Background and Objective: Identification of atypical mycobacterium (Non tuberculosis Mycobacterium NTM) is important because of the worldwide propagation of these organisms. Recently, molecular studies have identified the specific loci for mycobacterium species by DNA - finger printing methods, but these methods are time-consuming and expensive. In this study, in addition to hsp65 PCR-RFLP method, QUB3232 locus was evaluated for differentiation of atypical mycobacterium from mycobacterium tuberculosis complex. Materials and Methods: This study was performed on 371 pulmonary and non pulmonary specimens separated from patients with the symptoms of pulmonary tuberculosis (PTB). After the isolation and culturing of mycobacterium strains using the Lowenstein Jensen media, biochemical tests including production of Niacin, Catalase activity, Nitrate reduction, pigment production and growth rate were performed. Drug susceptibility testing was performed by proportional method. DNA extraction was performed by phenol-chloroform method. hsp65 gene was amplified by PCR. Subsequently the amplicons were digested with three restriction enzymes namely AvaII, HphI and HpaII and electrophoresed on 3% agarose gel. QUB3232 locus was also evaluated for differentiation of atypical mycobacterium and mycobacterium tuberculosis complex. Results: Out of 371 isolates, 32 (8.6%) were multi-drug resistant TB (MDR-TB), 184 (49.5%) were susceptible and 155 (42.5%) were non MDR (combined resistance) that 15% of MDR cases and 25% of non MDR cases were non tuberculosis mycobacterium. Out of 31 slow growing isolates, 58% were M. simiae and 19% were M. kansasii. The sensitivity of QUB3232 locus for differentiation of the atypical mycobacterium from mycobacterium tuberculosis complex was 80%. From the total of 43 NTM samples, 12 (27.9%) were rapid growing and 72% were slow growing. Conclusion: QUB3232 locus has the high discriminative power for differentiation of atypical mycobacterium from the mycobacterium tuberculosis complex, therefore, it can be used as a substitute for PCR-RFLP method.

Background and Objective: Spoligotyping is a method based on 36bp Direct Repeat (DR) chromosomal loci polymorphism which is connected to one or two 35-41 bp spacer sequences. There are 94 different intra DR spacer sequences which are identified so far and only 43 of them are used as usual. Mycobacterium tuberculosis complex strains can be identified based on lacking or having these sequences. Materials and Methods: Spoligotyping test was carried out on 238 TB smear positive patients. Primary separation of mycobacterium strains was done through Petrof 4% method and Lowenstein Jensen (LJ) media. Biochemical tests such as Niacin test/Catalase activity/Nitrate reduction were done in order to identify the strains. Drug sensitivity to INH (0.2Mg/ml)/ RIF (40Mg/ml)/ STM (10Mg/ml) and ETBl (2Mg/ml) identified by proportional method and according to that, the strains were divided into three groups: sensitive, multi drug resistance (MDR) and non MDR. Then DNA was extracted by CTAB method from the positive colonies. Sequences were amplified by PCR and after denaturizing, hybridization with Streptavidine peroxidase enzyme was performed by Line reverse blot method. Radiography was done after adding the Luminoscense and membrane onto the X-ray films. Results: Serotypes were divided into 9 groups (Beijing/ CAS1/ Haarlem / U/ T2/ T1/ EAI3/ EAI2 and CAS2). Most of the strains were from Haarlem (27%) and CAS1 (25%) groups. Two strains were also identified in this method that belonged to Mycobacterium bovis. Conclusion: Spoligotyping method is an easy, rapid and sensitive test in order to identify Mycobacterium tuberculosis complex strains.

Background and Objective: Tuberculosis (TB) caused by Mycobacterium tuberculosis, is an infectious disease in human which kills nearly three millions of people annually. Approximately, one - third of the world populations are infected with this bacteria and 5 - 10 % of them develop the active form of the disease. Individuals are different in susceptibility to TB infection. These differences might be due to the host characteristics especially genetic factors. TNF-α as a pro-inflammatory cytokine, play a key role in host defense against tuberculosis. Presence of mutation in this gene can influence the effectiveness, performance and capability of immune responses against TB infection. The Aim of this study was to investigate the frequency of TNF-α gene polymorphisms and its relation with susceptibility to the pulmonary TB. Materials and Methods: Sixty healthy controls and 60 TB patients were enrolled. Genotype of TNF-238, TNF -244, TNF-308, TNF -857 and TNF-863 were determined using PCR-RFLP method. The results were analyzed by Fisher Exact and χ 2 tests using SPSS v.14 and evaluated with Hardy–Weinberg equilibrium. Results: The results of this study showed a significant difference in TNF-308 and TNF -857 regions between the control and study groups (P < 0.05). Conclusion: Presence of mutation in TNF-308 and TNF -857 regions may increase the host susceptibility to mycobacterium tuberculosis and genotyping of these regions can be used for screening of the high risk individuals.

Background and Objective: In recent decades, epidemiology has significantly been considered in hygienic studies and disease control, and has made a way into all the programs and hygiene policies. By examining the convergence of harmful lineage genetic patterns, the common infectious resources among the patients can be inferred. The purpose of this study was to compare the Mycobacterium Tuberculosis genetic patterns convergence isolated from patients infected with Mycobacterium Tuberculosis by MIRU-VNTR technique.
Materials and Methods: After isolation the samples from Lowenstein Jensen culture environment and taking segregate tests and drug susceptibility, the DNA was extracted using CTAB/Nacl technique. The genetic patterns of lineages were calculated according to 12 loci format with MIRU-VNTR technique. Demographic and molecular information of patients was used for epidemiological purposes.
Results: After performing drug sensitivity test, 65/140 (64/4%) samples fall into MDR, 29 (20/7%) samples in non MDR category, and the rest of them were among drug – sensitive lineages. Lineage genetic pattern analysis indicated that 49 (35%) of samples related to Delhi/CAS, 28 (20%) to Uganda I, 16 (11/4%) to New I, 1 (0.7%) to EAI, 3(2/1%) to Haarlem, and 5(3/5%) to H37RV families.
Conclusion: The genetic pattern convergence comparison exhibited that the most common and variant genetic patterns was seen in Tehran province which were mostly connected to south (from the South of Tehran to Azadi Square) and to the border cities neighboring Afghanistan, Iraq, Turkmenistan and cities with extreme percentage of immigration, all of which signified shared polluted resources among patients.