[Molecular markers of M. tuberculosis virulence in lung tissue (experimental study)]. / Molekulyarnye markery virulentnosti M. tuberculosis v tkani legkikh (eksperimental'noe issledovanie).

More than a quarter of the world's population is infected with Mycobacterium tuberculosis. However, only about 10% of those infected develop active TB. This indicates a key role for innate immunity in limiting M. tuberculosis replication. Most often, bacteria can regulate the expression of host-specific molecules and weaken host immunity. OBJECTIVE: To use a biological model, in order to determine significant molecular immunohistochemical markers characterizing the virulence of the "Buryat" and "Omsk" subtypes of the M. tuberculosis Beijing genotype in lung tissue. MATERIAL AND METHODS: Lung samples of the C57BL/6 male mice were obtained during experimental infection with M. tuberculosis strains: the reference laboratory strain H37Rv, multidrug-resistant clinical strains 396 (highly lethal and hypervirulent «Buryat¼ genotype Beijing 14717-15) and 6691 (low-lethal and low-virulent "Omsk" genotype Beijing 1071-32) on days 14, 21, 60 and 120. They were studied by histological and immunohistochemical methods. The relative areas of expression of IL-6, IL-12A, iNOS, and TNF-α in the lung tissue of model animals were established. RESULTS: A study of strain 396 showed that both disease progression and damage to lung tissue are associated with a highly reactive immune response and increased synthesis of iNOS and strain characteristics that block the production of TNF-α. On the contrary, for strain 6691 a low reactivity of the immune response was revealed, with statistically significantly lower values of the relative area of expression of NOS and TNF-α during all observation periods (days 14-120). All animals that survived to day 120 showed a similar morphological picture with differences in cytokine levels, indicating a nonlinear relationship between proinflammatory factors and the damage substratum. CONCLUSION: The progression of the disease and damage of lung tissue were associated with a highly reactive immune response and increased synthesis of iNOS, strain properties that block the TNF-α production. Thus, iNOS and TNF-α can act as molecular markers characterizing the virulence of the "Buryat" and "Omsk" subtypes of M. tuberculosis in lung tissue.

Inhibition of the Expression of Inducible NO Synthase by Neuroactive Amino Acid Derivatives Phenibut and Glufimet In Vitro and Ex Vivo.

The effects of glufimet and phenibut (glutamic acid and GABA derivatives, respectively) on concentration of inducible NO synthase and cGMP in LPS-activated mouse peritoneal macrophages and on NO end products in their culture medium were examined in vitro and ex vivo. Addition of LPS into culture medium elevated concentration of NO metabolites in this medium and increased concentration of inducible NO synthase and cGMP in the lysates of peritoneal macrophages, whereas incubation of the cells with examined agents applied at concentration of 10-5 M diminished these indices. Similar results were obtained with intraperitoneal injection of LPS, glufimet, and phenibut. In culture medium containing peritoneal macrophages from the mice injected with LPS (100 µg/kg), the concentrations of inducible NO synthase and cGMP as well as the total concentration of nitrite and nitrate ions increased, whereas in culture medium with the cells from LPS-exposed mice treated with glufimet (28.7 mg/kg) and phenibut (50 mg/kg) these indices significantly decreased.

Effect of Phenibut and Glufimet, a Novel Glutamic Acid Derivative, on Respiration of Heart and Brain Mitochondria from Animals Exposed to Stress against the Background of Inducible NO-Synthase Blockade.

Increased oxygen consumption by heart and brain mitochondria in the absence of ADP and reduced mitochondrial respiration in the presence of ADP were observed in rats exposed to stress simulated by suspension by the dorsal neck skin fold for 24 h, which attests to uncoupling of substrate oxidation and ATP synthesis and can cause electron drain from the respiratory chain, formation of ROS, and oxidative damage to cell structures. Blockade of inducible NO synthase with aminoguanidine (single intraperitoneal dose of 50 mg/kg before stress exposure) increased coupling of respiration and oxidative phosphorylation in heart and brain mitochondria of rats exposed to immobilization-painful stress, which was especially pronounced in cardiomyocytes. The test compounds glufimet (single intraperitoneal dose of 29 mg/kg before stress exposure) and phenibut (single intraperitoneal dose of 50 mg/kg before stress exposure) limited stress-induced mitochondrial damage against the background of inducible NO synthase blockade and without it, which was seen from increased respiratory control ratio in comparison with that in untreated rats exposed to stress (control).

[MOLECULAR CHARACTERISTICS OF THE MULTIDRUG-RESISTANT MYCOBACTERIUM TUBERCULOSIS STRAINS IN THE NORTHWEST RUSSIA].

The goal of this work was to study the genotypic characteristics of the multidrug-resistant (MDR, i.e., resistant to at least rifampicine and isoniazid) Mycobacterium tuberculosis strains isolated in 2011-2012 from tuberculosis (TB) patients in the Northwest Russia. Spoligotyping of 195 M. tuberculosis isolates identified 14 different spoligotypes and assigned isolates to the genetic families Beijing (n = 162, 83%), LAM (n = 15), H3/URAL (n = 14), as well as T, Haarlem and X. Spoligotypes SIT1 (Beijing), SIT42 (LAM) and SIT262 (H3/URAL) were the most prevalent. Irrespective to the genotype, all the isolates were resistant to streptomycin. The multidrug resistance was accompanied by the resistance to ethionamide (56%), amikacin (31%), kanamycin (40%), and capreomycin (33%). The ethambutol resistance was found in 71% (n = 115) and 42% (n = 14) of the Beijing and non-Beijing strains, respectively (p < 0.05). In conclusion, the multidrug resistant M. tuberculosis population circulating in the Northwest Russia continues to be dominated by the Beijing family strains.

[The genome polymorphism of the Mycobacterium avium subsp. hominissuis strains].

The non-tuberculosis mycobacteria Mycobacterium avium subsp. hominissuis (MAH) are able to cause human mycobacteriosis. In this work, the results of the first comprehensive study of the genome polymorphism of the clinical strains of MAH were reported using the typing scheme by 13 loci MATR-VNTR (TR292, TRX3, TR25, TR47, MATR-1, MATR-4, MATR-5, MATR-6, MATR-8, MATR-11, MATR-14, MATR-15, MATR-16) containing tandem nucleotide sites and IS1245-RFLP-typing sites. A total of 90 MAH strains isolated from patients with lung mycobacteriosis without epidemiological connection (including HIV infected) were tested in 2008-2011. The inhomogeneity of the MAH strains by 36 profiles of 13 loci MATR-VNTR was observed. The majority of the strains (68.8%) were included in the 8 MATR-VNTR clusters; most large cluster contained 37 strains with 13-bitnumerical profile 2222223145443'. The nucleotide sequence of the MATR-16 (3') locus contains the long deletion (GenBank accession no. KF479191). The MAH strains of the MATR-VNTR clusters were found to be inhomogeneous by the IS1245 marker. The MATR-VNTR-typing method by 13 loci is recommended for preliminary differentiation of domestic MAH strains with further analysis of the MATR-VNTR clusters using the IS1245-RFLP-typing method.

Effect of GABA derivatives on the rate of thrombus formation, platelet aggregation, and plasma coagulation capacity in rats with experimental gestosis.

Experimental gestosis induced by replacement of drinking water with 1.8% NaCl promoted hypercoagulation, increased the rate and degree of platelet aggregation, and reduced clotting time in pregnant females. GABA derivatives, compounds RGPU-151, RGPU-152, and phenibut normalized parameters of hemostasis and platelet aggregation and the rate of thrombus formation in the animals. The efficiency of the test substances did not significantly differ from that of the reference drug sulodexide.

[Genotypic characteristics of Mycobacterium avium subsp. hominissuis strains].

Mycobacterium avium are typical environmental, non-tuberculosis microorganisms that occasionally cause mycotuberculosis, an infectious disease in wild and domestic animals, birds, and humans. Here, we report the results of the first study on the genetic diversity of the Russian population of M. avium. A total of 85 M. avium subsp. hominissuis (MAH) clinical strains were isolated from patients (including 30 HIV-positive individuals) with mycobacteriosis in St. Petersburg, 2008-2011. The biochemical identification of the microorganisms was carried out using the PCR detection of the mobile elements IS901 and IS900, as well as of the polymorphism of restriction fragments of the hsp65 gene. The genetic diversity of the isolates was evaluated by VNTR typing based on eight variable-number tandem repeats (VNTRs) (292, X3, 25, 47, 3, 7, 10, and 32 [Thibault et al., 2007]). The MAH population studied was characterized by 15 VNTR types, including nine unique patterns and six clusters of isolates with identical eight-digit profiles. The largest clusters (22221128 and 24221128) included 45 (59.2%) and 15 (19.7%) isolates, respectively; the others contained from 2-7 strains. The strains of the cluster 2533112'8 possessed a truncated TR10 locus (allele 2'). Taking into account the absence of the epidemiological links between the patients and the fact that the infection was delivered from the environment, the high rate of clustering of MAH isolates can be explained by the low discriminatory power of the eight-locus VNTR-typing scheme (HGDI 0-0.61).

[Molecular-genetic characteristics of Mycobacterium tuberculosis strains isolated from patients with tuberculous spondylitis].

AIM: Molecular-genetic characteristic of M. tuberculosis strains isolated from operation material of patients with tuberculous spondylitis. MATERIALS AND METHODS: 107 strains of M. tuberculosis isolated in 2007 - 2011 from patients with spine tuberculosis were studied by methods of spoligotyping and MIRU-VNTR by 12 and 24 loci. RESULTS: Strains of genetic family Beijing dominated (n = 80), 78% of those had multiple drug resistance (MDR). Strains of genetic families T, H3 (Ural), LAM, Manu, H4 and S were also detected. Differentiating of 80 strains of Beijing genotype by MIRU-VNTR method by 24 loci revealed 24 variants (HGI = 0.83) including 7 clusters, the largest of those (100-32) included 23 strains (87% MDR). CONCLUSION: The leading role of Beijing genotype M. tuberculosis strains in development of tuberculous spondylitis with multiple drug resistance of the causative agent is shown.

[Influence of RGPU-207 compound and amiodarone on ion currents in shellfish neurons].

RGPU-207 compound and amiodarone in concentrations of 1, 10, 100 and 1000 microM produce dose-dependent and reversible effects on trans-membrane sodium, calcium, and potassium ion currents of neurons in pond snail and orb snail shellfish. In concentration of 1 microM, both compounds increased the amplitude of potassium ion currents, while not affecting the amplitude of sodium and calcium ion currents. In concentrations of 100 and 1000 microM, dose-dependent suppression of all currents (with predominant potassium ion current suppression) was observed. Under the action of RGPU-207 compound, the kinetics of activation and inactivation of sodium and calcium ion currents was not changed, but the kinetics of activation of the potassium slow current was slowing down. Amiodarone decelerated the inactivation of calcium ion current and accelerated the inactivation of potassium slow current. RGPU-207 compound, in comparison to amiodarone, produces a similar membranotropic effect on the shellfish neurons.

[Effect of GABA derivative--RSMU-151 on the development of oxidative stress in rats with experimental gestosis].

Experimental gestosis induced in rats by drinking 1.8% sodium chloride solution instead of water during the entire period of pregnancy leads to activation of lipid peroxidation (LPO) process, as manifested by increased concentration of diene conjugates and malonic dialdehyde, decreased concentration of antioxidant enzymes (superoxide dismutase and glutathione peroxidase) in homogenates of rat brain, liver, uterus, and placenta. The GABA derivatives--RSMU-151 limits the damaging effect of gestosis, which is manifested by a decrease in the concentration of LPO products and by activation of the antioxidant system enzymes in all organs studied.

[Biodiversity and evolution of circulating bacteria and virus populations. Novel problems of medical microbiology].

Biodiversity and evolution of circulating bacteria and virus populations is a serious scientific problem, solving this problem is necessary for effective prophylaxis of infectious diseases. Principal trends of development in this field of science are described. Results of studies that were carried out and investigated biodiversity of principal pathogens in Russia and St. Petersburg in particular are presented. Risk of infectious security of society caused by increasing diversity of pathogenic microorganisms is described, and priority trends of research development in this field are specified.

[Characteristics of Mycobacterium tuberculosis strains circulating in Pskov region].

AIM: Characteristics of drug resistance (DR) and population structure of Mycobacterium tuberculosis in Pskov region. MATERIALS AND METHODS: In 90 strains of M. tuberculosis drug resistance was studied by culture method and by using "TB-BIOCHIP"; genotyping was determined by spoligotyping method. RESULTS: 55 (61.1%) of 90 M.tuberculosis strains had drug resistance, with 40 (44.4%) being multi-resistant. M. tuberculosis population was presented by SIT1 spoligotype strains of genetic families Beijing--44.4%, LAM--21.1%, T--14.4%, Haarlem--11.1% and Ural--5.6%, according to SpolDB4. Among M. tuberculosis strains circulating in Pskov region the most widespread (44.4%) was SIT1 spoligotype (p < 0.0001). DR and multi-resistant DR (MDR) in Beijing strains occurred more frequently than in "non-Beijing" strains (p < 0.001 and p = 0.03 respectively) and were determined by rpoB mutations Ser531-->Ley and katG Ser315-->Thr. All the SIT252 spoligotype strains were multi-resistant, and their resistance to rifampicin was determined by rpoB Asp516-->Ser substitution, to isoniazid --katG Ser315-->Thr and inhA_T15 substitutions. CONCLUSION: The data obtained gives evidence on tuberculosis epidemiological unfavorability and wide circulation of MDR M. tuberculosis strains in Pskov region.

A Comprehensive Map of Mycobacterium tuberculosis Complex Regions of Difference.

Mycobacterium tuberculosis complex (MTBC) species are classic examples of genetically monomorphic microorganisms due to their low genetic variability. Whole-genome sequencing made it possible to describe both the main species within the complex and M. tuberculosis lineages and sublineages. This differentiation is based on single nucleotide polymorphisms (SNPs) and large sequence polymorphisms in the so-called regions of difference (RDs). Although a number of studies have been performed to elucidate RD localizations, their distribution among MTBC species, and their role in the bacterial life cycle, there are some inconsistencies and ambiguities in the localization of RDs in different members of the complex. To address this issue, we conducted a thorough search for all possible deletions in the WGS data collection comprising 721 samples representing the full MTBC diversity. Discovered deletions were compared with a list of all previously described RDs. As with the SNP-based analysis, we confirmed the specificities of 79 regions at the species, lineage, or sublineage level, 17 of which are described for the first time. We also present RDscan (https://github.com/dbespiatykh/RDscan), an open-source workflow, which detects deletions from short-read sequencing data and correlates the results with high-specificity RDs, curated in this study. Testing of the workflow on a collection comprising ∼7,000 samples showed a high specificity of the found RDs. This study provides novel details that can contribute to a better understanding of the species differentiation within the MTBC and can help to determine how individual clusters evolve within various MTBC species. IMPORTANCE Reductive genome evolution is one of the most important and intriguing adaptation strategies of different living organisms to their environment. Mycobacterium offers several notorious examples of either naturally reduced (Mycobacterium leprae) or laboratory-reduced (Mycobacterium bovis BCG) genomes. Mycobacterium tuberculosis complex has its phylogeny unambiguously framed by large sequence polymorphisms that present unidirectional unique event changes. In the present study, we curated all known regions of difference and analyzed both Mycobacterium tuberculosis and animal-adapted MTBC species. For 79 loci, we have shown a relationship with phylogenetic units, which can serve as a marker for diagnosing or studying biological effects. Moreover, intersections were found for some loci, which may indicate the nonrandomness of these processes and the involvement of these regions in the adaptation of bacteria to external conditions.

[Molecular genetic tools for etiologic diagnostics of disseminated lung tuberculosis].

AIM: Improvement of etiologic diagnostics of disseminated lung tuberculosis (DLT) and determination of Mycobacterium tuberculosis (MBT) drug susceptibility on the basis of molecular genetic methods. MATERIALS AND METHODS: Samples from respiratory tract of patients with DLT were studied using real time polymerase chain reaction and the "TB-BIOCHIP" assay developed by Institute of Molecular Biology. Methods of spoligotyping and reverse hybridization were used for identification, genotyping and express-detection of drug resistance of MBT to rifampicin in sputum samples stained for bacterioscopy. RESULTS: In 76 (41.5%) of 183 patients with radiological signs of DLT, DNA of tuberculosis complex mycobacteria was detected in respiratory tract samples (specificity 87.7%); mutations in genes rpoB, katG, inhA as well as region ahpC-oxyR associated with resistance to rifampicin and isoniazide were revealed in 67% and 79.5% of patients with DLT respectively. In 48.8% of sputum samples, DNA of MBT of epidemically significant genotype Beijing associated with multidrug resistance of MBT in Russia was identified. CONCLUSION: Molecular genetic methods allow to use both fresh and archived respiratory tract specimens for rapid verification of DLT diagnosis during oligobacillar forms of tuberculosis as well as timely prescribe and correct the treatment regimen of the patient according to individual drug susceptibility spectrum of the agent.

Novel macroarray-based method of Corynebacterium diphtheriae genotyping: evaluation in a field study in Belarus.

In this study, we evaluated a novel macroarray-based spoligotyping method for Corynebacterium diphtheriae strain typing. A total of 20 C. diphtheriae biotype gravis toxigenic isolates collected in Belarus from suspected foci of diphtheria infection (diphtheria cases, carriers, or contacts) were subjected to DNA fingerprinting. All strains had an identical ribotyping profile that was identified as ribotype 'Rossija' by comparison with the international ribotype database at the Institut Pasteur of Paris. A spoligotyping method based on simultaneous reverse-hybridization analysis of two CRISPR (clustered, regularly interspaced short palindromic repeats) loci differentiated these strains into three spoligotypes. Comparison of the spoligotyping results with the epidemiological linkage network helped us to resolve suspected links in the chains of transmission. To conclude, the C. diphtheriae spoligotyping method demonstrated its utility in the field study, in particular, underlining the importance of the use of both CRISPR loci. The generated discrete data can be presented in digital binary format and be easily exchanged between laboratories and stored in local and global databases.

Early ancient sublineages of Mycobacterium tuberculosis Beijing genotype: unexpected clues from phylogenomics of the pathogen and human history.

OBJECTIVE: The Mycobacterium tuberculosis Beijing family is an epidemiologically important lineage subdivided into large-scale phylogenetic sublineages: ancient, endemic in East Asia, and global modern. Here, we analysed ancient sublineages of the Beijing genotype in the Omsk region of southwestern Siberia, an intriguing area at the intersection of European Russia, Siberia, and Central Asia. METHODS: The study included 423 M. tuberculosis strains isolated in 2013-2017 and subjected to drug susceptibility testing, genotyping, and whole genome sequencing. RESULTS: The Beijing genotype constituted 280 out of 423 strains. Forty Beijing strains belonged to the early ancient sublineage (wild type mutT4-48). Of these, 11 belonged to the 14717-15 MIRU-VNTR cluster and had intact RD181, 29 belonged to the 1071-32 cluster and had the RD181 deletion. Thirty-nine ancient strains were multidrug-resistant (MDR) and 20 pre-extensively drug resistant (XDR)/XDR. Comparison with global data demonstrated that these clones circulate mainly in Asian Russia with certain phylogenetic affinity to strains from Japan, Korea, and northeastern China. The genome-wide analysis revealed 29-37 single nucleotide polymorphism distances between isolates from different Russian regions within these two clusters. CONCLUSIONS: Based on phylogenetic, phylogeographic, genomic, and historical data, we hypothesize that these two clones or their direct ancestors were probably brought to Russia ∼70 years ago after the Second World War with Japanese prisoners of war and, until recently, were mainly circulating in Siberia and the Far East. Their elevated prevalence in Omsk along with the extremely strong association with not only MDR but also pre-XDR/XDR also observed in other locations highlight their epidemic potential and the need for monitoring and attention from health authorities.

Composition and cardioprotective effects of Primula veris L. solid herbal extract in experimental chronic heart failure.

BACKGROUND: High interest in chronic heart failure (CHF) is accounted for by its high incidence, poor prognosis, growing number of hospital admissions due to the heart failure relapse, and inadequate treatment. These facts necessitate a search for new pharmacological agents for the CHF correction. Herbal medicinal products appear to be very promising as they have a noticeable therapeutic effect and tend to be more harmless in comparison to the most of synthesized medications. PURPOSE: Our aim was to study the composition of the Primula veris L. solid herbal extract (PVSHE) and its effects on the myocardial contractile function in animals with experimental CHF. STUDY DESIGN: The study design involved the identification of the raw material composition of the P. veris L. extract. For the experimental part of our research, we used the model of CHF to elucidate the cardioprotective properties of PVSHE. METHODS: The active extract constituents were isolated by thin-layer chromatography and column chromatography; the extract components were identified by high-performance liquid chromatography, ultraviolet spectroscopy (UVS), and nuclear magnetic resonance spectroscopy (NMRS). To model CHF, L-isoproterenol at a dose of 2.5 mg/kg was intraperitoneally injected to the experimental rats twice a day for 21 days. Cardiac output was assessed with the loading test, adrenoreactivity test, and maximum isometric loading test; CHF markers adrenomedullin and copeptin were detected in blood plasma with ELISA kit for adrenomedullin and copeptin (Coud-Clone Corp., USA). RESULTS: P. veris L. solid herbal extract contains flavonoid aglycons (apigenin, quercetine, kaemferol), flavonoid glycosides (cinarozid, rutin, hyperozid), as well as polymethoxylated flavonoids acting as chemotaxonomic markers for the genus Primula (8-methoxy-flavone; 3',4'methylenedioxy-5'-methoxyflavone). The substance 3',4'methylenedioxy-5'-methoxyflavone has been isolated from the primrose herb for the first time. We showed that the PVSHE has a cardioprotective effect when it was administered at a dose of 30 mg/kg in the experimental CHF, as evidenced by a lower number of animal death, lower level of CHF markers in the blood plasma of the experimental animals, the higher increase in rate of myocardial contraction and relaxation, the higher level of left ventricular pressure (LVP) and of maximum intensity of structural performance (MISP), as compared to the control group. CONCLUSION: P. veris L. solid herbal extract contains flavonoid aglycons, flavonoid glycosides, and polymethoxylated flavonoids. The herbal agent increases the myocardial contractility in experimental CHF.

[Drug sensitivity in Mycobacterium tuberculosis versus its viability, cytotoxicity, genotype, and the course of the process in patients with pulmonary tuberculosis].

The authors studied drug sensitivity, mutations in the katG, in-hA, alpC, rpoB genes, virulence via the cytotoxicity test on THP-1 cells, and the viability and genetic affiliation of 53 clinical M. tuberculosis isolates versus data on the form and dynamics of a process. Sensitive and resistant strains did not significantly differ in viability and cytotoxicity. The highest death of infected macrophages was observed was seen with infection of M. tuberculosis of the Beijing B0 genotype, the least one seen with that of LAM with the similar rate of multiple drug resistance. There was a correlation of the changes in the count of lymphocytes in patients with the genetic affiliation of a causative agent. The severest course of the tuberculous process was observed in baseline lymphopenia (before treatment) in combination with multidrug resistance of mycobacteria, high and moderate cytotoxicity and high viability. Ser-Leu 531 mutation resulted in cross resistance to rifampicin and mycobutin in most cases.

[Differentiation of the Mycobacterium tuberculosis W-Beijing family strains from the Russian Federation by the VNTR-typing].

Recent phylogenetic studies allowed the Mycobacterium tuberculosis complex to be divided into a number of the strain families. The W-Beijing family is one of most widespread M. tuberculosis variants frequently causing epidemic outbreaks. This family is genetically homogenous and conserved so that ETR A, B, C, D, E - typing is insufficient for the W-Beijing differentiation. All W-Beijing isolates have common profile (42435). This led to the false clustering in the molecular epidemiology study, especially in the region of predominance of the W-Beijing family. In this investigation we searched for the VNTR loci with high evolution rate, which were polymorphic in the W-Beijing genome. Eleven VNTR-loci were assayed in the DNA panel of 99 M. tuberculosis isolates from the tuberculosis patients in North-West and West-Siberian regions of Russia during the period from 2000 to 2001. Ninety nine strains of M. tuberculosis were divided into 74 VNTR-types, 51 isolates of the W-Beijing family were subdivided into 30 VNTR-types. The Hunter-Gudson index (HGDI) for all studied loci (ETR-A, ETR-C, ETR-E, V, V2, V3, V4, V5, V6, V10, V11) was close to one of the IS6110 RFLP indices being "the gold standard" of the M. tuberculosis complex genotyping. The V2, V3 loci located in the sequences of the PPE gene family, were highly polymorphic and more discriminative then others (HGDI is about 0.8). The congruence between the IS6110 RFLP-typing and 11 loci VNTR-typing was measured during genotyping for 23 isolates of the W-Beijing family. The isolates were divided into 9 genotypes by the IS6110 RFLP and into 13 variants by the VNTR-typing. The profiles correlation coefficient was 0.767689 that reflected the differences in the rate and type of the given genome target evolution.

Molecular structure of Mycobacterium tuberculosis population in Russia and its interaction with neighboring countries.

BACKGROUND: The tuberculosis (TB) situation in Russia is aggravated by the emergence and the spread of multidrug-resistant strains, HIV co-infection and drawbacks in the health control system. The Mycobacterium tuberculosis population structure in Russia has been defined by the remarkable mass migration in the 20th century, the variable genetic background of different ethnic groups inhabiting Russia, and by the pathobiology of circulating strains. Here, I will review the phylogeography and pathobiology of: (i) the dominating Beijing family; (ii) the Latin-American Mediterranean (LAM) family, the second largest in Russia and MDR-associated in some regions; and (iii) the Ural family, endemic in Russia and thought to be less transmissible and drug resistant. REVIEW AND ANALYSIS: M. tuberculosis variant Beijing В0/W148 is regarded as a successful clone of M. tuberculosis widespread in the former Soviet Union. However, a closer look reveals a peculiar clinal gradient of its geographic distribution; it peaks in Siberia and, to a lesser extent, in the European part of the former USSR. In contrast, its rate is sharply decreased in the Asian part of the former Soviet Union, and it is absent in the autochthonous populations elsewhere in the world. Two interdependent hypotheses will be put forward. First, B0/W148 likely originated in Siberia and its primary dispersal was driven by a massive population outflow from Siberia to European Russia in the period 1960-1980. Second, a historically recent phylogenetically demonstrated successful dissemination of the Beijing B0/W148 strain was triggered by an advent and wide use of the modern anti-TB drugs and was due to its remarkable capacity to acquire drug resistance. Robust phylogenetic markers were used to study the evolution of LAM and its major sublineages in Russia and its neighboring countries. A total of 250 M. tuberculosis isolates were assigned to LAM based on analysis of LAM-specific SNP in Rv3062 and Rv0129c. The family status was rectified for 121 isolates mis-assigned by spoligotyping to non-LAM families (T1 or T5-RUS1). The re-estimated LAM rate increased twofold in Russia and Kazakhstan and fourfold in Belarus. The majority (>90%) of LAM isolates from all three countries belonged to the LAM-RUS sublineage. In contrast, Ibero-American LAM RD-Rio sublineage was identified in only 7 Russian isolates. These findings and further analysis suggest a monophyletic origin of the LAM-RUS subfamily that is endemic in Russia. In contrast, rare LAM RD-Rio isolates were likely brought to Russia through occasional human contact. The analysis of the Ural family showed its highest prevalence in the North/East Pontic (Black Sea) area that may have been an area of its origin and primary dispersal. Ural family strains are not marked by increased pathogenic capacities, association with drug resistance (although there is a trend towards MDR Ural strains in the European part of the former USSR) or increased transmissibility. This reflects their basically low contagiosity which is why the Ural family is still moderately widespread in central Eurasia. Large-scale SNP or WGS population-based studies targeting strains from indigenous populations and, eventually, analysis of ancient DNA will better test these hypotheses. Host genetics factors likely play the most prominent role in the differential dissemination of particular M. tuberculosis genotypes.