Anand Kumar Maurya

Role of genotype® mycobacterium common mycobacteria/additional species assay for rapid differentiation between Mycobacterium tuberculosis complex and different species of non-tuberculous mycobacteria

Background: Mycobacterium tuberculosis complex (MTBC) and non-tuberculous mycobacteria (NTM) may or may not have same clinical presentations, but the treatment regimens are always different. Laboratory differentiation between MTBC and NTM by routine methods are time consuming and cumbersome to perform. We have evaluated the role of GenoType® Mycobacterium common mycobacteria/additional species (CM/AS) assay for differentiation between MTBC and different species of NTM in clinical isolates from tuberculosis (TB) cases. Materials and Methods: A total of 1080 clinical specimens were collected from January 2010 to June 2012. Diagnosis was performed by Ziehl-Neelsen staining followed by culture in BacT/ALERT 3D system (bioMerieux, France). A total of 219 culture positive clinical isolates (BacT/ALERT® MP cultures) were selected for differentiation by p-nitrobenzoic acid (PNB) sensitivity test as and BIO-LINE SD Ag MPT64 TB test considering as the gold standard test. Final identification and differentiation between MTBC and different species of NTM were further confirmed by GenoType® Mycobacterium CM/AS assay (Hain Lifescience, Nehren, Germany). Results: Out of 219 BacT/ALERT® MP culture positive isolates tested by PNB as 153 MTBC (69.9%) and by GenoType® Mycobacterium CM/AS assay as 159 (72.6%) MTBC and remaining 60 (27.4%) were considered as NTM species. The GenoType® Mycobacterium CM/AS assay was proved 99.3% sensitive and 98.3% specific for rapid differentiation of MTBC and NTM. The most common NTM species were; Mycobacterium fortuitum 20 (33.3%) among rapid growing mycobacteria and Mycobacterium intracellulare 11 (18.3%) among slow growing mycobacteria. Conclusion: The GenoType® Mycobacterium assay makes rapid and accurate identification of NTM species as compared with different phenotypic and molecular diagnostic tool and helps in management of infections caused by different mycobacteria.

The advantage of using IS6110-PCR vs. BACTEC culture for rapid detection of Mycobacterium tuberculosis from pleural fluid in northern India.

Pleural tuberculosis is an extra-pulmonary disease which poses a diagnostic dilemma. The detection of mycobacterial DNA by IS6110 polymerase chain reaction (PCR) in clinical samples is a promising approach for the rapid diagnosis of pleural tuberculosis infections. The aim of the present study is to evaluate the advantage of using IS6110 PCR for rapid detection of Mycobacterium tuberculosis (M. tuberculosis) from pleural fluid. 102 clinically suspected cases of pleural tuberculosis cases were enrolled from inwards and outwards of the Department of Pulmonary Medicine at Chattrapati Shahuji Maharaj Medical University, Lucknow from April 2007 to April 2010. The pleural fluids were processed at the Mycobacteriology Laboratory of Department of Microbiology at Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow, Pleural fluid samples were processed and examined by Ziehl Neelsen (ZN) staining for acid fast bacilli and detection of M. tuberculosis by BACTEC culture. We applied IS6110 PCR to detect specific M. tuberculosis complex in pleural fluid samples. We found a significant difference in sensitivity of different tests, acid fast bacilli were detected in 17 (16.6%) samples by ZN Staining , 47 (46.1%) by BACTEC culture and using IS6110 PCR, 62 (60.7%) were positive for IS6110 PCR for M. tuberculosis. We found IS6110 PCR was much more sensitive than ZN staining and BACTEC culture. IS6110 PCR detection of M. tuberculosis may be very useful in cases that are highly suspect as pleural tuberculosis and those that are negative for AFB and culture. IS6110 PCR may gain an immense prospective to better clinicians ability to improve diagnosis of pleural tuberculosis.

Mixed Cutaneous Infection Caused by Mycobacterium szulgai and Mycobacterium intermedium in a Healthy Adult Female: A Rare Case Report.

Nontuberculous mycobacteria (NTMs) are ubiquitous and are being increasingly reported as human opportunistic infection. Cutaneous infection caused by mixed NTM is extremely rare. We encountered the case of a 46-year-old female, who presented with multiple discharging sinuses over the lower anterior abdominal wall (over a previous appendectomy scar) for the past 2 years. Microscopy and culture of the pus discharge were done to isolate and identify the etiological agent. Finally, GenoType Mycobacterium CM/AS assay proved it to be a mixed infection caused by Mycobacterium szulgai and M. intermedium. The patient was advised a combination of rifampicin 600 mg once daily, ethambutol 600 mg once daily, and clarithromycin 500 mg twice daily to be taken along with periodic follow-up based upon clinical response as well as microbiological response. We emphasize that infections by NTM must be considered in the etiology of nonhealing wounds or sinuses, especially at postsurgical sites.

Evaluation of GenoType® MTBDRplus assay for rapid detection of drug susceptibility testing of multi-drug resistance tuberculosis in Northern India.

BACKGROUND The problem of multi-drug resistance tuberculosis (MDR-TB) is growing in several hotspots throughout the world. Rapid and accurate diagnosis of MDR-TB is crucial to facilitate early treatment and to reduce its spread in the community. The aim of the present study was to evaluate the new, novel GenoType® MTBDRplus assay for rapid detection of drug susceptibility testing (DST) of MDR-TB cases in Northern India. MATERIALS AND METHODS A total of 550 specimens were collected from highly suspected drug resistant from pulmonary and extra-pulmonary TB cases. All the specimens were processed by Ziehl- Neelsen staining, culture, differentiation by the GenoType® CM assay, first line DST using BacT/ALERT 3D system and GenoType® MTBDRplus assay. The concordance of the GenoType® MTBDRplus assay was calculated in comparison with conventional DST results. RESULTS Overall the sensitivity for detection of rifampicin, isoniazid and MDR-TB resistance by GenoType® MTBDRplus assay was 98.0%, 98.4% and 98.2% respectively. Out of 55 MDR-TB strains, 45 (81.8%), 52 (94.5%) and 17 (30.9%) strains showed mutation in rpoB, katG and inhA genes respectively (P < 0.05). The most prominent mutations in rpoB, katG and inhA genes were; 37 (67.3%) in S531L, 52 (94.5%) in S315T1 and 11 (20%) in C15T regions respectively (P < 0.05). CONCLUSIONS Our study demonstrated a high concordance between the GenoType® MTBDRplus assay resistance patterns and those were observed by conventional DST with good sensitivity, specificity with short turnaround times and to control new cases of MDR-TB in countries with a high prevalence of MDR-TB.

Recent methods for diagnosis of nontuberculous mycobacteria infections: Relevance in clinical practice

Nontuberculous mycobacteria (NTM) infections are ever more important in recent years for leading causes of morbidity and mortality worldwide. Clinical appearance of Mycobacterium tuberculosis (TB) complex and NTM is same, but the treatment regimen is always different. NTM is challenging for both diagnostic and therapeutic with reason that it mimic pathological, microbiological, immunological, and radiological findings of TB. Newer molecular diagnostic methods allow for a better identification of NTM infections in patients not responding to antitubercular treatment and falsely categorized as drug-resistant TB. This article will explore the recent methods for the diagnosis and identification of NTM infections in clinical practice. In the future, the molecular-based diagnosis will significantly reduce the turnaround time of the diagnosis and thereby improving patient outcome.

OL-036 Incidence of multi drug resistant tuberculosis (MDR-TB) in extrapulmonary tuberculosis cases in Northern India

OL-036 Incidence of multi drug resistant tuberculosis (MDR-TB) in extrapulmonary tuberculosis cases in Northern India A.K. Maurya1 *, S. Kant1, V.L. Nag2, R.A.S. Kushwaha1, M. Kumar2, S. Jain2, T.N. Dhole2. 1Department of Pulmonary Medicine, Chhatrapati Shahuji Maharaj Medical University (Erstwhile King George Medical University) Lucknow, India, 2Department of Microbiology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow, India