age group of 41-60 years. Occupationally, the majority of the pulmonary TB patients were agricultural labourers (25.2%) while the majority of the extra-pulmonary TB patients were housewives or self employed (18.92%). CONCLUSION Though more importance is being given to HIV-TB coinfection, we cannot overlook DM, which showed a significantly higher prevalence in pulmonary TB patients compared to HIV. The rising prevalence of DM in high TB burden countries may adversely affect TB control.

Role of risk factors and socio-economic status in pulmonary tuberculosis: a search for the root cause in patients in a tertiary care hospital, South India.

Objective  To determine the frequency of underlying risk factors and the socio‐economic impact based on occupation in the development of tuberculosis.

Isolated tuberculous epididymo-orchitis: an unusual presentation of tuberculosis

Isolated epididymo-orchitis is an unusual presentation of tuberculosis. A case of bilateral epididymitis and right-sided orchitis with scrotal involvement in a 38-year-old male patient is presented. Strong clinical suspicion of tuberculous etiology was confirmed by appropriate investigations of epididymal biopsy. The patient improved clinically with antitubercular therapy.

Honey as an antimicrobial agent against Pseudomonas aeruginosa isolated from infected wounds

Background: As natural products garner attention in the medical field due to emergence of antibiotic resistant strains of bacteria, honey is valued for its antibacterial activity. Objective: Fifty strains of Pseudomonas aeruginosa isolated from infected wounds were evaluated for their antibacterial action using honey in comparison with different antibiotics and Dettol. Methodology and Results: All the strains were found to be sensitive to honey at a minimum inhibitory concentration of 20% in comparison with Dettol at 10% using agar dilution method. In the second step, the time kill assay was performed on five isolates of P. aeruginosa to demonstrate the bactericidal activity of honey at different dilutions of honey ranging from 20% to 100% at regular time intervals. All the isolates of P. aeruginosa tested were killed in 12-24 h depending on the dilutions of the honey tested. Thus, honey could prevent the growth of P. aeruginosa even if it was diluted by deionized water by fivefolds in vitro. Honey had almost uniform bactericidal activity against P. aeruginosa irrespective of their susceptibility to different classes of antibiotics. Conclusion: Honey which is a natural, non-toxic, and an inexpensive product has activity against the P. aeruginosa isolated from infected wounds may make it an alternative topical choice in the treatment of wound infections.

Rapid immunochromatographic test for the identification and discrimination of Mycobacterium tuberculosis complex isolates from non-tuberculous mycobacteria

BACKGROUND A new rapid Immunochromatographic test (ICT) kit (SDBioline TB Ag MPT64RAPID(®)) developed by Standard Diagnostics, South Korea was evaluated for rapid differentiation of M. tuberculosis from non tuberculous mycobacteria (NTM). It detects MPT 64 antigen in M. tuberculosis isolates using mouse monoclonal MPT 64 antibody. The kit was assessed for routine identification of the Acid Fast Bacilli(AFB) isolated in our laboratory. MATERIALS AND METHODS Two hundred eight culture isolates of Mycobacteria were tested using ICT test kit for detection of MPT 64 antigen from liquid and solid culture. H37Rv strain was employed as the positive reference control. Any negative result was referred for confirmation by Gen Probe Accu Probe assay for MTB Complex (Gen-Probe, San Diego, Calif.). Speciation of NTM was performed using genotypic Mycobacterium CM assay (Hains life sciences, Germany). RESULTS Of the 208 culture positive isolates tested, 182 (87.5%) were found positive for Mycobacterium tuberculosis Complex and remaining 26 (12.5%) were considered as NTM. These results were further confirmed by Gen Probe Accu probe assay that served as the reference method for detection of MTBC. H37Rv reference strain was taken as a control for ICT test and molecular tests. The reference strain showed the presence of MPT64 antigen band in the ICT test. Similar bands were formed in all MTBC (182) isolates tested, proving 100 per cent sensitivity and no bands were detected in 48 (100%) NTM isolates tested, proving 100 per cent specificity of the ICT kit. CONCLUSION Tuberculosis is a global pandemic. Rapid identification of Mycobacteria as MTB complex or non-tuberculous Mycobacteria from culture is important for treatment of infected cases and drug susceptibility testing of the culture isolate. MPT 64 TB antgen detection using SD Bioline Immunochromatographic test is a simple and cost effective method for differentiation of Mycobacterial cultures as MTB complex from non- tuberculous Mycobacteria.

Comparison among three cold staining methods in the primary diagnosis of tuberculosis: a pilot study

OBJECTIVE In developing countries, sputum smear microscopy is the main tool for pulmonary tuberculosis case finding. The objective of the present study was to evaluate the diagnostic efficacy of Gabbetts staining (GS) and modified cold staining (MCS), both of which are two-step methods, in comparison with that of fluorescent staining (FS), which is a three-step method, for the detection of AFB in sputum smears. METHODS Our sample comprised 260 sputum samples collected from individuals suspected of having pulmonary tuberculosis at Kasturba Hospital, in Manipal, India. Smears were prepared in triplicate: one each for FS, MCS, and GS. The smears were randomly numbered so that the examiner was blinded to the sample identities. RESULTS Of the 260 samples, 16 (6.15%), 15 (5.77%), and 13 (5.00%) showed positive AFB results with FS, MCS, and GS, respectively. The sensitivity of GS and MCS, in comparison with that of FS, was 81.25% and 93.75%, respectively. The concordance of GS and MCS with FS was good (0.988 and 0.996, respectively), and no statistically significant differences were found. CONCLUSIONS Although MCS and GS were found to be less sensitive than was FS, which is evaluated under fluorescence microscopy, the first two are promising methods for the diagnosis of tuberculosis.

Role of real-time PCR for detection of tuberculosis and drug resistance directly from clinical samples.

BACKGROUND Only a few studies done earlier in India reveal the utility of real-time PCR in detecting drug resistance in cases of pulmonary tuberculosis. OBJECTIVES The study was carried out to standardise real-time PCR (Quantitative real-time PCR, qPCR) targeting 16s RNA for the rapid detection of tuberculosis and its drug resistance from suspected TB patients. MATERIALS AND METHODS Sputum samples from 100 clinically suspected tuberculosis patients, after processing were subjected to microscopy, MGIT culture and qPCR. qPCR targeted 16sRNA for detecting Mycobacterium tuberculosis complex, KatG and rpoB genes for detection of resistance to isoniazid and rifampicin respectively. 1% proportionate method and Line probe assay (Hain Lifesciences, Nehren, Germany) were used to confirm the MDR isolates. RESULTS The study showed positivity of microscopy, culture and qPCR for M. tuberculosis as 37%, 44% and 46% respectively. Sensitivity of 100% and specificity of 96.5% in the detection of M. tuberculosis was observed for qPCR in comparison to culture. MDRTB was detected in 14 cases whereas monoresistance to rifampicin and isoniazid was detected in 1 and 3 samples respectively. CONCLUSION Real-time PCR targeting 16sRNA, KatG and rpoB is a sensitive, specific, rapid and reliable technique to detect pulmonary tuberculosis and its MDR status directly from the sputum samples.

Rational design and synthesis of novel diphenyl ether derivatives as antitubercular agents

A series of triclosan mimic diphenyl ether derivatives have been synthesized and evaluated for their in vitro antitubercular activity against Mycobacterium tuberculosis H37Rv. The binding mode of the compounds at the active site of enoyl-acyl carrier protein reductase of M. tuberculosis has been explored. Among them, compound 10b was found to possess antitubercular activity (minimum inhibitory concentration =12.5 µg/mL) comparable to triclosan. All the synthesized compounds exhibited low levels of cytotoxicity against Vero and HepG2 cell lines, and three compounds 10a, 10b, and 10c had a selectivity index more than 10. Compound 10b was also evaluated for log P, pKa, human liver microsomal stability, and % protein binding, in order to probe its druglikeness. Based on the antitubercular activity and druglikeness profile, it may be concluded that compound 10b could be a lead for future development of antitubercular drugs.

Diagnostic efficacy of Ziehl-Neelsen method against fluorescent microscopy in detection of acid fast bacilli

Objective To investigate the application of Ziehl-Neelsen (Z-N) and fluorescent microscopy in detection of acid fast bacilli (AFB). Methods Duplicate smears were prepared from 260 sputum samples and stained with Z-N and fluorescent staining (FS) methods. The efficiency of both methods in primary diagnosis of tuberculosis were evaluated. Results The smears were positive for AFB in 15 (5.77%) samples by Z-N staining method and in 16 (6.15%) samples by FS method. The sensitivity and specificity of Z-N staining method against FS method were 93.75% and 100% respectively. Conclusions Though lesser cost-effective than Z-N, FS method is a more sensitive and better case finding tool in detection of AFB.

Role of multiplex polymerase chain reaction in diagnosing tubercular meningitis

Tuberculous meningitis (TBM) is one of the most serious manifestations of extrapulmonary tuberculosis. Timely and accurate diagnosis provides a favorable prognosis in patients with TBM. The study evaluated the use of multiplex polymerase chain reaction (PCR) in the diagnosis of TBM. A study was conducted on 74 patients clinically suspected with TBM. The cerebrospinal fluid (CSF) specimens were processed for smear microscopy, middle brook 7H9 culture, and multiplex PCR using primers directed against IS6110 gene and 38 kD protein for detection of Mycobacterium tuberculosis. The results were analyzed to assess the role of multiplex PCR in the diagnosis of TBM. A total of 26 (35.1%) patients were diagnosed with TBM. Microscopy was negative in all while culture was positive in two cases only. Comparing with clinical diagnosis and CSF adenosine deaminase levels of ≥10 U/L, multiplex PCR showed sensitivity, specificity, positive predictive value, and negative predictive value of 71.4%, 89.6%, 83.3%, and 81.2%, respectively, in the diagnosis of TBM.