Mycobacterium Tuberculosis Beijing Genotype Research Articles

Effects of the Beijing genotype on latent tuberculosis infection, TB disease risk, and clustering of TB cases

BackgroundThe Beijing genotype of Mycobacterium tuberculosis (Mtb) has sparked debate regarding its virulence and transmissibility. This study contributes to this discussion by assessing its effect on the risk of latent tuberculosis infection (LTBI), active tuberculosis (TB) disease among contacts, and clustering of known TB cases. MethodsWe conducted a retrospective cohort study using the records of 4457 culture-confirmed TB patients and their contacts (20,448) reported to the Florida Department of Health between 2009 and 2023. Univariate and multivariate analyses were used to evaluate the effect of the Beijing strain on LTBI, active TB risk among contacts, and case clustering. ResultsOur study revealed no significant difference in transmissibility between the Beijing and non-Beijing genotypes among contacts. LTBI prevalence was 19.9%, slightly higher in non-Beijing than Beijing genotypes (20.2% vs. 15.5%, p < 0.001). The prevalence of active TB was 1.8%, with no significant difference between the Beijing and non-Beijing genotypes (1.4% vs. 1.8%, p = 0.296). Increased LTBI risk was associated with older age, male sex, Hispanic ethnicity, multidrug-resistant TB exposure, household exposure, and a longer exposure duration. Active TB risk was higher for males, HIV-positive individuals, and contacts with more prolonged exposure to index cases. The Beijing genotype was associated with increased TB case clustering (aOR = 1.98, 95%CI: 1.53, 2.55, p < 0.001) as compared to the non-Beijing genotypes. US birthplace (aOR = 2.75, 95%CI: 2.37, 3.19, p < 0.001), pulmonary disease (aOR = 1.27, 95%CI: 1.04, 1.56, p < 0.020), cavitary TB (aOR = 1.25, 95% CI: 1.08, 1.44, p < 0.003), previous year alcohol use (aOR = 1.68, 95%CI: 1.38, 2.04, p < 0.001), and recreational drug use (aOR = 1.32, 95%CI: 1.04, 1.67, p < 0.024) were also associated with an increased risk of TB case clustering. ConclusionWhile the Beijing genotype did not increase the risk of LTBI or active TB among contacts, it showed a higher tendency for case clustering. Hence, interventions should prioritize populations where this genotype is prevalent.

Insight into pathogenomics and phylogeography of hypervirulent and highly-lethal Mycobacterium tuberculosis strain cluster

Background. The Mycobacterium tuberculosis Beijing genotype is globally spread lineage with important medical properties that however vary among its subtypes. M. tuberculosis Beijing 14717-15-cluster was recently discovered as both multidrug-resistant, hypervirulent, and highly-lethal strain circulating in the Far Eastern region of Russia. Here, we aimed to analyze its pathogenomic features and phylogeographic pattern.Results. The study collection included M. tuberculosis DNA collected between 1996 and 2020 in different world regions. The bacterial DNA was subjected to genotyping and whole genome sequencing followed by bioinformatics and phylogenetic analysis. The PCR-based assay to detect specific SNPs of the Beijing 14717-15-cluster was developed and used for its screening in the global collections. Phylogenomic and phylogeographic analysis confirmed endemic prevalence of the Beijing 14717-15-cluster in the Asian part of Russia, and distant common ancestor with isolates from Korea (> 115 SNPs). The Beijing 14717-15-cluster isolates had two common resistance mutations RpsL Lys88Arg and KatG Ser315Thr and belonged to spoligotype SIT269. The Russian isolates of this cluster were from the Asian Russia while 4 isolates were from the Netherlands and Spain. The cluster-specific SNPs that significantly affect the protein function were identified in silico in genes within different categories (lipid metabolism, regulatory proteins, intermediary metabolism and respiration, PE/PPE, cell wall and cell processes).Conclusions. We developed a simple method based on real-time PCR to detect clinically significant MDR and hypervirulent Beijing 14717-15-cluster. Most of the identified cluster-specific mutations were previously unreported and could potentially be associated with increased pathogenic properties of this hypervirulent M. tuberculosis strain. Further experimental study to assess the pathobiological role of these mutations is warranted.

Association of Single-Nucleotide Polymorphisms in the VDR Gene with Tuberculosis and Infection of Beijing Genotype Mycobacterium tuberculosis.

The aim of the present study was to investigate the association between vitamin D receptor (VDR) gene polymorphism and tuberculosis susceptibility, as well as the potential interaction of host genetic factors with the heterogeneity of Mycobacterium tuberculosis in the population from Xinjiang, China. From January 2019 to January 2020, we enrolled 221 tuberculosis patients as the case group and 363 staff with no clinical symptoms as the control group from four designated tuberculosis hospitals in southern Xinjiang, China. The polymorphisms of Fok I, Taq I, Apa I, Bsm I, rs3847987 and rs739837 in the VDR were detected by sequencing. M. tuberculosis isolates were collected from the case group and identified as Beijing or non-Beijing lineage by multiplex PCR. Propensity score (PS), univariate analysis and multivariable logistic regression models were used to perform the analysis. Our results showed that the allele and genotype frequencies of Fok I, Taq I, Apa I, Bsm I, rs3847987 and rs739837 in VDR were not correlated with tuberculosis susceptibility or lineages of M. tuberculosis. Two out of six loci of the VDR gene formed one haplotype block, and none of the haplotypes was found to correlate with tuberculosis susceptibility or lineages of M. tuberculosis infected. Polymorphisms in the VDR gene may not indicate susceptibility to tuberculosis. There was also no evidence on the interaction between the VDR gene of host and the lineages of M. tuberculosis in the population from Xinjiang, China. Further studies are nonetheless required to prove our conclusions.

The mutation rate of rpoB gene showed an upward trend with the increase of MIRU10, MIRU39 and QUB4156 repetitive number

BackgroundMycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) is a frequently used typing method for identifying the Beijing genotype of Mycobacterium tuberculosis (Mtb), which is easily transformed into rifampicin (RIF) resistance. The RIF resistance of Mtb is considered to be highly related with the mutation of rpoB gene. Therefore, this study aimed to analyze the relationship between the repetitive number of MIRU loci and the mutation of rpoB gene.MethodsAn open-source whole-genome sequencing data of Mtb was used to detect the mutation of rpoB gene and the repetitive number of MIRU loci by bioinformatics methods. Cochran-Armitage analysis was performed to analyze the trend of the rpoB gene mutation rate and the repetitive number of MIRU loci.ResultsAmong 357 rifampicin-resistant tuberculosis (RR-TB), 304 strains with mutated rpoB genes were detected, and 6 of 67 rifampicin susceptible strains were detected mutations. The rpoB gene mutational rate showed an upward trend with the increase of MIRU10, MIRU39, QUB4156 and MIRU16 repetitive number, but only the repetitive number of MIRU10, MRIU39 and QUB4156 were risk factors for rpoB gene mutation. The Hunter-Gaston discriminatory index (HGDI) of MIRU10 (0.65) and QUB4156 (0.62) was high in the overall sample, while MIRU39 (0.39) and MIRU16 (0.43) showed a moderate discriminatory Power.ConclusionThe mutation rate of rpoB gene increases with the addition of repetitive numbers of MIRU10, QUB4156 and MIRU39 loci.

Recent Transmission and Prevalent Characterization of the Beijing Family Mycobacterium tuberculosis in Jiangxi, China.

The Beijing genotype is the most common type of tuberculosis in Jiangxi Province, China. The association of population characteristics and their prevalence in the development of recent transmission is still unclear. 1,433 isolates were subjected to drug-resistant tests and MIRU-VNTR analysis. We compared differences in demographic characteristics and drug resistance patterns between the Beijing and non-Beijing family strains. We also explored the association of the clustering rate with the Beijing genotype of Mycobacterium tuberculosis. The Beijing genotype was dominant (78.16%). The results of MIRU-VNTR showed that 775 of 1,433 strains have unique patterns, and the remaining gather into 103 clusters. A recent transmission rate was 31.54% (452/1,433). The Beijing genotype strains were more likely to spread among the recurrent population (p = 0.004), people less than 50 years of age (p = 0.02 or 0.003), and the personnel in the northern regions (p = 0.03). Drug resistance patterns did not show significant differences between Beijing and non-Beijing genotype isolates. Furthermore, we found that HIV-positive cases had a lower clustering rate (p = 0.001). Our results indicated that the recurrent population and people under 50 years of age were more likely to be infected with the Beijing genotype of M. tuberculosis. The strains from the Beijing family were easier to cluster compared to strains isolated from the non-Beijing family. Social activity and AIDS substantially impacted the clustering rate of the Beijing genotype of M. tuberculosis. Multidrug resistant M. tuberculosis affected Beijing genotype transmission.

Molecular epidemiological analysis of Mycobacterium tuberculosis modern Beijing genotype strains isolated in Chiba Prefecture over 10 years

IntroductionThe prevalence of the phylogenetic groups of Mycobacterium tuberculosis Beijing genotype has been reported to be similar in different areas of Japan. However, recent reports from rural areas of Japan show a low prevalence of modern Beijing strains, suggesting that the distribution of modern Beijing strains may have changed recently. Therefore, multi-locus variable number of tandem repeats analysis (MLVA) and draft whole genome sequence (DWGS) analysis were carried out to investigate the prevalence of particular genotype strains. MethodsNine hundred and ninety modern Beijing strains were studied using minimum spanning tree (MST) analysis and neighbor-net analysis of MLVA and WGS data. ResultsAn MST of M. tuberculosis Beijing genotype strains reconstructed from 12 loci-MLVA data showed two large complexes with the J12-0006 MLVA pattern. In one of the complexes, strains with the pECT07 pattern produced by 24 loci-MLVA and its SLVs were most prevalent. DWGS analysis was carried out for pECT07 and its SLV strains. Neighbor-net and MST analyses of the DWGS data showed that pECT07 and its SLV strains were grouped in separate clusters. When all the combinations of two of the tested strains were analyzed, MST analysis showed that only 9 (1.7%) of the 528 pairs of tested strains had 5 or less SNPs. ConclusionsThe results of this study suggested that pECT07 and its variants were prevalent among M. tuberculosis modern Beijing strains in Chiba Prefecture, but the prevalence of those strains may not have been due to an earlier large-scale latent outbreak.

Practical approach to detection and surveillance of emerging highly resistant Mycobacterium tuberculosis Beijing 1071-32-cluster

Ancient sublineage of the Mycobacterium tuberculosis Beijing genotype is endemic and prevalent in East Asia and rare in other world regions. While these strains are mainly drug susceptible, we recently identified a novel clonal group Beijing 1071-32 within this sublineage emerging in Siberia, Russia and present in other Russian regions. This cluster included only multi/extensive drug resistant (MDR/XDR) isolates. Based on the phylogenetic analysis of the available WGS data, we identified three synonymous SNPs in the genes Rv0144, Rv0373c, and Rv0334 that were specific for the Beijing 1071-32-cluster and developed a real-time PCR assay for their detection. Analysis of the 2375 genetically diverse M. tuberculosis isolates collected between 1996 and 2020 in different locations (European and Asian parts of Russia, former Soviet Union countries, Albania, Greece, China, Vietnam, Japan and Brazil), confirmed 100% specificity and sensitivity of this real-time PCR assay. Moreover, the epidemiological importance of this strain and the newly developed screening assay is further stressed by the fact that all identified Beijing 1071-32 isolates were found to exhibit MDR genotypic profiles with concomitant resistance to additional first-line drugs due to a characteristic signature of six mutations in rpoB450, rpoC485, katG315, katG335, rpsL43 and embB497. In conclusion, this study provides a set of three concordant SNPs for the detection and screening of Beijing 1071-32 isolates along with a validated real-time PCR assay easily deployable across multiple settings for the epidemiological tracking of this significant MDR cluster.

MDR and Pre-XDR Clinical Mycobacterium tuberculosis Beijing Strains: Assessment of Virulence and Host Cytokine Response in Mice Infectious Model.

Mycobacterium tuberculosis Beijing genotype associated with drug resistance is a growing public health problem worldwide. The aim of this study was the assessment of virulence for C57BL/6 mice after infection by clinical M. tuberculosis strains 267/47 and 120/26, which belong to the modern sublineages B0/W148 and Central Asia outbreak of the Beijing genotype, respectively. The sublineages were identified by the analysis of the strains’ whole-genomes. The strains 267/47 and 120/26 were characterized as agents of pre-extensively drug-resistant (pre-XDR) and multidrug-resistant (MDR) tuberculosis, respectively. Both clinical strains were slow-growing in 7H9 broth compared to the M. tuberculosis H37Rv strain. The survival rates of C57BL/6 mice infected by 267/47, 120/26, and H37Rv on the 150th day postinfection were 10%, 40%, and 70%, respectively. Mycobacterial load in the lungs, spleen, and liver was higher and histopathological changes were more expressed for mice infected by the 267/47 strain compared to those infected by the 120/26 and H37Rv strains. The cytokine response in the lungs of C57BL/6 mice after infection with the 267/47, 120/26, and H37Rv strains was different. Notably, proinflammatory cytokine genes Il-1α, Il-6, Il-7, and Il-17, as well as anti-inflammatory genes Il-6 and Il-13, were downregulated after an infection caused by the 267/47 strain compared to those after infection with the H37Rv strain.

Probable long-term prevalence for a predominant Mycobacterium tuberculosis clone of a Beijing genotype in Colon, Panama.

Beijing genotype Mycobacterium tuberculosis strains associate with increased virulence, resistance and/or higher transmission rates. This study describes a specific Beijing strain predominantly identified in the Panamanian province of Colon with one of the highest incidences of tuberculosis in the country. Retrospective mycobacterial interspersed repetitive unit/variable number of tandem repeats analysis of 42 isolates collected between January and August 2018 allowed to identify a cluster (Beijing A) with 17 (40.5%) Beijing isolates. Subsequent prospective strain-specific PCR-based surveillance from September 2019 to March 2020 confirmed the predominance of the Beijing A strain (44.1%) in this province. Whole-genome sequencing revealed higher-than-expected diversity within the cluster, suggesting long-term prevalence of this strain and low number of cases caused by recent transmission. The Beijing A strain belongs to the Asian African 3 (Bmyc13, L2.2.5) branch of the modern Beijing sublineage, with their closest isolates corresponding to cases from Vietnam, probably introduced in Panama between 2000 and 2012.

Comparison of two molecular diagnostic methods for identifying Beijing genotype of Mycobacterium tuberculosis

The Beijing family of Mycobacterium tuberculosis has been identified as a severe pathogen among this species and found in many clinical isolates during the last decade. Early identification of such genotype is important for better prevention and treatment of tuberculosis. The present study performed to compare the efficiency of Real-Time PCR and IS6110-Based Inverse PCR methods to identify the Beijing family. This study was carried out on 173 clinical isolates of Mycobacterium tuberculosis complex in Golestan Province, northern Iran. DNA extraction performed by boiling and determining the Beijing and non-Beijing strains carried out using Real-Time PCR and IS6110-Based Inverse PCR. In both Real-Time PCR and IS6110-Based Inverse PCR method, 24 specimens (13.9%) of the Beijing family were identified and the result of the IS6110-Based Inverse PCR method showed that all the Beijing strains in this region belonged to the Ancient Beijing sub-lineage. Although the efficacy of the two methods in the diagnosis of the Beijing family is similar, the IS6110-Based Inverse PCR is more applicable to the ability to detect new and old Beijing family.

Genomic signatures of drug resistance in highly resistant Mycobacterium tuberculosis strains of the early ancient sublineage of Beijing genotype in Russia

The Mycobacterium tuberculosis Beijing genotype is a clinically and epidemiologically important lineage that is subdivided into ancient/ancestral and modern strains. In our previous study in western Siberia, we identified variable number of tandem repeats (VNTR)-based clusters within the early ancient sublineage of the Beijing genotype characterized by an unexpectedly high rate of extensive drug resistance (XDR). In the current study, next generation sequencing data were analysed to gain insight into genomic signatures underlying drug resistance of these strains. A total of 184 genomes of the Beijing early ancient sublineage from Russia (16), China (15), Japan (36), Korea (25), Vietnam (18), Thailand (73), and the USA (1) were used for phylogenetic analysis. The drug-resistant profile was deduced genotypically. The Russian isolates were distributed into two clusters and were all drug resistant, mainly pre-XDR and XDR. The largest of these clusters included only Russian isolates from remote locations in both Asian and European parts of the country. All its isolates had a quadruple drug resistance (to isoniazid, rifampin, ethambutol and streptomycin) due to the 6-mutation signature (KatG Ser315Thr, KatG Ile335Val, RpoB Ser450Leu, RpoC Asp485Asn, EmbB Gln497Arg, and RpsL Lys43Arg). In most samples, it was complemented with additional and different pncA, gyrA and rrs mutations leading to the pre-XDR/XDR genotype. Phylogenomic analysis indicates a distant origin of this Russian resistant cluster in the early 1970s but location and circumstances are yet to be clarified.

Genotyping, Assessment of Virulence and Antibacterial Resistance of the Rostov Strain of Mycobacterium tuberculosis Attributed to the Central Asia Outbreak Clade.

The Central Asia Outbreak (CAO) clade is a growing public health problem for Central Asian countries. Members of the clade belong to the narrow branch of the Mycobacterium tuberculosis Beijing genotype and are characterized by multidrug resistance and increased transmissibility. The Rostov strain of M. tuberculosis isolated in Russia and attributed to the CAO clade based on PCR-assay and whole genome sequencing and the laboratory strain H37Rv were selected to evaluate the virulence on C57Bl/6 mice models by intravenous injection. All mice infected with the Rostov strain succumbed to death within a 48-day period, while more than half of the mice infected by the H37Rv strain survived within a 90-day period. Mice weight analysis revealed irreversible and severe depletion of animals infected with the Rostov strain compared to H37Rv. The histological investigation of lung and liver tissues of mice on the 30th day after injection of mycobacterial bacilli showed that the pattern of pathological changes generated by two strains were different. Moreover, bacterial load in the liver and lungs was higher for the Rostov strain infection. In conclusion, our data demonstrate that the drug-resistant Rostov strain exhibits a highly virulent phenotype which can be partly explained by the CAO-specific mutations.

Study on the prevalence of Beijing genotype Mycobacterium tuberculosis and its relationship with second-line anti-tuberculosis drug resistance

Objective: To explore the prevalence risk factors of Beijing genotype Mycobacterium tuberculosis (MTB) in Beijing and its correlation with second-line anti tuberculosis drug resistance. Methods: A total of 1 140 clinical MTB positive strains were collected from various districts in Beijing, and the drug sensitivity was detected by proportion method. Beijing genotype and non Beijing genotype MTB were identified by the method of Spoligotyping. Using SPSS 22.0 statistical software, chi square test or Fisher exact probability test was used to analyze the experimental data. Results: Among 1 140 MTB clinical isolates, 941 (82.5%) were Beijing genotype MTB, 199 were non Beijing genotype MTB. There were 663 males (70.5%) in Beijing genotype and 124 males (62.3%) in non Beijing genotype strains. There were significant differences in the proportion of males between the two genotypes [P=0.021, OR (95% CI):1.442 (1.048-1.985)]. There were 441 floating population (46.9%) in Beijing genotype MTB and 78 floating population (39.2%) in non Beijing genotype MTB. There was a significant difference in the proportion of floating population between the two genotypes [P=0.048,OR (95%CI):1.368(1.001-1.869)]. There were 129 patients (13.7%) aged 65 or older in Beijing genotype MTB, 40 patients (20.1%) aged 65 or older in non Beijing genotype MTB. The difference was statistically significant [P=0.021, or (95% CI): 0.631 (0.426-0.936)]. The resistance drug rates of Levofloxacin (Lfx), Amikacin (Am), Capreomycin (Cm), Para-aminosalicylic (PAS) in Beijing genotypes were 5.5% (52/941), 1.3% (12/941), 3.2% (30/941) and 3.0% (28/941), respectively, and those of non Beijing genotypes were 10.6% (21/199), 8.5% (17/199, 12.6% (25/199) and 11.6% (23/199), the difference was statistically significant (all P<0.05). There were 58 (6.2%) multidrug-resistant (MDR) strains in Beijing genotype MTB and 19 (9.5%) multidrug-resistant strains in non Beijing genotype. There was no significant difference in the proportion of MDR strains between Beijing genotype and non Beijing genotype (P>0.05). Conclusions: Beijing genotype MTB is widespread in Beijing and has a higher proportion in male population and floating population. Compared with non Beijing genotype, Beijing genotype MTB has a lower resistance rate to Lfx, Am, Cm and PAS, and there is no significant difference in the proportion of MDR-TB patients between the two genotypes.

Outbreak of Mycobacterium tuberculosis Beijing Strain in a High School in Yunnan, China.

In this study, we report an investigation of a tuberculosis (TB) outbreak in a high school in China. Eleven students with active TB were identified. A culture-negative 17-year-old girl was considered as index case affected by pulmonary and meningeal TB. Screening results indicated latent TB in 32.8% of the students in the classroom of index case, whereas a significantly decreased prevalence of TB infection was found among students on the same floor, ranging from 1.3 to 8.2%. Genotyping revealed that all the Mycobacterium tuberculosis (MTB) isolates belonged to Beijing spoligotype international types 1 (SIT1). In conclusion, a diagnostic delay for the culture-negative index case played an important role in the transmission of Beijing genotype MTB strain in the boarding school in Yunnan. The separate locations of classrooms and sufficient air ventilation contributed to the significant difference in proportions of TB infection between classmates and other students in this outbreak.

A landscape of genomic alterations at the root of a near-untreatable tuberculosis epidemic

BackgroundAtypical Beijing genotype Mycobacterium tuberculosis strains are widespread in South Africa and have acquired resistance to up to 13 drugs on multiple occasions. It is puzzling that these strains have retained fitness and transmissibility despite the potential fitness cost associated with drug resistance mutations.MethodsWe conducted Illumina sequencing of 211 Beijing genotype M. tuberculosis isolates to facilitate the detection of genomic features that may promote acquisition of drug resistance and restore fitness in highly resistant atypical Beijing forms. Phylogenetic and comparative genomic analysis was done to determine changes that are unique to the resistant strains that also transmit well. Minimum inhibitory concentration (MIC) determination for streptomycin and bedaquiline was done for a limited number of isolates to demonstrate a difference in MIC between isolates with and without certain variants.ResultsPhylogenetic analysis confirmed that two clades of atypical Beijing strains have independently developed resistance to virtually all the potent drugs included in standard (pre-bedaquiline) drug-resistant TB treatment regimens. We show that undetected drug resistance in a progenitor strain was likely instrumental in this resistance acquisition. In this cohort, ethionamide (ethA A381P) resistance would be missed in first-line drug-susceptible isolates, and streptomycin (gidB L79S) resistance may be missed due to an MIC close to the critical concentration. Subsequent inadequate treatment historically led to amplification of resistance and facilitated spread of the strains. Bedaquiline resistance was found in a small number of isolates, despite lack of exposure to the drug. The highly resistant clades also carry inhA promoter mutations, which arose after ethA and katG mutations. In these isolates, inhA promoter mutations do not alter drug resistance, suggesting a possible alternative role.ConclusionThe presence of the ethA mutation in otherwise susceptible isolates from ethionamide-naïve patients demonstrates that known exposure is not an adequate indicator of drug susceptibility. Similarly, it is demonstrated that bedaquiline resistance can occur without exposure to the drug. Inappropriate treatment regimens, due to missed resistance, leads to amplification of resistance, and transmission. We put these results into the context of current WHO treatment regimens, underscoring the risks of treatment without knowledge of the full drug resistance profile.

Establishment of a multiplex PCR for rapid identification of Mycobacterium species

Objective To establish and evaluate a multiplex PCR method for rapid identification of Mycobacterium species in order to provide an approach for rapid diagnosis of pathogens causing tuberculosis. Methods Four genes including 16S rRNA, Rv0577, RD9 and mtbk_20680 were selected to establish the multiplex PCR method. Specific primers were designed and the reaction system and conditions were optimized. The sensitivity and specificity of the multiplex PCR method were evaluated through identifying Mycobacterium africanum (M.africanum), Mycobacterium bovis (M.bovis), M. bovis BCG, seven common non-tuberculosis Mycobacterium (NTM), seven reference species of common respiratory bacteria and 93 clinical strains of Mycobacterium tuberculosis (MTB) isolated from patients with tuberculosis in Gansu Province of China. Results The fragments of 16S rRNA, Rv0577, RD9 and mtbk_20680 genes were 543 bp, 786 bp, 369 bp and 231 bp in length, respectively. MTB strains of the Beijing genotype were positive for all of the four genes, while the non-Beijing genotype strains were negative for mtbk_20680. M. africanum, M. bovis and M. bovis BCG strains were negative for 16S rRNA and Rv0577. NTM strains only carried 16S rRNA and none of four genes were detected in the seven species of respiratory bacteria. Among the 93 clinical MTB strains, 80 (86.02%) belonged to the Beijing genotype and the other 13 (13.98%) were non-Beijing genotype strains. The specificity of the multiplex PCR method was 100%. Conclusions The established multiplex PCR method could accurately distinguish Mycobacterium, Mycobacterium tuberculosis complex (MTBC), NTM, MTB, and Beijing and non-Beijing genotype MTB with high sensitivity and specificity. Key words: Multiplex PCR; Mycobacterium; Mycobacterium tuberculosis; Beijing genotype

Beijing genotype of Mycobacterium tuberculosis is less associated with drug resistance in south China

Mycobacterium tuberculosis Beijing genotype strains are widespread globally. However, there has been no systematic study on the association between Beijing genotype and the characteristics of drug resistance. In this study, 359 M. tuberculosis isolates from south China were collected and their background information, genotype diversity and drug resistance was investigated. The results revealed that 66.0% of strains (237/359) were categorised as Beijing genotype. There was no statistical difference between Beijing and non-Beijing genotype strains in terms of patient sex, age, place of residence and treatment history. Drug resistance testing showed that 34.8% (125/359) of isolates were resistant to at least one of the seven drugs tested. The proportions of multidrug-resistant tuberculosis and extensively drug-resistant tuberculosis were 17.0% and 1.4%, respectively. Previously treated patients presented a significantly higher risk of developing drug resistance than new cases. Although the prevalence of drug resistance was higher in Beijing genotype than in non-Beijing genotype strains, there was no significant difference between these two genotypes in the multivariate analysis. Even in re-treated patients, the association of Beijing genotype with drug resistance was not significant. This study provides an insight into genotype diversity and demonstrates the characteristics of drug resistance in Beijing genotype strains, which will be useful in generating efficient tuberculosis prevention and control strategies in China.

In vivo virulence of Beijing genotype Mycobacterium tuberculosis

Mycobacterium tuberculosis Beijing genotype strains comprise 5080% in Russian Federation, which are divided into the main B0/W148, CladeA, and CAO clusters based on VNTR and SNP analysis. It should be noted that such phylogenetically highly close MTB strains belong to the modern Beijing family, generally demonstrating high transmissibility, association with drug resistance, and prevalence among patients with severe forms of the disease. However, studies on MTB genetic cluster strain-related virulence are scarce and contradictory. Here, we investigated virulence of diverse Mycobacterium tuberculosis strains belonging to the B0/W148, CladeA and CAO clusters and nonclustered strain NK of the Beijing family as well as laboratory strain H37Rv in C57BL/6 mice. It was found that mice infected with NK and B0/W148 vs. CladeA strains revealed the peak and the lowest mortality, respectively, while assessing survival rate in various groups (20 mice per MTB strain examined). Analyzing experimental data in mice demonstrated that all MTB strains were able to cause typical tuberculosis-related pathogenic signs. In particular, time-dependent evaluation of pathological changes (on 1, 3, 7, 14, 21, 28, 60 and 120 day post infection) in the lungs and spleen revealed significant differences among various strains. Tuberculosis progression was observed in the mice infected with B0/W148 and NK strains, whereas CladeA, CAO and H37Rv strains resulted in stabilized course and less marked organs damage. Moreover, we found that bacterial load after infection with Beijing family clustering strains was lower compared to that of the reference H37Rv strain, except NK strain demonstrating the peak bacterial load among the Beijing family comparable to H37Rv strain at 120 dpi. Thus, it was found that the level of virulence between most virulent B0/W148 cluster strain vs. NK strain was similar. Overall, the data obtained indicate that Beijing genotype strains are characterized by a diverse range of phenotypic virulence in vivo.

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

ObjectiveThe 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. MethodsThe study included 423 M. tuberculosis strains isolated in 2013–2017 and subjected to drug susceptibility testing, genotyping, and whole genome sequencing. ResultsThe 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. ConclusionsBased 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.

Genetic diversity of drug resistant Mycobacterium Tuberculosis in local area of Southwest China: a retrospective study

BackgroundBy 2014 although tuberculosis (TB) incidence had fallen by an average of 1.5% per year since 2000 and was 18% lower than the level of in 2000, 1.5 million people died for TB in that year. One of reason was that drug resistant Mycobacterium tuberculosis (DRTB) spread. This study aims to determine drug resistant characteristics and genotype of DRTB that isolated from patients in a tuberculosis referral hospital of southwest China.MethodsFive hundred thirty-eight drug resistant tuberculosis samples were collected from July 2013 to March 2015. All the isolates were identified by genomic deletions in region of difference 105 (RD105) and genotyped by mycobacterial interspersed repetitive unit-variable number tandem repeat typing (MIRU-VNTR). Polymorphism and cluster analysis of each locus was carried out using Bionumerics Version 3.0 and phyloviz software.ResultsFive hundred thirty-eight TB strains included 503 Mycobacterium tuberculosis (MTB) isolates and 35 non Mycobacterium tuberculosis (NMTB) isolates. Of 503 isolates Beijing family type was 447 (88.9%, 447/503) and non-Beijing family type was 56 (11.1%, 56/503). Five hundred three DRTB isolates were divided into 345 genotypes, of which 265 isolates were single genotype and the remaining 238 strains were classified into 80 clusters with cluster rate of 47.3% and cluster ratio of 31.4%. Sixty-nine clusters belonged to Beijing family with cluster rate was 48.3% and clustering ratio was 32.9%. The non - Beijing family had 11 clusters with a cluster rate of 39.3% and the clustering ratio of 19.6%. Beijing genotype had a significant correlation with the age (P < 0.05), the retreatment patients (P < 0.05) and the city of Chongqing (P < 0.05), not with gender (P > 0.05). In the 9 Beijing genotype clusters each cluster contained some patients who lived in the same region.ConclusionsBeijing genotype was the predominant in the patients with DRTB in our hospital. In Chongqing retreatment patients with Beijing genotype MTB may be patient with DRTB. Drug resistance test (DST), regular medication and strict follow-up are very important for patients with Beijing genotype MTB. In Chongqing control and treatment of DRTB should be paid attention. Transmission and relations of patients with DRTB need to be further research.