Discordant results of Xpert MTB/Rif assay and BACTEC MGIT 960 liquid culture to detect Mycobacterium tuberculosis in community screening in Vietnam.

BACKGROUND: Xpert MTB/Rif, a molecular test to detect tuberculosis (TB), has been proven to have high sensitivity and specificity when compared with liquid culture in clinical settings. However, little is known about its performance in community TB screening. METHODS: In Vietnam, a national TB prevalence survey was conducted in 2017. Survey participants who screened positive by chest X-ray, cough symptoms and/or recent history of tuberculosis were requested to provide at least two sputum samples that were tested for Mycobacterium tuberculosis by Xpert MTB/Rif G4 (Xpert) and BACTEC MGIT960 culture (MGIT). RESULTS: There were 4,649 eligible participants provided both samples for testing. Among them, 236 (5.1%) participants tested positive for TB by Xpert, 244 (5.3%) tested positive by MGIT and 317 tested positive by at least one test; 163 (51.4%) had discordant test results. Of the positive Xpert, 162 (68.6%) showed a low or very low bacterial load. In multivariate logistic regression comparing discordant with Xpert-MGIT concordant positive results, discordant Xpert-positive results occurred more often among participants who had low sputum bacterial load, male sex, a history of TB treatment, or night sweats. The associated factors were male sex, abnormal chest X-ray and having night sweats when the logistic model was against those with both Xpert and MGIT negative. CONCLUSIONS: We found high rates of discordance in the performance of Xpert and MGIT for community-based TB case finding. In situations where the majority of TB cases are expected to have a low bacterial load, multiple diagnostic tests and/or multiple samples are required to reach sufficient sensitivity.

Social determinants of the changing tuberculosis prevalence in Viet Nam: Analysis of population-level cross-sectional studies.

BACKGROUND: An ecological relationship between economic development and reduction in tuberculosis prevalence has been observed. Between 2007 and 2017, Viet Nam experienced rapid economic development with equitable distribution of resources and a 37% reduction in tuberculosis prevalence. Analysing consecutive prevalence surveys, we examined how the reduction in tuberculosis (and subclinical tuberculosis) prevalence was concentrated between socioeconomic groups. METHODS AND FINDINGS: We combined data from 2 nationally representative Viet Nam tuberculosis prevalence surveys with provincial-level measures of poverty. Data from 94,156 (2007) and 61,763 (2017) individuals were included. Of people with microbiologically confirmed tuberculosis, 21.6% (47/218) in 2007 and 29.0% (36/124) in 2017 had subclinical disease. We constructed an asset index using principal component analysis of consumption data. An illness concentration index was estimated to measure socioeconomic position inequality in tuberculosis prevalence. The illness concentration index changed from -0.10 (95% CI -0.08, -0.16; p = 0.003) in 2007 to 0.07 (95% CI 0.06, 0.18; p = 0.158) in 2017, indicating that tuberculosis was concentrated among the poorest households in 2007, with a shift towards more equal distribution between rich and poor households in 2017. This finding was similar for subclinical tuberculosis. We fitted multilevel models to investigate relationships between change in tuberculosis prevalence, individual risks, household socioeconomic position, and neighbourhood poverty. Controlling for provincial poverty level reduced the difference in prevalence, suggesting that changes in neighbourhood poverty contribute to the explanation of change in tuberculosis prevalence. A limitation of our study is that while tuberculosis prevalence surveys are valuable for understanding socioeconomic differences in tuberculosis prevalence in countries, given that tuberculosis is a relatively rare disease in the population studied, there is limited power to explore socioeconomic drivers. However, combining repeated cross-sectional surveys with provincial deprivation estimates during a period of remarkable economic growth provides valuable insights into the dynamics of the relationship between tuberculosis and economic development in Viet Nam. CONCLUSIONS: We found that with equitable economic growth and a reduction in tuberculosis burden, tuberculosis became less concentrated among the poor in Viet Nam.

High Rates of Recurrent Tuberculosis Disease: A Population-level Cohort Study.

BACKGROUND: Retreatment tuberculosis (TB) disease is common in high-prevalence settings. The risk of repeated episodes of recurrent TB is unknown. We calculated the rate of recurrent TB per subsequent episode by matching individual treatment episodes over a period of 13 years. METHODS: All recorded TB episodes in Cape Town between 2003 and 2016 were matched by probabilistic linkage of personal identifiers. Among individuals with a first episode notified in Cape Town and who completed their prior treatment successfully we estimated the recurrence rate stratified by subsequent episode and HIV status. We adjusted person-time to background mortality by age, sex, and HIV status. RESULTS: A total of 292 915 TB episodes among 263 848 individuals were included. The rate of recurrent TB was 16.4 per 1000 person-years (95% CI, 16.2-16.6), and increased per subsequent episode (8.4-fold increase, from 14.6 to 122.7 per 1000 from episode 2 to 6, respectively). These increases were similar stratified by HIV status. Rates among HIV positives were higher than among HIV negatives for episodes 2 and 3 (2- and 1.5-fold higher, respectively), and the same thereafter. CONCLUSIONS: TB recurrence rates were high and increased per subsequent episode, independent of HIV status. This suggests that HIV infection is insufficient to explain the high burden of recurrence; it is more likely due to a high annual risk of infection combined with an increased risk of infection or progression to disease associated with a previous TB episode. The very high recurrence rates would justify increased TB surveillance of patients with >1 episode.

Correction: The second national tuberculosis prevalence survey in Vietnam.

[This corrects the article DOI: 10.1371/journal.pone.0232142.].

The second national tuberculosis prevalence survey in Vietnam.

INTRODUCTION: Tuberculosis (TB) remains a significant cause of morbidity and mortality in Vietnam. The current TB burden is unknown as not all individuals with TB are diagnosed, recorded and notified. The second national TB prevalence survey was conducted in 2017-2018 to assess the current burden of TB disease in the country. METHOD: Eighty-two clusters were selected using a multistage cluster sampling design. Adult (≥15 years of age) residents having lived for 2 weeks or more in the households of the selected clusters were invited to participate in the survey. The survey participants were screened for TB by a questionnaire and digital chest X-ray after providing written informed consent. Individuals with a positive symptom screen and/or chest X-ray suggestive of TB were asked to provide sputum samples to test for Mycobacterium tuberculosis by Ziehl-Neelsen direct light microscopy, Xpert MTB/RIF G4, BACTEC MGIT960 liquid culture and Löwenstein-Jensen solid culture. Bacteriologically confirmed TB cases were defined by an expert panel following a standard decision tree. RESULT: Of 87,207 eligible residents, 61,763 (70.8%) participated, and 4,738 (7.7%) screened positive for TB. Among these, 221 participants were defined as bacteriologically confirmed TB cases. The estimated prevalence of bacteriologically confirmed adult pulmonary TB was 322 (95% CI: 260-399) per 100,000, and the male-to-female ratio was 4.0 (2.8-5.8, p<0.001). In-depth interviews with the participants with TB disease showed that only 57.9% (95% CI: 51.3-64.3%) reported cough for 2 weeks or more and 32.1% (26.3-38.6%) did not report any symptom consistent with TB, while their chest X-ray results showed that 97.7% (95% CI: 94.6-99.1) had abnormal chest X-ray images suggesting TB. CONCLUSION: With highly sensitive diagnostics applied, this survey showed that the TB burden in Vietnam remains high. Half of the TB cases were not picked up by general symptom-based screening and were identified by chest X-ray only. Our results indicate that improving TB diagnostic capacity and access to care, along with reducing TB stigma, need to be top priorities for TB control and elimination in Vietnam.

How Well Do Routine Molecular Diagnostics Detect Rifampin Heteroresistance in Mycobacterium tuberculosis?

Rifampin heteroresistance-where rifampin-resistant and -susceptible tuberculosis (TB) bacilli coexist-may result in failed standard TB treatment and potential spread of rifampin-resistant strains. The detection of rifampin heteroresistance in routine rapid diagnostic tests (RDTs) allows for patients to receive prompt and effective multidrug-resistant-TB treatment and may improve rifampin-resistant TB control. The limit of detection (LOD) of rifampin heteroresistance for phenotypic drug susceptibility testing by the proportion method is 1% and, yet, is insufficiently documented for RDTs. We, therefore, aimed to determine, for the four RDTs (XpertMTB/RIF, XpertMTB/RIF Ultra, GenoTypeMTBDRplusv2.0, and GenoscholarNTM+MDRTBII), the LOD per probe and mutation, validated by CFU counting and targeted deep sequencing (Deeplex-MycTB). We selected one rifampin-susceptible and four rifampin-resistant strains, with mutations D435V, H445D, H445Y, and S450L, respectively, mixed them in various proportions in triplicate, tested them with each RDT, and determined the LODs per mutation type. Deeplex-MycTB revealed concordant proportions of the minority resistant variants in the mixtures. The Deeplex-MycTB-validated LODs ranged from 20% to 80% for XpertMTB/RIF, 20% to 70% for Xpert Ultra, 5% to 10% for GenoTypeMTBDRplusv2.0, and 1% to 10% for GenoscholarNTM+MDRTBII for the different mutations. Deeplex-MycTB, GenoTypeMTBDRplusv2.0, and GenoscholarNTM+MDRTBII provide explicit information on rifampin heteroresistance for the most frequently detected mutations. Classic Xpert and Ultra report rifampin heteroresistance as rifampin resistance, while Ultra may denote rifampin heteroresistance through "mixed patterns" of wild-type and mutant melt probe, melt peak temperatures. Overall, our findings inform end users that the threshold for reporting resistance in the case of rifampin heteroresistance is the highest for Classic Xpert and Ultra to resolve phenotypic and genotypic discordant rifampin-resistant TB results.

BCG vaccination protects against infection with Mycobacterium tuberculosis ascertained by tuberculin skin testing.

OBJECTIVES: There is conflicting evidence as to whether Bacillus Calmette-Guérin (BCG) vaccination offers protection against Mycobacterium tuberculosis infection ascertained by a positive tuberculin skin test (TST). We investigated the association between BCG vaccination status and TST results in a set of surveys at increasing TST cut-off values to take cross-reactions to BCG vaccination into account. METHODS: Secondary analysis of data from three consecutive tuberculin surveys done among schoolchildren in Tanzania between 1990 and 2002. BCG vaccination status was ascertained by the presence of a typical scar. RESULTS: We analyzed data of 277,588 children of whom 77.7% were BCG vaccinated and 8.5% had TST indurations ≥ 15 mm, 5.1% ≥ 17 mm and 2.8% ≥ 19 mm. In the combined analysis, odds ratios for a positive TST were > 1 for children with BCG up to TST cut-off values of 16 mm. For cut-off values > 17 mm crude and adjusted odds ratios were significantly < 1, and decreased with further increasing cut-off values. CONCLUSIONS: Using a methodology that makes use of the differences in TST reaction sizes between specific and non-specific responses, we showed that BCG vaccination was associated with reduced prevalence of M. tuberculosis infection as measured by the tuberculin skin test, suggesting a protective effect.

Solar ultraviolet B exposure and global variation in tuberculosis incidence: an ecological analysis.

Epidemiological evidence supports vitamin D deficiency as a risk factor for tuberculosis. Differences in solar ultraviolet B (UV-B) exposure, the major source of vitamin D, might therefore partially explain global variation in tuberculosis incidence.In a global country-based ecological study, we explored the correlation between vitamin D-proxies, such as solar UV-B exposure, and other relevant variables with tuberculosis incidence, averaged over the period 2004-2013.Across 154 countries, annual solar UV-B exposure was associated with tuberculosis incidence. Tuberculosis incidence in countries in the highest quartile of UV-B exposure was 78% (95% CI 57-88%, p<0.001) lower than that in countries in the lowest quartile, taking into account other vitamin D-proxies and covariates. Of the explained global variation in tuberculosis incidence, 6.3% could be attributed to variations in annual UV-B exposure. Exposure to UV-B had a similar, but weaker association with tuberculosis notification rates in the multilevel analysis with sub-national level data for large countries (highest versus lowest quartile 29% lower incidence; p=0.057).The potential preventive applications of vitamin D supplementation in high-risk groups for tuberculosis merits further investigation.

Variation and risk factors of drug resistant tuberculosis in sub-Saharan Africa: a systematic review and meta-analysis.

BACKGROUND: Prevalence of multidrug resistant tuberculosis (MDR-TB), defined as in vitro resistance to both rifampicin and isoniazid with or without resistance to other TB drugs, in sub-Saharan Africa (SSA) is reportedly low compared to other regions. These estimates are based on data reported to the World Health Organization (WHO) on drug resistance surveys, which may suffer from a reporting bias. We set out to evaluate the variation in prevalence of drug resistant tuberculosis (DR-TB) and its determinants across SSA countries among new and previously treated TB patients. METHODS: The aim was to perform a systematic review and meta-analysis of DR-TB prevalence and associated risk factors in SSA. PubMed, EMBASE, Cochrane and bibliographies of DR-TB studies were searched. Surveys at national or sub-national level, with reported DR-TB prevalence (or sufficient data to calculate a prevalence) to isoniazid (INH), rifampicin (RMP), ethambutol (EMB), and streptomycin (SM) conducted in SSA excluding the Republic of South Africa, published between 2003 and 2013 with no language restriction were considered. Two authors searched and reviewed the studies for eligibility and extracted the data in pre-defined forms. Forest plots of all prevalence estimates by resistance outcome were performed. Summary estimates were calculated using random effects models, when appropriate. Associations between any DR-TB and MDR-TB with potential risk factors were examined through subgroup analyses stratified by new and previously treated patients. RESULTS: A total of 726 studies were identified, of which 27 articles fulfilled the inclusion criteria. Studies reported drug susceptibility testing (DST) results for a total of 13,465 new and 1,776 previously treated TB patients. Pooled estimate of any DR-TB prevalence among the new cases was 12.6% (95% CI 10.6-15.0) while for MDR-TB this was 1.5% (95% CI 1.0-2.3). Among previously treated patients, these were 27.2% (95% CI 21.4-33.8) and 10.3% (95% CI 5.8-17.4%), respectively. DR-TB (any and MDR-TB) did not vary significantly with respect to study characteristics. CONCLUSIONS: The reported prevalence of DR-TB in SSA is low compared to WHO estimates. MDR-TB in this region does not seem to be driven by the high HIV prevalence rates.

The Mycobacterium tuberculosis Uganda II family and resistance to first-line anti-tuberculosis drugs in Uganda.

BACKGROUND: The global increase in the burden of multidrug-resistant tuberculosis (MDR-TB) underscores an urgent need for data on factors involved in generation and spread of TB drug resistance. We performed molecular analyses on a representative sample of Mycobacterium tuberculosis (MTB) isolates. Basing on findings of the molecular epidemiological study in Kampala, we hypothesized that the predominant MTB strain lineage in Uganda is negatively associated with anti-TB drug resistance and we set out to test this hypothesis. METHODS: We extracted DNA from mycobacterial isolates collected from smear-positive TB patients in the national TB drug resistance survey and carried out IS6110-PCR. To identify MTB lineages/sub lineages RT-PCR SNP was performed using specific primers and hybridization probes and the 'melting curve' analysis was done to distinguish the Uganda II family from other MTB families. The primary outcome was the distribution of the Uganda II family and its associations with anti-TB drug resistance and HIV infection. RESULTS: Out of the 1537 patients enrolled, MTB isolates for 1001 patients were available for SNP analysis for identification of Uganda II family, of which 973 (97%) had conclusive RT-PCR results. Of these 422 (43.4%) were of the Uganda II family, mostly distributed in the south west zone (55.0%; OR = 4.6 for comparison with other zones; 95% CI 2.83-7.57; p < 0.001) but occurred in each of the other seven geographic zones at varying levels. Compared to the Uganda II family, other genotypes as a group were more likely to be resistant to any anti-TB drug (OR(adj) =2.9; 95% CI 1.63-5.06; p = 0.001) or MDR (OR(adj) 4.9; 95% CI, 1.15-20.60; p = 0.032), even after adjusting for geographic zone, patient category, sex, residence and HIV status. It was commonest in the 25-34 year age group 159/330 (48.2%). No association was observed between Uganda II family and HIV infection. CONCLUSION: The Uganda II family is a major cause of morbidity due to TB in all NTLP zones in Uganda. It is less likely to be resistant to anti-TB drugs than other MTB strain lineages.

A household survey on screening practices of household contacts of smear positive tuberculosis patients in Vietnam.

BACKGROUND: Close contacts of tuberculosis (TB) patients are at increased risk of developing tuberculosis. Although passive contact screening guidelines are incorporated in the national TB control program, currently it is unknown how frequent close contacts are screened for TB in Vietnam. This study assesses current contact screening practices in Vietnam and determines the proportion of household contacts screened of newly registered TB patients. METHOD: Survey of household contacts of smear-positive TB patients (index patients) registered for treatment in 2008 in three Vietnamese cities. Households were interviewed in 2010 about screening for TB since treatment registration date of the index patient. RESULTS: We interviewed 4,118 household contacts of 1,091 identified index cases. Contact screening mainly relied on self-referral by household contacts. Of the 4,118 household contacts, 474 (11.5%) self-referred for TB screening, while this screening proportion was only 5.5% among contacts under 5 years old (16/293). Sputum examinations were performed in 374 (78.9%) of the screened contacts. Contact screening identified 27 cases of pulmonary TB (0.7%; or 656 cases/100,000 contacts), of which 20 were detected by sputum smear. CONCLUSIONS: The low proportion of household TB contacts screened for TB illustrates the limitations of passive contact screening as currently practiced in Vietnam. Children under 5 years of age are particularly neglected with this approach. Active contact screening with fixed follow-up times of close contacts of newly diagnosed TB patients should be considered in Vietnam, particularly in case of young children and drug-resistant TB.

The T2 Mycobacterium tuberculosis genotype, predominant in Kampala, Uganda, shows negative correlation with antituberculosis drug resistance.

Surveillance of the circulating Mycobacterium tuberculosis complex (MTC) strains in a given locality is important for understanding tuberculosis (TB) epidemiology. We performed molecular epidemiological studies on sputum smear-positive isolates that were collected for anti-TB drug resistance surveillance to establish the variability of MTC lineages with anti-TB drug resistance and HIV infection. Spoligotyping was performed to determine MTC phylogenetic lineages. We compared patients' MTC lineages with drug susceptibility testing (DST) patterns and HIV serostatus. Out of the 533 isolates, 497 (93.2%) had complete DST, PCR, and spoligotyping results while 484 (90.1%) participants had results for HIV testing. Overall, the frequency of any resistance was 75/497 (15.1%), highest among the LAM (34.4%; 95% confidence interval [CI], 18.5 to 53.2) and lowest among the T2 (11.5%; 95% CI, 7.6 to 16.3) family members. By multivariate analysis, LAM (adjusted odds ratio [OR(adj)], 5.0; 95% CI, 2.0 to 11.9; P < 0.001) and CAS (OR(adj), 2.9; 95% CI, 1.4.0 to 6.3; P = 0.006) families were more likely to show any resistance than was T2. All other MTC lineages combined were more likely to be resistant to any of the anti-TB drugs than were the T2 strains (OR(adj), 1.7; 95% CI, 1.0 to 2.9; P = 0.040). There were no significant associations between multidrug resistance and MTC lineages, but numbers of multidrug-resistant TB strains were small. No association was established between MTC lineages and HIV status. In conclusion, the T2 MTC lineage negatively correlates with anti-TB drug resistance, which might partly explain the reported low levels of anti-TB drug resistance in Kampala, Uganda. Patients' HIV status plays no role with respect to the MTC lineage distribution.

Epidemiology of isoniazid resistance mutations and their effect on tuberculosis treatment outcomes.

Isoniazid resistance is highly prevalent in Vietnam. We investigated the molecular and epidemiological characteristics and the association with first-line treatment outcomes of the main isoniazid resistance mutations in Mycobacterium tuberculosis in codon 315 of the katG and in the promoter region of the inhA gene. Mycobacterium tuberculosis strains with phenotypic resistance to isoniazid from consecutively diagnosed smear-positive tuberculosis patients in rural Vietnam were subjected to Genotype MTBDRplus testing to identify katG and inhA mutations. Treatment failure and relapse were determined by sputum culture. In total, 227 of 251 isoniazid-resistant strains (90.4%) had detectable mutations: 75.3% in katG codon 315 (katG315) and 28.2% in the inhA promoter region. katG315 mutations were significantly associated with pretreatment resistance to streptomycin, rifampin, and ethambutol but not with the Beijing genotype and predicted both unfavorable treatment outcome (treatment failure or death) and relapse; inhA promoter region mutations were only associated with resistance to streptomycin and relapse. In tuberculosis patients, M. tuberculosis katG315 mutations but not inhA mutations are associated with unfavorable treatment outcome. inhA mutations do, however, increase the risk of relapse, at least with treatment regimens that contain only isoniazid and ethambutol in the continuation phase.

For whom the bell tolls: isoniazid preventive therapy and tuberculosis drug resistance.

The expected increase in drug-resistant tuberculosis due to large-scale preventive treatment in people living with HIV calls for reconsidering the "double use" of isoniazid for prophylaxis and curative treatment (Mills et al., this issue).

Alignment of new tuberculosis drug regimens and drug susceptibility testing: a framework for action.

New tuberculosis drug regimens are creating new priorities for drug susceptibility testing (DST) and surveillance. To minimise turnaround time, rapid DST will need to be prioritised, but developers of these assays will need better data about the molecular mechanisms of resistance. Efforts are underway to link mutations with drug resistance and to develop strain collections to enable assessment of new diagnostic assays. In resource-limited settings, DST might not be appropriate for all patients with tuberculosis. Surveillance data and modelling will help country stakeholders to design appropriate DST algorithms and to decide whether to change drug regimens. Finally, development of practical DST assays is needed so that, in countries where surveillance and modelling show that DST is advisable, these assays can be used to guide clinical decisions for individual patients. If combined judiciously during both development and implementation, new tuberculosis regimens and new DST assays have enormous potential to improve patient outcomes and reduce the burden of disease.

Clustering of Beijing genotype Mycobacterium tuberculosis isolates from the Mekong delta in Vietnam on the basis of variable number of tandem repeat versus restriction fragment length polymorphism typing.

BACKGROUND: In comparison to restriction fragment length polymorphism (RFLP) typing, variable number of tandem repeat (VNTR) typing is easier to perform, faster and yields results in a simple, numerical format. Therefore, this technique has gained recognition as the new international gold standard in typing of Mycobacterium tuberculosis. However, some reports indicated that VNTR typing may be less suitable for Beijing genotype isolates. We therefore compared the performance of internationally standardized RFLP and 24 loci VNTR typing to discriminate among 100 Beijing genotype isolates from the Southern Vietnam. METHODS: Hundred Beijing genotype strains defined by spoligotyping were randomly selected and typed by RFLP and VNTR typing. The discriminatory power of VNTR and RFLP typing was compared using the Bionumerics software. RESULTS: Among 95 Beijing strains available for analysis, 14 clusters were identified comprising 34 strains and 61 unique profiles in 24 loci VNTR typing ((Hunter Gaston Discrimination Index (HGDI = 0.994)). 13 clusters containing 31 strains and 64 unique patterns in RFLP typing (HGDI = 0.994) were found. Nine RFLP clusters were subdivided by VNTR typing and 12 VNTR clusters were split by RFLP. Five isolates (5%) revealing double alleles or no signal in two or more loci in VNTR typing could not be analyzed. CONCLUSIONS: Overall, 24 loci VNTR typing and RFLP typing had similar high-level of discrimination among 95 Beijing strains from Southern Vietnam. However, loci VNTR 154, VNTR 2461 and VNTR 3171 had hardly added any value to the level of discrimination.

Tuberculosis relapse in Vietnam is significantly associated with Mycobacterium tuberculosis Beijing genotype infections.

BACKGROUND: In Vietnam, the Mycobacterium tuberculosis Beijing genotype is associated with multi-drug resistance and is emerging. A possible explanation for this genotype's success is an increased rate of relapse. METHODS: In a prospective cohort study, isolates from patients with smear-positive tuberculosis were subjected to drug susceptibility testing and to spoligotyping and variable number of tandem repeats typing before treatment and after recurrence of tuberculosis. RESULTS: Among 1068 patients who were actively followed up over 18 months for recurrence, 23 relapse cases occurred (1.39 cases/100 person-years). After adjustment for genotype, tuberculosis treatment history, and drug resistance, relapse was significantly associated with the Beijing genotype (adjusted hazard ratio [aHR], 5.48; 95% confidence interval [CI], 2.06-14.55) and isoniazid resistance (aHR, 5.91; 95% CI, 2.16-16.16). CONCLUSIONS: The strongly increased relapse rate in tuberculosis cases caused by Beijing strains probably contributes to the successful spread of this genotype in Vietnam and elsewhere.