Drug Resistance Patterns of Mycobacterium tuberculosis – Isolates from Indore, India

Introduction: Tuberculosis (TB) is an Infectious disease existing pandemically in our world.In parallel, the prevalence of multidrug-resistant tuberculosis (MDR-TB) is also increasing. The TB control programs were not successful due to the emergence of multidrug resistance in M. tuberculosis strains. Objective of the present study was to detect the rate of MDR-MTB in the central state of India. Materials and Methods: The study included all new & old cases of pulmonary & extra pulmonary tuberculosis enrolled between January 2013 and December 2014 carried out in our Indore, lab, India. All the patients’ samples found to be TB positive in our lab were tested by Mycobacterium culture and MTB isolated strains were put for first-line drug-susceptibility testing (DST). MDR-TB was defined as TB caused by bacilli showing resistance to at least isoniazid and rifampicin. Results: A total of 60 MTB isolated strains; fourteen were MDRs (23.3%). Resistance to INH, RIF, PYRA, ETHAM,streptomycin (STREPTO) was found to be 41%, 36.6%, 23.3%, 30%, 25% respectively. Conclusions: MDR-TB prevalence was found to be high among both new & old cases of pulmonary & extra pulmonary tuberculosis cases. Nation-wide and State-wide representative data on prevalence of MDR-TB are lacking in the state of Madhya Pradesh, in India. Efforts needs to be directed towards further continuous surveillance for MDR-TB among newly diagnosed TB cases and old diagnosed cases.


INTRODUCTION
Tuberculosis is a common, though preventable infectious disease of the world. It is known that India accounts for about 30 per cent of the total global TB burden [1] Currently, tuberculosis management and control is potentially devastating threat worldwide due to Did not profve to be successful though the therapy. Thus lead to formation of drug resistance bacteria spread in population there by entitling MTB Strains as manmade development of resistance as a consequence of suboptimal regimens and treatment interruptions by patients themselves [2].
Drug resistance in TB is classified in group of primary or that of acquired type when drug resistance is demonstrated in a patient, who has never received anti-TB treatment previously, it is termed primary resistance. Acquired resistance is that which occurs as a result of specific previous treatment [3].
Resistant to at least two major anti tuberculosis drugs; INH and RIF with or without resistance to other anti-TB drugs has been termed MDR-TB. MDR-TB is more difficult to treat than drug susceptible TB, requiring the use of less effective second line anti tubercular drugs which are often associated with major side effects [4].
Recently, WHO has estimated that around 3.7% of new TB cases are developed as MDRs. MDR-TB global average rate is 20%. About 9% of these cases also are resistant to at least one injectable second line antitubercular drugs. Extensively drug resistant (XDR) TB cases as reported earlier [5].
Major concern over drug resistance is a fear of spread of drug resistant organisms and ineffectiveness of chemotherapy in patients infected with them. The distribution and rate of MDR and XDR-TB are not uniform and vary in different places, regions, populations and countries. Limited data are available on the trends of prevalence of drug resistance in Indore region and therefore the find existence of Drug Resistant Mycobacterium. In our city in the Central State of India during the years 2013-2014.

Study Setting
This study was done at Central Lab Onquest, Indore (M.P.), which is a private NABL accredited lab; the data presented are from January 2013 to December 2014.
We tested a total of 440 samples from all the age groups of clinical TB suspects of TB suspects from higher socioeconomic structure capable to efford private lab Indore & its surrounding region. Patient demographic data like age, gender, address were obtained. We received &processed all the pulmonary or extra pulmonary samples. As our lab is private lab, some of the data some samples are submitted without prior history of patient. so we received direct samples in our lab and status of all patients whether new or old treated cases are not received in all cases. The patient samples were studied by viewing morphology using staining and under microscope, culture, along with culturing them identification and Drug Sensitivity testing (D.S.T). All tests were performed using patient's consentunder Medical supervision, according to guidelines for Good Clinical laboratoy practice (GCLP) in our NABL (National board for accreditation for testing and calibration for laboratory. Samples obtained in the laboratory in a cold box and were processed on the same day or were kept at +4 degrees Celcius. in refrigerator, until their processing was done. The tissue samples were first decontaminated using N-Acetyl-L-Cysteine, 2 % S od i u m h ydr o xid e and sodium citrate, PBS (pH 6.8). After they were decontaminated the smears were stained by Ziehl-Neelsen method and examined by trained technical staff for acid fast bacilli. and cultures were done on the Lowenstein -Jensen (LJ) egg based medium were carried out as per the standard methods.
All sample were processed according as instructed (Hi-Media) on the readymade Lowenstein-Jensen (LJ) medium (Hi-Media) and incubated for 8 weeks. All the isolates were identified as M. tuberculosis by their slow growth rate, colony morphology, inability to grow on L-J media containing p-nitrobenzoic acid (PNB), niacin and catalase test [6-8] DST was carried out on ready prepared Hi-media LJ medium first line drug kit according to kit information (Proportion method). First line drug include INH, RIF, PYRA, ETHAM and STREPTO. MDR-TB was defined as TB caused by bacilli showing resistance to at least INH and RIF. An uninoculated tube of LJ medium was used as negative control and M. tuberculosis A.T.C.C. H37Rv was used as positive control.

RESULTS
We found Out that of small number of test samples of 440 samples, relatively small number, 60 showed growth of Mycobacterium tuberculosis isolates whereas 3 were identified as atypical mycobacteria (MOTT).   Among 60 isolates in our lab, 33 were from male patients and 27 were from female patients. Maximum isolates were found in the age group 30-40 years followed by 40-50 years as shown in the Table 2.

DISCUSSION
Increasing drug resistance to commonly used drugs Rifampicin (RIF) and INH in isolates of M. tuberculosis is a major cause of concern. Because of these trends it is important to know the rate of primary and acquired drug resistance in an area at regular intervals.
Drug resistant tuberculosis is either acquired due to poor management of treatment or transmission from infectious drug resistant TB patients. As found in many other studies history of antitubercular treatment has been consistently associated with risk of MDR-TB [10]. One most important limitation of this study is previous treatment histories, and information about second line drug susceptibility like quinolones were not available being a private lab for analysis, restricted our ability to derive concrete conclusions also including outdoor patients' tests done by our specialized clinicians in our lab. Present data also limits as representative of the whole community and are limited to few hospitals and outdoor registered patients for diagnosis. Our data can be well added to the community based multicenter study, from indore, with other studies including all parts of the country using the full spectrum of drugs, to describe the true prevalence of MDRTB in the state and country.
The pattern of drug resistance differs from place to place and at different periods of time. For a drug regimen to be effective with complete restriction of further growth or developing further resistant variations of Mycobacterium, it is important to know the drug resistance pattern of existing and spreading in that area, that needs to be conducted in periodic surveys of antibiotic drug resistance and to establish a continuous drug resistance surveillance program. As several of the studies cited in here including our present, study may have flaws with sampling, carefully planned long term studies with sufficient quality assurance are further required.

CONCLUSION
The study reported herewith lacks the detailed information about the previous treatment histories of medical treatment of patient and the information about second line drug susceptibility of Mycobacterium in the populations, which were not available for analysis, restricted us to derive concrete conclusions. Other limitations are of data as they are not representative of the whole community and are limited to few hospitals of Indore. A community based multicenter study, which includes all parts of the country and uses the full spectrum of drugs, is needed to further describe the true existence of MDRTB in Country

CONSENT
It is not applicable.