Hypoxia as a Factor Involved in the Regulation of the apoA-1, ABCA1, and Complement C3 Gene Expression in Human Macrophages.

Hypoxia plays a critical role in progression of atherosclerosis. Local oxygen deficiency in a plaque creates a specific microenvironment that alters the transcriptome of resident cells, particularly of macrophages. Reverse cholesterol transport from plaque to liver is considered a main mechanism for regression of atherosclerosis. Ubiquitously expressed ATP-binding cassette transporter A1 (ABCA1) and liver- and small intestine-derived apolipoprotein A-1 (ApoA-1) are two main actors in this process. We recently reported endogenous apoA-1 expression in human macrophages. While ABCA1 and ApoA-1 have antiatherogenic properties, the role of complement factor C3 is controversial. Plasma C3 level positively correlates with the risk of cardiovascular diseases. On the other hand, C3 gene knockout in a murine atherosclerosis model increases both plaque size and triglycerides level in blood. In the present study, we show for the first time that a hypoxia-mimicking agent, CoCl2, induces the upregulation of the apoA-1 and C3 genes and the accumulation of intracellular and membrane protein ApoA-1 in THP-1 macrophages. The MEK1/2-Erk1/2 and MKK4/7-JNK1/2/3 cascades are involved in upregulation of ABCA1 and C3 via activation of transcription factor NF-κB, which interacts with the HIF-1α subunit of hypoxia-inducible factor 1 (HIF-1). The three major MAP-kinase cascades (Erk1/2, JNK1/2/3, and p38) and the NF-κB transcription factor are involved in the hypoxia-induced expression of the apoA-1 gene in THP-1 macrophages.

[The Spectrum of Mutations in Genes Associated with Resistance to Rifampicin, Isoniazid, and Fluoroquinolones in the Clinical Strains of M. tuberculosis Reflects the Transmissibility of Mutant Clones].

To study the transmissibility of drug resistant mutant clones, M. tuberculosis samples were isolated from the patients of the clinical department and the polyclinic of the Central TB Research Institute (n = 1455) for 2011-2014. A number of clones were phenotypically resistant to rifampicin (n = 829), isoniazid (n = 968), and fluoroquinolones (n = 220). We have detected 21 resistance-associated variants in eight codons of rpoB, six variants in three codons of katG, three variants in two positions of inhA, four variants in four positions of ahpC, and nine variants in five codons of gyrA, which were represented in the analyzed samples with varied frequencies. Most common mutations were rpoB 531 Ser→Leu (77.93%), katG 315 (Ser→Thr) (94.11%), and gyrA 94 (Asp→Gly) (45.45%). We found that the mutations at position 15 of inhA (C→T) (frequency of 25.72%) are commonly associated with katG 315 (Ser→Thr). This association of two DNA variants may arise due to the double selection by coexposure of M. tuberculosis to isoniazid and ethionamide. The high transmissibility of mutated strains was observed, which may be explained by the minimal influence of the resistance determinants on strain viability. The high transmissibility of resistant variants may also explain the large populational prevalence of drug-resistant TB strains.

[[Length polymorphism of minisatellite repeat B2-VNTR of the bradykinin B2 receptor gene in healthy Russians and in patients with coronary heart disease].

Bradykinin B2 receptor is involved in many processes, including the regulation of blood pressure and smooth muscle contraction, vasodilation, inflammation, edema, cell proliferation, pain. It is suggested that this receptor may be one of the factors that have cardioprotective and infarct-limiting effects. It is assumed that certain genetic variants in both coding and non-coding regions ofBDKRB2 gene may influence its expression. In the 3'-untranslated region of BDKRB2 exon 3 the minisatellite repeat B2-VNTR is located. B2-VNTR has previously been shown to affect the BDKRB2 mRNA stability. Therefore, it is important to perform the molecular genetic analysis of this minisatellite in patients with different forms of coronary heart disease in order to reveal possible associations between specific B2-VNTR alleles and certain clinical forms of coronary heart disease. In the present study, a comparative analysis of the allele and genotype frequencies of B2-VNTR was carried out in groups of healthy individuals and patients with two clinical forms of coronary heart disease (angina pectoris and myocardial infarction), ethnically Russian. The results of the B2-VNTR length polymorphism analysis indicate that this tandem repeat may be attributed to a class of low polymorphic and non-hypervariable minisatellite. In all analyzed groups we revealed three B2-VNTR alleles, consisting of 43, 38 and 33 repeat units. Alleles of 43 and 33 repeats were major in all investigated groups. No statistically significant differences were found in the B2-VNTR allele and genotype frequencies between men and women in control group, and also between healthy men and men with angina pectoris and myocardial infarction. Thus, B2-VNTR length polymorphism was not associated with these clinical forms of coronary heart disease in Russian men. However, we do not exclude the possibility of association between the B2-VNTR short alleles (38 and 33 repeats) and cardioprotective effects of bradykinin B2 receptor in women with coronary heart disease. This hypothesis requires further investigation.

Performance of Cepheid ® Xpert MTB/RIF ® and TB-Biochip ® MDR in two regions of Russia with a high prevalence of drug-resistant tuberculosis.

The purpose of this study was to assess the performance of Cepheid® Xpert MTB/RIF® ("Xpert") and TB-Biochip® MDR ("TB-Biochip"). Sputum specimens from adults with presumptive tuberculosis (TB) were homogenized and split for: (1) direct Xpert and microscopy, and (2) concentration for Xpert, microscopy, culture [Lowenstein-Jensen (LJ) solid media and Mycobacteria Growth Indicator Tube® (MGIT)], indirect drug susceptibility testing (DST) using the absolute concentration method and MGIT, and TB-Biochip. In total, 109 of 238 (45.8 %) specimens were culture-positive for Mycobacterium tuberculosis complex (MTBC), and, of these, 67 isolates were rifampicin resistant (RIF-R) by phenotypic DST and 64/67 (95.5 %) were isoniazid resistant (INH-R). Compared to culture of the same specimen, a single direct Xpert was more sensitive for detecting MTBC [95.3 %, 95 % confidence interval (CI), 90.0-98.3 %] than direct (59.6 %, 95 % CI, 50.2-68.5 %) or concentrated smear (85.3 %, 95 % CI, 77.7-91.1 %) or LJ culture (80.8 %, 95 % CI, 72.4-87.5 %); the specificity was 86.0 % (95 % CI, 78.9-91.3 %). Compared with MGIT DST, Xpert correctly identified 98.2 % (95 % CI, 91.5-99.9 %) of RIF-R and 95.5 % (95 % CI, 85.8-99.2 %) of RIF-susceptible (RIF-S) specimens. In a subset of 104 specimens, the sensitivity of TB-Biochip for MTBC detection compared to culture was 97.3 % (95 % CI, 91.0-99.5 %); the specificity was 78.1 % (95 % CI, 61.5-89.9 %). TB-Biochip correctly identified 100 % (95 % CI, 94.2-100 %) of RIF-R, 94.7 % (95 % CI, 76.7-99.7 %) of RIF-S, 98.2 % (95 % CI, 91.4-99.9 %) of INH-R, and 78.6 % (95 % CI, 52.1-94.2 %) of INH-S specimens compared to MGIT DST. Xpert and Biochip were similar in accuracy for detecting MTBC and RIF resistance compared to conventional culture methods.

Molecular typing of Mycobacterium tuberculosis circulated in Moscow, Russian Federation.

The present study investigates epidemiological diversity and multidrug resistance spreading among Mycobacterium tuberculosis strains circulating in Moscow, Russian Federation. Among 115 M. tuberculosis strains selected randomly from the sputum of epidemiologically unrelated tuberculosis (TB) patients, multidrug-resistant (MDR) strains predominated. Mutations in the RRDR of the rpoB gene were detected in 64 (83.1%) of 77 rifampicin (RIF)-resistant strains. The Ser531→Leu substitution was prevalent among them (76.5%). Aberrations in the Ser315 codon of katG and/or in the inhA promoter region were found in 79 (84.0%) of 94 isoniazid (INH)-resistant strains. Strains belonging to the Beijing family prevailed. Seventy-one different patterns were identified using the 24-VNTR loci typing scheme. Three main 24-loci VNTR clusters included 34 strains which belonged to the Beijing family. The spoligotyping and 24-loci VNTR typing combination demonstrated maximal discriminatory power. Among the Beijing strains, the MDR phenotype was revealed more frequently than among the others. High genetic heterogeneity of the studied population was shown by the assessment of VNTR loci variability in the analyzed group and in the strains from other parts of Russia. Comparison of the 24-VNTR locus typing and spoligotyping data with revealed resistance-associated mutation allows us to make a suggestion that the active transmission of MDR strains and the independent appearance of drug resistance during chemotherapy occurred in the studied population simultaneously.

[Regulatory effect of complex of natural cytokines and antimicrobial peptides on murine macrophages infected with multidrug-resistant Mycobacterium tuberculosis].

AIM: To study the effect of complex of natural cytokines and antimicrobial peptides (CNCAP) included in preparation Superlymph on growth of multidrug-resistant Mycobacterium tuberculosis CN-37 on the modem of murine peritoneal macrophages (MPh) cultivated in vitro. MATERIALS AND METHODS: Effect of CNCAP on peritoneal MPh of tuberculosis-susceptible mice C57BL/6 infected by M. tuberculosis CN-37 was studied using ex vivo model. Macrophages were preliminary incubated with CNCAP during one day. M. tuberculosis growth was assessed on 7th day by PCR. RESULTS: Preliminary incubation of infected MPh with CNCAP resulted in inhibition of M. tuberculosis CN-37 growth. CONCLUSION: Superlymph activates macrophages which lead to enhanced bactericidal action of MPh on M. tuberculosis CN-37.

[Detection of mutations in codon 306 of the embB gene for molecular genetic characterization of clinical Mycobacterium tuberculosis strains].

A total of 254 Mycobacterium tuberculosis strains were used in the study. Among them, there were 183 ethambutol (EMB)-resistant strains, 13 multidrug resistant ones, but EMB-sensitive, and 39 strains sensitive to rifampicin (RIF), isoniazid (INZ), and EMB. All the strains were analyzed for genetic changes in three loci: embB306, rpoB, and katG/inhA promoter, which were associated with the formation of resistance to EMB, RIF, and INZ, respectively. The Mycobacterium tuberculosis strains were obtained from pulmonary tuberculosis patients living in the Central Region of the Russian Federation. Resistance to RIF, INZ, and EMB was revealed by the absolute concentration test. The inhibitory concentration (IC) of EMB was determined for all the strains. Genetic changes in the above loci were estimated by mini-sequencing, followed by mass-spectrometry recording MALDI-TOF products. The relative low frequency of embB306 mutations was observed among the EMB-resistant strains (about 41.5%). Mutations in codon 306 were detected only in strains with EMB IC > or = 2 mg/L. A statistical significant association was found between the frequency of embB306 mutations and the multidrug resistant phenotype. A combination of these mutations with the traditional genetic markers of multidrug resistance may be used for the more effective detection of multidrug-resistant strains.

[Biological properties of M. Tuberculosis W cluster strains].

To study the biological characteristics of M. tuberculosis W cluster strains, the authors carried out in vitro, ex vivo, and in vivo experiments on 18 clinical strains and 2 laboratory ones, which had been clustered by a standardized methodology. Comparison was made in experiments in vitro and in macrophageal inoculation (ex vivo) from the 5,6-[3H]-uracil incorporation that reflected mycobacterial replication. The in vivo experiments estimated mycobacterial survival and cultivation and the lung pathomorphological pattern of infected animals. The results showed that M. tuberculosis W cluster strains had in vitro the mean multiplication rate, showing a high viability in the macrophages; the in vivo experiments demonstrated that the W cluster strains did not differ in virulence and might be both more and less virulent than the laboratory strain H37Rv. Notwithstanding the fact that the assumption on the hypervirulence of M. tuberculosis W cluster has not been supported, the distinguishing characteristic of the strains of this cluster is their enhanced capacity to survive in the macrophages no matter what infective dose.

Detection of mutations in Mycobacterium tuberculosis genome determining resistance to fluoroquinolones by hybridization on biological microchips.

We developed a method of identification of Mycobacterium tuberculosis with simultaneous evaluation of the sensitivity to fluoroquinolones on a biological microchip array. The method of multiplex two-staged PCR followed by hybridization of a biochip makes it possible to detect 8 mutant variants of gyrA gene occurring in fluoroquinolone-resistant strains (approximately 85% all resistant forms) within 1 day. Using this method we analyzed 107 cultures isolated from patients with tuberculosis and 78 sputum samples. Mutations in gyrA gene were detected in 48 (92%) resistant strains. Natural S95T polymorphism in gyrA gene was detected in all resistant and in 76% sensitive strains. The sensitivity and specificity of the proposed method calculated on the basis of the analysis of sputum samples (n=78) were 94 and 100%, respectively.

[Inhibiting effect of complex of natural cytokines and cationic antimicrobial peptides on the growth of Mycobacterium tuberculosis H37Rv in vitro].

AIM: To assess direct antimicrobial effect of complex of natural cytokines (CNC) and antimicrobial peptides (Syperlymph preparation; CNC) on Mycobacterium tuberculosis H37Rv (H37Rv) and effect mediated by macrophages (MP) treated with the preparation. MATERIALS AND METHODS: Direct effect of CNC was studied during cultivation of H37Rv in the presence of preparation, whereas indirect effects--during simultaneous cultivation of H37Rv and mice peritoneal MP C57B1/6. Assessment of growth was performed on the 7th day using PCR. RESULTS: It was shown that CNC directly inhibits growth of H37Rv in vitro. Cultivation of H37Rv in culture of MP resulted in inhibition of M. tuberculosis. The most evident inhibition was noted after extension of time of preliminary treatment of MP with Syperlymph and simultaneous increase of its concentration. CONCLUSION: Antimycobacterial effect of Syperlymph preparation related to complex effect of cytokines and antimicrobial peptides directed to M. tuberculosis and macrophages, which forms the conditions for killing of mycobacteria.

[Impact of M. tuberculosis genotype on survival in mice with experimental tuberculosis].

To study the virulence of Mycobacterium tuberculosis strains of the W cluster, C57B1/6 mice were intravenously inoculated with a lethal dose (5x10(6) CFU) of 14 clinical M. tuberculosis strains (11 drug-sensitive and 3 multidrug-resistant) belonging to different RFDP IS6110 genotypic clusters and two laboratory M. tuberculosis strains H37Rv and H37Ra. The virulence was evaluated by the survival of mice after infection and by the trends in body weight loss. The study indicated that the mice inoculated with different M. tuberculosis strains differed in survival rates and in the trend in body weight loss. A minor HD cluster strain and 2 non-clustered strains were most virulent, next were 2 AI cluster strains. W cluster strains had both higher (n = 2) and lower (n = 3), and comparable (n = 2) H37Rv virulence. A KQ cluster strain had the least virulence. An attenuated H37Ra strain caused no animal death. Inoculation with three multidrug-resistant strains belonging to the W cluster (n = 2) and one non-clustered strain demonstrated no relationship of virulence to the sensitivity of a strain to antituberculous agents. The findings argue against the opinion on W cluster M. tuberculosis strains as hypervirulent.

[Mycobacterium tuberculosis strain transmission caused by migratory processes in the Russian Federation (in case of populational migration from the Caucasian Region to Moscow and the Moscow Region)].

The investigation was carried out on 134 M. tuberculosis isolated from 134 patients treated at the Central Research Institute of Tuberculosis, Russian Academy of Medical Sciences. The patients were divided into 2 groups: 1) those who were natives of Moscow and the Moscow Region (MR patients); 2) those who were migrants to the Moscow Region from Azerbaijan, Daghestan, Chechnya, Ingushetia, Karachai-Cherkessia, North Ossetia (the Caucasian Region) (CR patients) who had fallen in the place of birth. Genotyping by the polymorphism of lengths of the restriction fragments containing the insertion sequence IS6110 revealed a genetic diversity of M. tuberculosis strains. The examined M. tuberculosis strains belonged to 13 genotypic families. The W and AI families were prevalent. The family W M. tuberculosis strains isolated from the Caucasians were highly clustered, as confirmed by the overwhelming predominance of the strain variant W148 (19.7%). The spectrum of the strain variants of the W family, and those of the AI family in particular, greatly differed in MR and CR patients. Only one strain variant AI12 occurring both in MR and CR patients was detected. A study of the transmission activity coefficient (TAC) of the families W and AI indicated that the transmission activity of W strains was significantly higher than that of M. tuberculosis strains of the AI family. A comparative analysis of the TAC of M. tuberculosis strains of the AI family demonstrated that the transmission activity of the strains of this family was identical no matter where a patient had fallen ill (1.59 and 1.41% in the Moscow and Caucasian Regions, respectively). Unlike M. tuberculosis strains of the AI family, the TAC of W strains isolated from the patients infected in the Moscow Region (28.17 and 19.05%, respectively), which suggests the more intensive transmission of the pathogen M. tuberculosis of the W family in the Caucasian Region.

[Examining the ex vivo growth in the macrophages of Mycobacterium tuberculosis of various genotypic clusters].

To study the specific features of replication of Mycobacterium tuberculosis (MBT) strains of the W cluster in the macrophages (MP) on the ex vivo model of peritoneal MP of MBT-infected C57B1/6 mice, the authors estimated the viability of 6 antituberculous drug-sensitive MBT strains of the W-cluster from the incorporation of 5,6-[3H]-uracil into the mycobacterial cells and their induced specific MP from the level of LDH. Eight sensitive MBT strains of other genotypes clustered by the restriction fragment length polymorphism (PDRF) IS6110 and 2 laboratory strains M. tuberculosis H37Rv and M. tuberculosis H37Ra were taken as a control. The study indicated that, cultured in vitro, MBT strains belonging to different genotypic clusters differed in the level of 5,6-[3H]-uracil inclusion. When grown in MP, the MBT population of all genotypes showed a diminished viability as compared with that cultured without MP. The MBT clusters of W and AI clusters, unclustered strains, and M. tuberculosis H37Rv displayed a higher inclusion of 5,6-[3H]-uracil and the strains of KQ and HD clusters and M. tuberculosis H37Ra exhibited a significantly lower inclusion of 5,6-[3H]-uracil. W-cluster strains, the unclustered strain R807, and M. tuberculosis H37Rv showed the highest fitness (adaptability when grown in MP). The virulent strain M. tuberculosis H37Rv and avirulent strain M. tuberculosis H37Ra differed in MP viability by almost 5 times. Evaluation of the cytopathogenic effect indicated that the clinical MBT strains led to a specific MP lysis greater than 40%, the highest effect was produced by the MBT strains of the W cluster (more than 93%) and the HD cluster (96.45%). The control laboratory strains M. tuberculosis H37Rv and M. tuberculosis H37Ra contrasted sharply in their induced specific MP lysis (93.35 and 5.93%, respectively).

[The association of tuberculosis with the specificities of the HLA gene DRB1 in different regions of Tuva].

The fact that genetic factors largely determine susceptibility to different diseases, including those of infectious nature is beyond question now. Tuberculosis is not an exception in this respect. HLA genes that determine different immunological phenomena make a certain contribution to tuberculosis susceptibility. This paper presents the results of typing using the polymerase chain reaction from the specificities of the HLA gene DRB1 in patients with pulmonary tuberculosis and healthy individuals in different regions of the Republic of Tuva. The studies in these regions of Tuva have revealed a significant positive association of tuberculosis with the specificities of HLA DRb1 13(6) and HLA DRB1 14(6). Analyzing 14 families of patients with tuberculosis has shown that HLA haplotypes from the sick parents who carry the specificities of HLA DRB1 13 and/or DRb1 14 are more frequently transmitted to sick children than to healthy ones. High morbidity in the indigenous dwellers of the Republic of Tuva may be associated with these specificities of the HLA gene DRB1, which is due to the national peculiarities of the native population of this republic.

[Genotypic characteristics of Mycobacterium tuberculosis strains from the Republic of Tyva]. / Genotipicheskaia kharakteristika shtammov Mycobacterium tuberculosis iz Respubliki Tyva.

To determine the genetic heterogenicity of Mycobacterium tuberculosis isolated from patients with tuberculosis from different districts of the Republic of Tyva, 71 M. tuberculosis strains IS6110 according to the polymorphism of restriction fragment lengths were typed; of them 32 strains were spoligotyped. The study could genetically characterize the groups of M. tuberculosis strains circulating among patients with tuberculosis in the Republic of Tyva. There was a predominance of the W family of mycobacteria (60.56%). The spoligotyping permitted identification of M. tuberculosis with the spoligotype Beijing in the Republic of Tyva. The mycobacteria belonging to this spoligotype were prevalent and accounted for 50% of the strains. The high proportion of strains with the unique spoligotype (25%) that have not earlier been encountered in the international database is a specific feature of the Mycobacterium population from the Republic of Tyva.

[Comparative characterization of molecular and microbiological methods for controlling chemotherapy in new cases of pulmonary tuberculosis]. / Sravnitel'naia kharakteristika molekuliarnykh i mikrobiologicheskikh metodov kontrolia khimioterapii u vpervye vyiavlennykh bol'nykh tuberkulezom legkikh.

The purpose of the study was to define a role of molecular genetic techniques in the evaluation of the efficacy of short-term chemotherapy in patients with pulmonary tuberculosis. A total of 203 patients with first detected tuberculosis patients who isolated Mycobacterium tuberculosis (MBT) were examined. Microbiological methods (bacterioscopy and seeding) and polymerase chain reaction (PCR) were used to detect MBT in new cases. During chemotherapy, there was a reduction in the number of positive results of both bacterioscopy and PCR, which serves as a sign of abacillation. Following 2 months of chemotherapy, 50% of the patients with negative bacterioscopy and seeding were found to have MBT DNA. In the patients with negative bacterioscopy and seeding, the results of PCR were also negative by month 6. In a group of patients without bacterial isolation, but with positive sputum tests for MBT DNA, the efficiency of chemotherapy was confirmed by the negative results of PCR. The latter is shown to enhance the sensitivity of bacteriological tests for MBT and to reduce the time of obtaining their results.

[Significance of molecular genetic and immunological techniques in risk groups and in children with tuberculosis]. / Znachenie molekuliarno-geneticheskikh i immunologicheskikh metodov issledovaniia v gruppakh riska i u bol'nykh tuberkulezom detei.

A total of 188 children and adolescents were examined. In all the children, blood Mycobacterium tuberculosis (MBT) DNA was determined by polymerase chain reaction (PCR) and MBT antigens (AG) and antibodies (AB) were by enzyme immunoassay. The studies have shown that it is expedient to concurrently determine MBT DNA and MBT AT in order to identify local forms of tuberculosis in children from risk groups. If the tests are positive, a comprehensive examination for tuberculosis is required; the presence of the syndrome of common disturbances is generally associated with tuberculous infection. When a local form of tuberculosis is excluded, preventive chemotherapy should be performed. Further negative tests for MBT DNA and lower MBT AT may be a criterion for the efficiency of preventive treatment. In children with tuberculosis, the results of repeated blood and urine tests for MBT DNA provide a way of evaluating the course of a tuberculous process and the efficiency of chemotherapy. PCR used to determine blood and urine MBT DNA is a highly specific test as positive results were in 79% of the children with tuberculosis.

[Role of PCR analysis in complex bacteriological studies in phthisiology]. / Rol' PTsR-analiza v kompleksnykh bakteriologicheskikh issledovaniiakh vo ftiziatrii.

The study was undertaken to assess the role of polymerase chain reaction (PCR) analysis in complex bacteriological studies in diagnosing pulmonary tuberculosis in 197 patients by bacterioscopy, inoculation, and PCR. It was shown that in addition to conventional bacteriological methods, PCR might be used as an additional laboratory study in making a diagnosis in patients with restrictive pulmonary tuberculosis. A combination of cultural inoculation and PCR analysis enhances the sensitivity of bacteriological diagnosis and reduces its duration in oligo- and abacillary patients with tuberculosis. PCR analysis enhances the efficiency of laboratory control over antituberculous chemotherapy.

[Molecular characteristics of multiresistant clinical strains of Mycobacterium tuberculosis isolated in Russia]. / Molekuliarnaia kharakteristika polirezistentnykh klinicheskikh shtammov Mycobacterium tuberculosis iz Rossii.

The efficiency of tuberculosis control programs is largely determined by methods for rapid diagnosis of the agent. In comparison with the traditional methods, new molecular technologies for characterization of mycobacteria appear to be more promising, because the result can be obtained in almost no time. Sixty-five strains of M. tuberculosis isolated in various regions of Russia were investigated. Drug resistance and strain appurtenance of this sample were determined by classical (absolute concentrations method, IS6110-RFLP) and modern molecular genetic methods (detection of mutations in rpo B gene, DRE-PCR). The spectrum of mutations of the rpoB gene associated with rifampicin resistance was evaluated by direct sequencing. Mutations involving codons 531 (62.7%), 526 (18.6%), and 516 (10.2%) of rpoB gene predominated in the studied sample. The studied strains were discriminated into 52 individual strains by IS6110-RFLP and DRE-PCR typing. Analysis of the resultant genetic variants showed the predominance of M. tuberculosis family W. The efficiency of combined approach to screening for M. tuberculosis is discussed.