Beijing Lineage Research Articles

Hyper transmission of Beijing lineage Mycobacterium tuberculosis: Systematic review and meta-analysis

The globally distributed "Beijing" lineage of Mycobacterium tuberculosis has been associated with outbreaks worldwide. Laboratory based studies have suggested that Beijing lineage may have increased fitness; however, it has not been established whether these differences are of epidemiological significance with regards to transmission. Therefore, we undertook a systematic review of epidemiological studies of tuberculosis clustering to compare the transmission dynamics of Beijing lineages versus the non-Beijing lineages. We systematically searched Embase and MEDLINE before 31st December 2018, for studies which provided information on the transmission dynamics of the different M. tuberculosis lineages. We included articles that conducted population-based cross-sectional or longitudinal molecular epidemiological studies reporting information about extent of transmission of different lineages. The protocol for this systematic review was prospectively registered with PROSPERO (CDR42018088579). Of 2855 records identified by the search, 46 were included in the review, containing 42,700 patients from 27 countries. Beijing lineage was the most prevalent and highly clustered strain in 72.4% of the studies and had a higher likelihood of transmission than non-Beijing lineages (OR 1·81 [95% 1·28-2·57], I2 = 94·0%, τ2 = 0·59, p < 0·01). Despite considerable heterogeneity across epidemiological contexts, Beijing lineage appears to be more transmissible than other lineages.

Genotypes and genetic characters of Mycobacterium tuberculosis from Myanmar using three typing methods

Knowledge on basic characteristics of Mycobacterium tuberculosis (MTB) is helpful to understand the disease epidemiology and support the prediction of clinical outcome of the disease. The aim of this study was to detect the genotypes and genotypic characters of clinical Mycobacterium tuberculosis (MTB) isolates from new and retreatment rifampicin-resistant patients using three different genotyping methods. Mycobacterial interspersed repetitive units-variable number tandem repeat (MIRU-VNTR) typing was used to determine the diversity of 222 clinical isolates. Spoligotyping and IS6110-restriction fragment length polymorphism (RFLP) typing were also used to investigate the genetic characters of 105 MTB strains. Among the 15 genotypes detected by MIRU-VNTR, Beijing strains were the most prevalent of all strains (54.8%); new cases (40.5%) and retreatment cases (69.4%), followed by EAI strain. Spoligotyping categorized the strains into 11 lineages and 13 orphans whereas 96 different IS6110 patterns were identified using RFLP method. The mode number of IS6110 was 18 and 20. Higher band numbers were found in Beijing genotype (p < 0.001). Clustering rates by spoligotyping, MIRU-VNTR and IS6110-RFLP typing were 0.714, 0.004 and 0.085, respectively. Discriminatory powers of spoligotyping, MIRU-VNTR typing and IS6110-RFLP typing were 0.637, 1.000 and 0.997, respectively. Dominant Beijing genotype in both new and retreatment cases denoting that prevailing tuberculosis in Myanmar changed from EAI to Beijing lineage.

Extensive global movement of multidrug-resistant M. tuberculosis strains revealed by whole-genome analysis

BackgroundWhile the international spread of multidrug-resistant (MDR) Mycobacterium tuberculosis strains is an acknowledged public health threat, a broad and more comprehensive examination of the global spread of MDR-tuberculosis (TB) using...

Retrospective Analysis of False-Positive and Disputed Rifampin Resistance Xpert MTB/RIF Assay Results in Clinical Samples from a Referral Hospital in Hunan, China.

Concerns about the specificity of the Xpert MTB/RIF (Xpert) assay have arisen, as false-positive errors in the determination of Mycobacterium tuberculosis complex (MTBC) infection and rifampin (RIF) resistance in clinical practice have been reported. Here, we investigated 33 cases where patients were determined to be RIF susceptible using the Bactec MGIT 960 (MGIT) culture system but RIF resistant using the Xpert assay. Isolates from two of these patients were found not to have any mutations in the rifampin resistance determining region (RRDR) region of rpoB and had good treatment outcomes with first-line antituberculosis (anti-TB) drugs. The remaining 31 patients included 5 new cases and 26 previously treated patients. A large number of well-documented disputed mutations, including Leu511Pro, Asp516Tyr, His526Asn, His526Leu, His526Cys, and Leu533Pro, were detected, and mutations, including a 508 to 509 deletion and His526Gly, were described here as disputed mutations for the first time. Twenty-one (81%) of the 26 previously treated patients had poor treatment outcomes, and isolates from 19 (90%) of these 21 patients were resistant to isoniazid (INH) as determined using the MGIT culture system. Twenty-seven of the 31 isolates with disputed rpoB mutations were phenotypically resistant to INH, 21 (78%) being predicted by GenoType MTBDRplus to have a high level of INH resistance. Most (77.4%) of the isolates with disputed mutations were of the Beijing lineage. These findings have implications for the interpretation of false-positive and disputed rifampin resistance Xpert MTB/RIF results in clinical samples and provide guidance on how clinicians should manage patients carrying isolates with disputed rpoB mutations.

Evidence for Highly Variable, Region-Specific Patterns of T-Cell Epitope Mutations Accumulating in Mycobacterium tuberculosis Strains.

Vaccines that confer protection through induction of adaptive T-cell immunity rely on understanding T-cell epitope (TCE) evolution induced by immune escape. This is poorly understood in tuberculosis (TB), an ancient, chronic disease, where CD4 T-cell immunity is of recognized importance. We probed 905 functionally validated, curated human CD4 T cell epitopes in 79 Mycobacterium tuberculosis (Mtb) whole genomes from India. This screen resulted in identifying 64 mutated epitopes in these strains initially using a computational pipeline and subsequently verified by single nucleotide polymorphism (SNP) analysis. SNP based phylogeny revealed the 79 Mtb strains to cluster to East African Indian (EAI), Central Asian Strain (CAS), and Beijing (BEI) lineages. Eighty-nine percent of the mutated T-cell epitopes (mTCEs) identified in the 79 Mtb strains from India has not previously been reported. These mTCEs were encoded by genes with high nucleotide diversity scores including seven mTCEs encoded by six antigens in the top 10% of rapidly divergent Mtb genes encoded by these strains. Using a T cell functional assay readout, we demonstrate 62% of mTCEs tested to significantly alter CD4 T-cell IFNγ and/or IL2 secretion with associated changes in predicted HLA-DR binding affinity: the gain of function mutations displayed higher predicted HLA-DR binding affinity and conversely mutations resulting in loss of function displayed lower predicted HLA-DR binding affinity. Most mutated antigens belonged to the cell wall/cell processes, and, intermediary metabolism and respiration families though all known Mtb proteins encoded mutations. Analysis of the mTCEs in an SNP database of 5,310 global Mtb strains identified 82% mTCEs to be significantly more prevalent in Mtb strains isolated from India, including 36 mTCEs identified exclusively in strains from India. These epitopes had a significantly higher predicted binding affinity to HLA-DR alleles that were highly prevalent in India compared to HLA-DR alleles rare in India, highlighting HLA-DR maybe an important driver of these mutations. This first evidence of region-specific TCE mutations potentially employed by Mtb to escape host immunity has important implications for TB vaccine design.

High-resolution genotyping of Mycobacterium tuberculosis isolates from syria using mycobacterial interspersed repetitive unit-variable-number tandem repeat

Background: We aimed to evaluate the utility of 24 loci mycobacterial interspersed repetitive unit-variable-number tandem repeat (MIRU-VNTR) genotyping method for discrimination of clinical Mycobacterium tuberculosis isolates in Syria. Methods: We studied 68 clinical tuberculosis (TB) isolates originating from unrelated Syrian TB patients from different regions in Syria. Genetic types (consisting of 24 digits) were determined and used to construct a dendrogram. Results: Fifty-six distinct MIRU patterns were revealed, from which 52 patterns were represented by unique isolates. Sixteen isolates were distributed into 4 clusters, 3 of which consisted of isolates belonging to the TUR lineage. Nine MIRUs showed high Hunter-Gaston index (HGI) values (>0.6), with QUB-26 having the highest discriminatory power (HGI = 0.821), followed by MIRU10, MIRU26, MIRU16, and Mtub39. The cumulative HGI value of the 24-MIRU set was 0.985. Interestingly, using the reduced 15-MIRU, set resulted in the same HGI. The TUR lineage was the most frequent in our sample (23.5%), and it appears that it is widespread in Syria as in Turkey. The discriminatory power of MIRU-VNTR among the subset belonging to the TUR lineage was extremely low due to the high clustering rate (62.5%) of TUR isolates, indicating that this method is inappropriate to discriminate isolates of this lineage. On the other hand, the Beijing lineage was not represented in our isolates. Conclusion: We demonstrated the high prevalence of the TUR lineage and the low prevalence of the Beijing lineage among Syrian clinical TB isolates. The MIRU-VNTR method was highly discriminative among non-TUR TB isolates, but it was inappropriate to discriminate isolates of the TUR lineage.

Socio-Demographic Characteristics and Risk Factors Contributing Pulmonary Tuberculosis Infection and Recent Transmission

Host factors, environmental factors, genetic diversity and distinct phylogeographic distribution of Mycobacterium tuberculosis (MTB) contribute to regional differences in drug resistance. Bangladesh remains among the top 20 high Multi drug resistant tuberculosis (MDR-TB) burden countries of the world. This cross sectional study was conducted to identify the socio demographic characteristics and the risk factors contributing Pulmonary tuberculosis (PTB) infection. These characteristics and risk factors were further investigated among the clustered isolates. Total 60 culture isolates consist of 40 RR and 20 rifampicin sensitive (RS) isolates were enrolled in this study. Laboratory works were done in National Tuberculosis Reference Laboratory (NTRL) and Department of Microbiology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka, Bangladesh. All 60 samples were confirmed as MTB by MPT 64 antigen detection. Two samples were excluded for possible cross contamination and two for failing to give PCR product for most of the locus. So, finally 56 samples were further analyzed for results. Four isolates were distributed within two clusters which were belonged to Beijing lineage. Socio demographic data and risk factors analysis of this study found significant male predominance (p = 0.04) with history of smoking (p = 0.01) and previous anti-TB treatment (p = 0.012) as the significant risk factors for RR TB.

Country-wide rapid screening for the Mycobacterium tuberculosis Beijing sublineage in Ecuador using a single-nucleotide polymorphism-polymerase chain reaction method.

Strains of the Beijing sublineage of Mycobacterium tuberculosis have caused large outbreaks of tuberculosis, often involving multidrug resistance strains and this genetically highly conserved family of strains predominates in some geographic areas. For most of the countries of Latin America, no country-wide studies about the prevalence of the Beijing lineage are available. In this study, we determine the prevalence of the Beijing sublineage in Ecuador, using a large nation-wide sample of 991 isolates from the years 2014-2016 and with the strains, in case-related-proportional representation, emerging from most of the provinces of the country. The isolates were genotyped with asinglenucleotidespecific polymorphism (SNP) polymerase chain reaction for the Beijing sublineage. SNPpositive strains were confirmed as belonging to this lineage with 24 mycobacterial interspersed repetitive unitvariable number of tandem repeat and DNA sequencing. We identified only four Beijing isolates in this collection of 991 strains and calculated a prevalence rate of 0.43%. Our study shows a limited dissemination of the Beijing strains in the Ecuadorian population. This in contrast with the neighbor countries of Peru and Colombia were locally a prevalence of up to 16% has been reported.

Mycobacterium tuberculosis Drug Resistance and Transmission among Human Immunodeficiency Virus-Infected Patients in Ho Chi Minh City, Vietnam.

Vietnam has a high burden of tuberculosis (TB) and multidrug-resistant (MDR) TB, but drug resistance patterns and TB transmission dynamics among TB/human immunodeficiency virus (HIV) coinfected patients are not well described. We characterized 200 Mycobacterium tuberculosis isolates from TB/HIV coinfected patients diagnosed at the main TB referral hospital in Ho Chi Minh City, Vietnam. Phenotypic drug susceptibility testing (DST) for first-line drugs, spoligotyping, and 24-locus mycobacterial interspersed repetitive unit (MIRU-24) analysis was performed on all isolates. The 24-locus mycobacterial interspersed repetitive unit clusters and MDR isolates were subjected to whole genome sequencing (WGS). Most of the TB/HIV coinfected patients were young (162/174; 93.1% aged < 45 years) males (173; 86.5% male). Beijing (98; 49.0%) and Indo-Oceanic (70; 35.0%) lineage strains were most common. Phenotypic drug resistance was detected in 84 (42.0%) isolates, of which 17 (8.5%) were MDR; three additional MDR strains were identified on WGS. Strain clustering was reduced from 84.0% with spoligotyping to 20.0% with MIRU-24 typing and to 13.5% with WGS. Whole genome sequencing identified five additional clusters, or members of clusters, not recognized by MIRU-24. In total, 13 small (two to three member) WGS clusters were identified, with less clustering among drug susceptible (2/27; 7.4%) than among drug-resistant strains (25/27; 92.6%). On phylogenetic analysis, strains from TB/HIV coinfected patients were interspersed among strains from the general community; no major clusters indicating transmission among people living with HIV were detected. Tuberculosis/HIV coinfection in Vietnam was associated with high rates of drug resistance and limited genomic evidence of ongoing M. tuberculosis transmission among HIV-infected patients.

Virulence of Mycobacterium tuberculosis Clinical Isolates Is Associated With Sputum Pre-treatment Bacterial Load, Lineage, Survival in Macrophages, and Cytokine Response.

It is uncertain whether differences in Mycobacterium tuberculosis (Mtb) virulence defined in vitro influence clinical tuberculosis pathogenesis, transmission, and mortality. We primarily used a macrophage lysis model to characterize the virulence of Mtb isolates collected from 153 Vietnamese adults with pulmonary tuberculosis. The virulence phenotypes were then investigated for their relationship with sputum bacterial load, bacterial lineages, bacterial growth, and cytokine responses in macrophages. Over 6 days of infection, 34 isolates (22.2%) showed low virulence (< 5% macrophages lysed), 46 isolates (30.1%) showed high virulence (≥90% lysis of macrophages), and 73 isolates (47.7%) were of intermediate virulence (5–90% macrophages lysed). Highly virulent isolates were associated with an increased bacterial load in patients' sputum before anti-tuberculosis therapy (P = 0.02). Isolate-dependent virulence phenotype was consistent in both THP-1 and human monocyte-derived macrophages. High virulence isolates survived better and replicated in macrophages one hundred fold faster than those with low virulence. Macrophages infected with high virulence isolates produced lower concentrations of TNF-α and IL-6 (P = 0.002 and 0.0005, respectively), but higher concentration of IL-1β (P = 5.1 × 10−5) compared to those infected with low virulence isolates. High virulence was strongly associated with East Asian/Beijing lineage [P = 0.002, Odd ratio (OR) = 4.32, 95% confident intervals (CI) 1.68–11.13]. The association between virulence phenotypes, bacterial growth, and proinflammatory cytokines in macrophages suggest the suppression of certain proinflammatory cytokines (TNF-α and IL-6) but not IL-1β allows better intracellular survival of highly virulent Mtb. This could result in rapid macrophage lysis and higher bacterial load in sputum of patients infected with high virulence isolates, which may contribute to the pathogenesis and success of the Beijing lineage.

Association of gyrA and rrs gene mutations detected by MTBDRsl V1 on Mycobacterium tuberculosis strains of diverse genetic background from India

There is limited data on the use of Genotype MTBDRslVersion 1 (MTBDRsl V1) as an initial rapid screening test to rule out XDR-TB and most importantly its performance in various genotypes of Mycobacterium tuberculosis is scarcely studied. A total of 359 MDR-TB isolates were tested for gene mutations representing second line drug resistance, using the MTBDRsl_V.1 and the results were compared with phenotypic method (Bactec MGIT-960 system) for second-line drug (SLD) susceptibility testing. Genetic lineages of all these isolates were also determined using spoligotyping and SITVIT2 WEB database. The MTBDRsl V1 detected mutations in the gyrA, rrs, and emb genes in 108 (30%), 2 (0.5%) and 129 (35.9%) isolates, respectively. Remaining 120 (33.4%) had no second line drug (SLD) resistance. In 17 (4.7%) isolates mutations were detected in both gyrA and rrs genes. Its concordance with MGIT-960 culture drug susceptibility testing (DST) was 97% and 94.1%, 93.5%, 60.5% and 50% for the detection of XDR-TB, pre-XDR, Ethambutol, and Aminoglycosides/Cyclopeptides resistance. The Beijing lineage was predominant (46%) between both the pre-XDR/XDR-TB isolates. We conclude that MTBDRsl is useful for rapid detection of SLD resistance. Also in pre-XDR and XDR-TB isolates the frequency of relevant genetic mutations was significantly higher in the Beijing strains.

A complete high-quality MinION nanopore assembly of an extensively drug-resistant Mycobacterium tuberculosis Beijing lineage strain identifies novel variation in repetitive PE/PPE gene regions.

A better understanding of the genomic changes that facilitate the emergence and spread of drug-resistant Mycobacterium tuberculosis strains is currently required. Here, we report the use of the MinION nanopore sequencer (Oxford Nanopore Technologies) to sequence and assemble an extensively drug-resistant (XDR) isolate, which is part of a modern Beijing sub-lineage strain, prevalent in Western Province, Papua New Guinea. Using 238-fold coverage obtained from a single flow-cell, de novo assembly of nanopore reads resulted into one contiguous assembly with 99.92 % assembly accuracy. Incorporation of complementary short read sequences (Illumina) as part of consensus error correction resulted in a 4 404 064 bp genome with 99.98 % assembly accuracy. This assembly had an average nucleotide identity of 99.7 % relative to the reference genome, H37Rv. We assembled nearly all GC-rich repetitive PE/PPE family genes (166/168) and identified variants within these genes. With an estimated genotypic error rate of 5.3 % from MinION data, we demonstrated identification of variants to include the conventional drug resistance mutations, and those that contribute to the resistance phenotype (efflux pumps/transporter) and virulence. Reference-based alignment of the assembly allowed detection of deletions and insertions. MinION sequencing provided a fully annotated assembly of a transmissible XDR strain from an endemic setting and showed its utility to provide further understanding of genomic processes within Mycobacterium tuberculosis.

Genetic Diversity and Drug Susceptibility of Mycobacterium tuberculosis Isolates in a Remote Mountain Area of China

ObjectiveWe determined the genetic diversity of Mycobacterium tuberculosis (MTB) in a remote mountainous area of southwest China and evaluated the resolving ability of single nucleotide polymorphism (SNP) genotyping combined with variable number tandem repeat (VNTR) genotyping for Beijing family strains in association with drug resistance status. MethodsThree hundred thirty-one MTB strains were isolated from patients living in mountainous regions of southwest China, and 8-loci SNP, VNTR-15 genotyping assays, and drug susceptibility testing of 9 drugs were performed. ResultsA total of 183 [55.29% (183/331)] strains were classified into the Beijing family. Of the 183 strains, 111 (60.66%) were defined as modern Beijing strains. The most predominant modern Beijing sub-lineage and ancient Beijing sub-lineage were Bmyc10 [39.34% (72/183)] and Bmyc25 [20.77% (38/183)], respectively. Of the isolates, 19.64% (65/331) were resistant to at least 1 of the 9 anti-TB drugs and 17 [4.98% (17/331)] MTB isolates were multi-drug resistant tuberculosis (MDR-TB). Two hundred sixty-one isolates showed a clustering rate of 14.18% (37/261) and a discriminatory index of 0.9990. The Beijing lineage exhibited a significantly higher prevalence of MDR-TB, as well as resistance to isoniazid (INH), rifampin (RIF), and para-aminosalicylic acid (PAS) when analyzed independently (P = 0.005, P = 0.017, P = 0.014, and P = 0.006 respectively). The Beijing lineage was not associated with genetic clustering or resistance to any drug. In addition, genetic clustering was not associated with drug resistance. ConclusionMTB strains demonstrate high genetic diversity in remote mountainous areas of southwest China. Beijing strains, especially modern Beijing strains, are predominant in remote mountainous area of China. The combination of 8-loci SNPs and VNTR-15 genotyping is a useful tool to study the molecular epidemiology of MTB strains in this area.

Single nucleotide polymorphisms of Beijing lineage Mycobacterium tuberculosis toxin-antitoxin system genes: Their role in the changes of protein activity and evolution

The article investigates SNP in genes of toxin-antitoxin systems type II in Mycobacterium tuberculosis Beijing lineage strains and their possible role in the development and formation of new sublineages. We established the catalog of SNPs in 142 TA systems genes in 1349 sequenced genomes of the M. tuberculosis Beijing lineage. Based on the catalog, 15 new sublineages were identified as part of Beijing lineages by non-synonymous SNP in 21 genes of TA systems. We discovered three toxin genes with mutations specific for epidemiologically dangerous sublineages Beijing-modern (vapC37 A46G, vapC38 T143C) and Beijing-B0/W148 (vapC12 A95G). We proved the functional significance of these polymorphisms by cloning these genes wild-type and with marker mutations for the Beijing lineage vapC12 (A95G), vapC37 (A46G), vapC38 (T143C). In vitro study of their activities revealed effect of mutations on the RNase activity of toxin proteins. Mutations in vapC37 and vapC38 decreased toxin activity, and mutation in the vapC12 increased it. We cloned the toxin vapC37 gene of Mycobacterium smegmatis mc2 155 in both allelic variants: without mutation and with A46G mutation, specific for the Beijing-modern lineage. It was shown that this mutation leads to a loss of toxicity.

Frequent transmission of the Mycobacterium tuberculosis Beijing lineage and positive selection for the EsxW Beijing variant in Vietnam.

To examine transmission dynamics of Mtb isolated from TB patients in Ho Chi Minh City, Vietnam we sequenced whole genomes of 1,635 isolates and compared these with 3,144 isolates from elsewhere. The data reveal an underlying burden of disease caused by endemic Mtb Lineage 1 associated with activation of long-term latent infection, and a three-fold higher burden associated with more recently introduced Beijing lineage and Lineage 4 Mtb strains. We find that Beijing lineage Mtb is frequently transferred between Vietnam and other countries, and detect higher levels of transmission of Beijing lineage strains within this host population than endemic Lineage 1 Mtb. Screening for parallel evolution of Beijing lineage-associated SNPs in other Mtb lineages as a signal of positive selection, we identify a mutation in the ESX-5 type VII secreted protein EsxW, which could potentially contribute to the enhanced transmission of Beijing lineage Mtb in Vietnamese and other host populations.

Mutations in ppe38 block PE_PGRS secretion and increase virulence of Mycobacterium tuberculosis.

Mycobacterium tuberculosis requires a large number of secreted and exported proteins for its virulence, immune modulation and nutrient uptake. Most of these proteins are transported by the different type VII secretion systems1,2. The most recently evolved type VII secretion system, ESX-5, secretes dozens of substrates belonging to the PE and PPE families, which are named for conserved proline and glutamic acid residues close to the amino terminus3,4. However, the role of these proteins remains largely elusive 1 . Here, we show that mutations of ppe38 completely block the secretion of two large subsets of ESX-5 substrates, that is, PPE-MPTR and PE_PGRS, together comprising >80 proteins. Importantly, hypervirulent clinical M. tuberculosis strains of the Beijing lineage have such a mutation and a concomitant loss of secretion 5 . Restoration of PPE38-dependent secretion partially reverted the hypervirulence phenotype of a Beijing strain, and deletion of ppe38 in moderately virulent M. tuberculosis increased virulence. This indicates that these ESX-5 substrates have an important role in virulence attenuation. Phylogenetic analysis revealed that deletion of ppe38 occurred at the branching point ofthe 'modern' Beijing sublineage and is shared by Beijing outbreak strains worldwide, suggesting that this deletion may have contributed to their success and global distribution6,7.

Infection of multiple Mycobacterium tuberculosis strains among tuberculosis/human immunodeficiency virus co-infected patients: A molecular study in Myanmar.

Background:Appearance of Mycobacterium tuberculosis (MTB) in the sputum of a tuberculosis (TB)/human immunodeficiency virus (HIV) co-infected patient under treatment may indicate either failure or new infection. This study aims to evaluate whether TB treatment failure among TB/HIV co-infected patients is a real failure.Methods:A prospective cohort study was conducted among 566 TB/HIV co-infected patients who started TB treatment in 12 townships in the upper Myanmar. Among the 566 participants, 16 (2.8%) resulted in treatment failure. We performed a molecular study using mycobacterial interspersed repetitive-unit-variable number of tandem repeat (MIRU-VNTR) genotyping for them. The MIRU-VNTR profiles were analyzed using the web server, MIRU-VNTRplus. All data were entered into EpiData version 3.1 and analyzed using R version 3.4.3.Results:Among 16 failure patients, seven had incomplete laboratory results. Of the nine remaining patients, nobody had exactly the same MIRU-VNTR pattern between the initial and final isolates. Four patients had persistent East-African Indian (EAI) lineages and one each had persistent Beijing lineage, changing from EAI to Beijing, from Beijing to EAI, NEW-1 to Beijing, and NEW-1 to X strains. Female patients have significantly larger genetic difference between MTB of the paired isolates than male patients (t-test, P = 0.04).Conclusion:Thus, in our study patients, infection of multiple MTB strains is a possible cause of TB treatment failure. Explanation for the association between gender and distance of genotypes from the initial to subsequent MTB infection needs further studies.

Assessing the Combined Antibacterial Effect of Isoniazid and Rifampin on Four Mycobacterium tuberculosis Strains Using In Vitro Experiments and Response-Surface Modeling.

While isoniazid and rifampin have been the cornerstone of tuberculosis therapy caused by drug-susceptible Mycobacterium tuberculosis for more than 40 years, their combined action has never been thoroughly assessed by modern quantitative pharmacology approaches. The aims of this work were to perform in vitro experiments and mathematical modeling of the antibacterial effect of isoniazid and rifampin alone and in combination against various strains of Mycobacterium tuberculosis After MIC determination of H37Rv and three strains belonging to the Beijing, Euro-American, and Indo-Oceanic lineages, the antibacterial effects of isoniazid and rifampin alone and in combination were studied in static time-kill experiments. A sigmoidal maximum effect model (Hill equation) and a response-surface model were used to describe the effect of the drugs alone and in combination, respectively. The killing effect of isoniazid and rifampin alone were well described by the Hill equation. Rifampin displayed a more concentration-dependent effect than isoniazid around the MIC. The pharmacodynamics parameters of each drug (maximal effect, median effect concentration, and coefficient of sigmoidicity) were quite similar between the four strains. The response-surface model from Minto et al. fit data of combined effect very well with low bias and imprecision (C. F. Minto, T. W. Schnider, T. G. Short, K. M. Gregg, A. Gentilini, Anesthesiology 92:1603-1616, 2000, https://doi.org/10.1097/00000542-200006000-00017). Response-surface modeling showed that the combined action of isoniazid and rifampin was synergistic for the H37Rv, Beijing, and Euro-American strains but only additive for the Indo-Oceanic strain. This study can serve as a motivating example for preclinical evaluation of combined action of antituberculous drugs.

ФИЛОГЕОГРАФИЯ MYCOBACTERIUM TUBERCULOSIS В ЦЕНТРАЛЬНОЙ И ПО-ГРАНИЧНОЙ МОНГОЛИИ

The distribution of Mycobacterium tuberculosis and scope of tuberculosis morbidity in the world are extremely considerable. Among regions with high burden of tuberculosis (according to the World Health Organization data for 2015) Mongolia is one of the most disadvantaged countries. This research was continuation of studying of phylogenetic structure of M. tuberculosis for the determination of evolutionary and migration processes in pathogen's northeastern population and was devoted to the study of the spatial distribution of epidemically significant genotypes and subtypes, circulating in central and border (northern and southern) Mongolia. The DNAs of 309 clinical isolates of the pathogen were genotyped by the 24-locus MIRU-VNTR and LSP methods. Strains of the Beijing lineage were additionally analyzed for the presence of specific SNPs and its 24-MIRU-VNTR profiles were compared with reference database of M. Merker et al. (2015). The study indicated significant predominance of strains of the Beijing lineage (subtype CC4). This genotype is characterized by high frequency of clustering, which indicates its epidemic danger and the relatively recent introduction of «successful» strains into the territory of Mongolia. Also, a relatively homogeneous distribution structure of genotypes and subtypes of the pathogen was revealed within the studied region, which differs from the structure of the Russian population of M. tuberculosis by its subtypical composition. Thus, we assuming that CC4-strains of the Beijing lineage spread into Mongolia from neighboring East Asian regions, most likely from the Northern provinces of China (Inner Mongolia) or nearest neighboring countries of Southeast Asia.

The intrinsic antibiotic resistance to β-lactams, macrolides, and fluoroquinolones of mycobacteria is mediated by the whiB7 and tap genes

The M. tuberculosis resistome includes a number of genes involved in intrinsic drug resistance. One of the major members of this system is the whiB7 gene encoding a transcription factor, which regulates expression of a number of genes—its regulon. In this study, we analyze the impact on intrinsic drug resistance levels of the whiB7 and tap genes and their prevalent mutants. The mutations found within whiB7 and tap genes in the genomes of strains belonging to Beijing and EAI-Manila lineages allowed us to investigate the new role of these genes in the intrinsic drug resistance of mycobacteria to β-lactams, fluoroquinolones, and macrolides and to assume their significance in the development of M. tuberculosis genotypes. Here we also established that the mutation in the tap gene–insC581, previously described, as a marker of the Beijing lineage, cannot be the marker due to its absence in a number of Beijing strains.