Clinical and operational value of the extensively drug-resistant tuberculosis definition.

Currently, no information is available on the effect of resistance/susceptibility to first-line drugs different from isoniazid and rifampicin in determining the outcome of extensively drug-resistant tuberculosis (XDR-TB) patients, and whether being XDR-TB is a more accurate indicator of poor clinical outcome than being resistant to all first-line anti-tuberculosis (TB) drugs. To investigate this issue, a large series of multidrug-resistant TB (MDR-TB) and XDR-TB cases diagnosed in Estonia, Germany, Italy and the Russian Federation during the period 1999-2006 were analysed. Drug-susceptibility testing for first- and second-line anti-TB drugs, quality assurance and treatment delivery was performed according to World Health Organization recommendations in all study sites. Out of 4,583 culture-positive TB cases analysed, 361 (7.9%) were MDR and 64 (1.4%) were XDR. XDR-TB cases had a relative risk (RR) of 1.58 to have an unfavourable outcome compared with MDR-TB cases resistant to all first-line drugs (isoniazid, rifampicin ethambutol, streptomycin and, when tested, pyrazinamide), and an RR of 2.61 compared with "other" MDR-TB cases (those susceptible to at least one first-line anti-TB drug among ethambutol, pyrazinamide and streptomycin, regardless of resistance to the second-line drugs not defining XDR-TB). The emergence of extensively drug-resistant tuberculosis confirms that problems in tuberculosis management are still present in Europe. While waiting for new tools which will facilitate management of extensively drug-resistant tuberculosis, accessibility to quality diagnostic and treatment services should be urgently ensured and adequate public health policies should be rapidly implemented to prevent further development of drug resistance

Performance of the Genotype® MTBDRPlus assay in the diagnosis of tuberculosis and drug resistance in Samara, Russian Federation

<p>Abstract</p> <p>Background</p> <p>Russia is a high tuberculosis (TB) burden country with a high prevalence of multidrug resistant tuberculosis (MDRTB). Molecular assays for detection of MDRTB on clinical specimens are not widely available in Russia.</p> <p>Results</p> <p>We performed an evaluation of the GenoType^®MTBDRplus assay (HAIN Lifescience GmbH, Germany) on a total of 168 sputum specimens from individual patients at a public health laboratory in Central Russia, as a model of a middle income site in a region with high levels of drug resistance. Phenotypic drug resistance tests (DST) were performed on cultures derived from the same sputum specimens using the BACTEC 960 liquid media system.</p> <p>Interpretable GenoType^®MTBDRplus results were obtained for 154(91.7%) specimens with readability rates significantly higher in sputum specimens graded 2+ and 3+ compared to 1+ (RR = 1.17 95%CI 1.04–1.32). The sensitivity and specificity of the assay for the detection of rifampicin (RIF) and isoniazid (INH) resistance and MDR was 96.2%, 97.4%, 97.1% and 90.7%, 83.3%, 88.9% respectively. Mutations in codon 531 of the <it>rpoB </it>gene and codon 315 of the <it>katG </it>gene dominated in RIF and INH resistant strains respectively. Disagreements between phenotypical and molecular tests results (12 samples) could be explained by the presence of rare mutations in strains circulating in Russia and simultaneous presence of resistant and sensitive bacilli in sputum specimens (heteroresistance).</p> <p>Conclusion</p> <p>High sensitivity, short turnaround times and the potential for screening large numbers of specimens rapidly, make the GenoType^®MTBDRplus assay suitable as a first-line screening assay for drug resistant TB.</p

Molecular epidemiology and evolutionary genetics of Mycobacterium tuberculosis in Taipei

<p>Abstract</p> <p>Background</p> <p>The control of tuberculosis in densely populated cities is complicated by close human-to-human contacts and potential transmission of pathogens from multiple sources. We conducted a molecular epidemiologic analysis of 356 <it>Mycobacterium tuberculosis </it>(MTB) isolates from patients presenting pulmonary tuberculosis in metropolitan Taipei. Classical antibiogram studies and genetic characterization, using mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing and spoligotyping, were applied after culture.</p> <p>Methods</p> <p>A total of 356 isolates were genotyped by standard spoligotyping and the strains were compared with in the international spoligotyping database (SpolDB4). All isolates were also categorized using the 15 loci MIRU-VNTR typing method and combin with <it>NTF </it>locus and RD deletion analyses.</p> <p>Results</p> <p>Of 356 isolates spoligotyped, 290 (81.4%) displayed known spoligotypes and 66 were not identified in the database. Major spoligotypes found were Beijing lineages (52.5%), followed by Haarlem lineages (13.5%) and EAI plus EAI-like lineages (11%). When MIRU-VNTR was employed, 140 patterns were identified, including 36 clusters by 252 isolates and 104 unique patterns, and the largest cluster comprised 95 isolates from the Beijing family. The combination of spoligotyping and MIRU-VNTR revealed that 236 (67%) of the 356 isolates were clustered in 43 genotypes. Strains of the Beijing family was more likely to be of modern strain and a higher percentage of multiple drug resistance than other families combined (P = 0.08). Patients infected with Beijing strains were younger than those with other strains (mean 58.7 vs. 64.2, p = 0.02). Moreover, 85.3% of infected persons younger than 25 years had Beijing modern strain, suggesting a possible recent spread in the young population by this family of TB strain in Taipei.</p> <p>Conclusion</p> <p>Our data on MTB genotype in Taipei suggest that MTB infection has not been optimally controlled. Control efforts should be reinforced in view of the high prevalence of the Beijing strain in young population and association with drug resistance.</p

A novel approach : the propensity to propagate (PTP) method for controlling for host factors in studying the transmission of Mycobacterium tuberculosis

RATIONALE: Understanding the genetic variations among Mycobacterium tuberculosis (MTB) strains with differential ability to transmit would be a major step forward in preventing transmission. OBJECTIVES: To describe a method to extend conventional proxy measures of transmissibility by adjusting for patient-related factors, thus strengthening the causal association found with bacterial factors. METHODS: Clinical, demographic and molecular fingerprinting data were obtained during routine surveillance of verified MTB cases reported in the Netherlands between 1993 and 2011, and the phylogenetic lineages of the isolates were inferred. Odds ratios for host risk factors for clustering were used to obtain a measure of each patient's and cluster's propensity to propagate (CPP). Mean and median cluster sizes across different categories of CPP were compared amongst four different phylogenetic lineages. RESULTS: Both mean and median cluster size grew with increasing CPP category. On average, CPP values from Euro-American lineage strains were higher than Beijing and EAI strains. There were no significant differences between the mean and median cluster sizes among the four phylogenetic lineages within each CPP category. CONCLUSIONS: Our finding that the distribution of CPP scores was unequal across four different phylogenetic lineages supports the notion that host-related factors should be controlled for to attain comparability in measuring the different phylogenetic lineages' ability to propagate. Although Euro-American strains were more likely to be in clusters in an unadjusted analysis, no significant differences among the four lineages persisted after we controlled for host factors.Portuguese Foundation for Science and Technology (FCT) (SFRH/BD/33902/2009 to HN-G). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Reduced Transmissibility of East African Indian Strains of Mycobacterium tuberculosis

BACKGROUND: Mycobacterium tuberculosis (MTB) has been classified into 4 main lineages. Some reports have associated certain lineages with particular clinical phenotypes, but there is still insufficient information regarding the clinical and epidemiologic implications of MTB lineage variation. METHODS: Using large sequence polymorphisms we classified MTB isolates from a population-based study in Montreal, Canada into the 4 major lineages, and identified the associated clinical and epidemiologic features. In addition, IS6110-RFLP and spoligotyping were used as indicators of recent TB transmission. The study population was divided into a derivation cohort, diagnosed between 2001 and 2007, and a separate validation cohort, diagnosed between 1996 and 2000. RESULTS: In the derivation cohort, when compared to the other MTB lineages, the East African-Indian (EAI) lineage was associated with lower rates of TB transmission, as measured by: positive TST among close contacts of pulmonary TB cases (adjusted odds ratio 0.6: [95% confidence interval 0.4-0.9]), and clustered TB cases (0.3: [<0.001-0.6]). Severe forms of TB were also less likely among the EAI group (0.4: [<0.001-0.8]). There were no significant differences when comparing patients with the other MTB lineages. In the validation cohort, the EAI lineage was associated with lower rates of positive TST among contacts (0.5: [0.3-0.9]) and a trend towards less clustered TB cases (0.5: [0.1-1.8]) when compared to the other lineages. Disease severity among the different groups was not significantly different in the validation cohort. CONCLUSIONS: We conclude that in Montreal, EAI strains were associated with reduced transmission compared to other MTB lineages

Tuberculosis associated with Mycobacterium tuberculosis Beijing and non-Beijing genotypes: a clinical and immunological comparison

BACKGROUND: The Mycobacterium tuberculosis Beijing genotype is biologically different from other genotypes. We aimed to clinically and immunologically compare human tuberculosis caused by Beijing and non-Beijing strains. METHODS: Pulmonary tuberculosis patients were prospectively enrolled and grouped by their M. tuberculosis genotypes. The clinical features, plasma cytokine levels, and cytokine gene expression levels in peripheral blood mononuclear cells (PBMC) were compared between the patients in Beijing and non-Beijing groups. RESULTS: Patients in the Beijing group were characterized by significantly lower frequency of fever (odds ratio, 0.12, p = 0.008) and pulmonary cavitation (odds ratio, 0.2, p = 0.049). Night sweats were also significantly less frequent by univariate analysis, and the duration of cough prior to diagnosis was longer in Beijing compared to non-Beijing groups (medians, 60 versus 30 days, p = 0.048). The plasma and gene expression levels of interferon (IFN) γ and interleukin (IL)-18 were similar in the two groups. However, patients in the non-Beijing group had significantly increased IL-4 gene expression (p = 0.018) and lower IFN-γ : IL-4 cDNA copy number ratios (p = 0.01). CONCLUSION: Patients with tuberculosis caused by Beijing strains appear to be less symptomatic than those who have disease caused by other strains. Th1 immune responses are similar in patients infected with Beijing and non-Beijing strains, but non-Beijing strains activate more Th2 immune responses compared with Beijing strains, as evidenced by increased IL-4 expression

Drug Resistant Mycobacterium tuberculosis of the Beijing Genotype Does Not Spread in Sweden

BACKGROUND: Drug resistant (DR) and multi-drug resistant (MDR) tuberculosis (TB) is increasing worldwide. In some parts of the world 10% or more of new TB cases are MDR. The Beijing genotype is a distinct genetic lineage of Mycobacterium tuberculosis, which is distributed worldwide, and has caused large outbreaks of MDR-TB. It has been proposed that certain lineages of M. tuberculosis, such as the Beijing lineage, may have specific adaptive advantages. We have investigated the presence and transmission of DR Beijing strains in the Swedish population. METHODOLOGY/PRINCIPAL FINDINGS: All DR M. tuberculosis complex isolates between 1994 and 2008 were studied. Isolates that were of Beijing genotype were investigated for specific resistance mutations and phylogenetic markers. Seventy (13%) of 536 DR strains were of Beijing genotype. The majority of the patients with Beijing strains were foreign born, and their country of origin reflects the countries where the Beijing genotype is most prevalent. Multidrug-resistance was significantly more common in Beijing strains than in non-Beijing strains. There was a correlation between the Beijing genotype and specific resistance mutations in the katG gene, the mabA-inhA-promotor and the rpoB gene. By a combined use of RD deletions, spoligotyping, IS1547, mutT gene polymorphism and Rv3135 gene analysis the Beijing strains could be divided into 11 genomic sublineages. Of the patients with Beijing strains 28 (41%) were found in altogether 10 clusters (2-5 per cluster), as defined by RFLP IS6110, while 52% of the patients with non-Beijing strains were in clusters. By 24 loci MIRU-VNTR 31 (45%) of the patients with Beijing strains were found in altogether 7 clusters (2-11 per cluster). Contact tracing established possible epidemiological linkage between only two patients with Beijing strains. CONCLUSIONS/SIGNIFICANCE: Although extensive outbreaks with non-Beijing TB strains have occurred in Sweden, Beijing strains have not taken hold, in spite of the proximity to high prevalence countries such as Russia and the Baltic countries. The Beijing sublineages so far introduced in Sweden may not be adapted to spread in the Scandinavian population

The Beijing genotype and drug resistant tuberculosis in the Aral Sea region of Central Asia

BACKGROUND: After the collapse of the Soviet Union, dramatically increasing rates of tuberculosis and multidrug-resistant tuberculosis (MDR-TB) have been reported from several countries. This development has been mainly attributed to the widespread breakdown of TB control systems and declining socio-economic status. However, recent studies have raised concern that the Beijing genotype of Mycobacterium tuberculosis might be contributing to the epidemic through its widespread presence and potentially enhanced ability to acquire resistance. METHODS: A total of 397 M. tuberculosis strains from a cross sectional survey performed in the Aral Sea region in Uzbekistan and Turkmenistan have been analysed by drug susceptibility testing, IS6110 fingerprinting, and spoligotyping. RESULTS: Fifteen isolates showed mixed banding patterns indicating simultaneous infection with 2 strains. Among the remaining 382 strains, 152 (40%) were grouped in 42 clusters with identical fingerprint and spoligotype patterns. Overall, 50% of all isolates were Beijing genotype, with 55% of these strains appearing in clusters compared to 25% of non-Beijing strains. The percentage of Beijing strains increased with increasing drug resistance among both new and previously treated patients; 38% of fully-susceptible isolates were Beijing genotype, while 75% of MDR-TB strains were of the Beijing type. CONCLUSION: The Beijing genotype is a major cause of tuberculosis in this region, it is strongly associated with drug resistance, independent of previous tuberculosis treatment and may be strongly contributing to the transmission of MDR-TB. Further investigation around the consequences of Beijing genotype infection for both tuberculosis transmission and outcomes of standard short course chemotherapy are urgently needed

Distribution of Spoligotyping Defined Genotypic Lineages among Drug-Resistant Mycobacterium tuberculosis Complex Clinical Isolates in Ankara, Turkey

Background: Investigation of genetic heterogeneity and spoligotype-defined lineages of drug-resistant Mycobacterium tuberculosis clinical isolates collected during a three-year period in two university hospitals and National Tuberculosis Reference and Research Laboratory in Ankara, Turkey. Methods and Findings: A total of 95 drug-resistant M. tuberculosis isolates collected from three different centers were included in this study. Susceptibility testing of the isolates to four major antituberculous drugs was performed using proportion method on Löwenstein–Jensen medium and BACTEC 460-TB system. All clinical isolates were typed by using spoligotyping and IS6110-restriction fragment length polymorphism (RFLP) methods. Seventy-three of the 95 (76.8%) drug resistant M. tuberculosis isolates were isoniazid-resistant, 45 (47.4%) were rifampicin-resistant, 32 (33.7%) were streptomycinresistant and 31 (32.6%) were ethambutol-resistant. The proportion of multidrug-resistant isolates (MDR) was 42.1%. By using spoligotyping, 35 distinct patterns were observed; 75 clinical isolates were grouped in 15 clusters (clustering rate of 79%) and 20 isolates displayed unique patterns. Five of these 20 unique patterns corresponded to orphan patterns in th

Shedding light on the performance of a pyrosequencing assay for drug-resistant tuberculosis diagnosis

BACKGROUND: Rapid molecular diagnostics, with their ability to quickly identify genetic mutations associated with drug resistance in Mycobacterium tuberculosis clinical specimens, have great potential as tools to control multi- and extensively drug-resistant tuberculosis (M/XDR-TB). The Qiagen PyroMark Q96 ID system is a commercially available pyrosequencing (PSQ) platform that has been validated for rapid M/XDR-TB diagnosis. However, the details of the assay’s diagnostic and technical performance have yet to be thoroughly investigated in diverse clinical environments. METHODS: This study evaluates the diagnostic performance of the PSQ assay for 1128 clinical specimens from patients from three areas of high TB burden. We report on the diagnostic performance of the PSQ assay between the three sites and identify variables associated with poor PSQ technical performance. RESULTS: In India, the sensitivity of the PSQ assay ranged from 89 to 98 % for the detection of phenotypic resistance to isoniazid, rifampicin, fluoroquinolones, and the injectables. In Moldova, assay sensitivity ranged from 7 to 94 %, and in South Africa, assay sensitivity ranged from 71 to 92 %. Specificity was high (94–100 %) across all sites. The addition of eis promoter sequencing information greatly improved the sensitivity of kanamycin resistance detection in Moldova (7 % to 79 %). Nearly all (89.4 %) sequencing reactions conducted on smear-positive, culture-positive specimens and most (70.8 %) reactions conducted on smear-negative, culture-positive specimens yielded valid PSQ reads. An investigation into the variables influencing sequencing failures indicated smear negativity, culture negativity, site (Moldova), and sequencing of the rpoB, gyrA, and rrs genes were highly associated with poor PSQ technical performance (adj. OR > 2.0). CONCLUSIONS: This study has important implications for the global implementation of PSQ as a molecular TB diagnostic, as it demonstrates how regional factors may impact PSQ diagnostic performance, while underscoring potential gene targets for optimization to improve overall PSQ assay technical performance. TRIAL REGISTRATION: ClinicalTrials.gov (#NCT02170441). Registered 12 June 2014. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12879-016-1781-y) contains supplementary material, which is available to authorized users