Mycobacterial Interspersed Repetitive Units-variable Number Of Tandem Repeats Research Articles

Recurrence Is a Noticeable Cause of Rifampicin-Resistant Mycobacterium tuberculosis in the Elderly Population in Jiangxi, China.

Setting: Rifampicin-resistant tuberculosis (RR-TB) in elderly people in Jiangxi Province, China.Objective: To investigate the incidence of RR-TB and risk factors in elderly people within a hospital setting in China.Design: Retrospective cohort study.Methods: A comparative study was performed to analyze RR-TB and rifampicin-susceptible TB (RS-TB). The 15-locus mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTR) method was used to distinguish between relapse and reinfection.Results: Twenty-three recurrent cases occurred in 151 elderly patients with RR-TB, and 24 recurrent cases occurred in 466 elderly patients with RS-TB during this period. TB recurrence was significantly different in the RR-TB and RS-TB groups (OR = 0.35, 95% CI: 0.14–0.88; χ2 = 5.28, P = 0.03). Comparing the risk factors for RR-TB and RS-TB, we found that educational level, age, and pulmonary cavity were inextricably linked to RR-TB in elderly patients. Furthermore, pulmonary cavity, HIV status, and alcohol consumption were associated with recurrence in elderly RR-TB patients.Conclusions: Recurrence is an important source of RR-TB in the elderly population. It is necessary to promptly treat tuberculosis patients, prevent the spread of AIDS, and reduce alcohol intake to control recurrent RR-TB in the elderly population.

Highly successful treatment outcome of multidrug‐resistant and genetic diversity of multidrug‐resistant Mycobacterium tuberculosis strains in Rwanda

To determine prevalent MDR-TB genotypes and describe treatment outcome and bacteriology conversion in MDR-TB patients. Review of laboratory records of 173 MDR-TB patients from all over Rwanda who initiated treatment under programmatic management of MDR-TB (PMDT) between 2014 and 2015. Fifty available archived isolates were genotyped by mycobacterial interspersed repetitive units - variable number of tandem repeats (MIRU-VNTR) genotyping. Of the 170 patients whose outcome was known, 114 (66.3%) were cured and 36 (21%) completed the treatment, giving a successful outcome (cured and completed) of 150 (87.3%) patients. Of 20 MDR-TB patients with unfavourable treatment outcome, 18 died, one failed and one defaulted/stopped treatment. Of the 18 patients who died, 11 (61%) were HIV-coinfected. The treatment outcome was successful for 93.9% among HIV negative and 81.8% among HIV-coinfected patients (P=0.02). Sputum smear conversion occurred in 3, 46, 57 and 78 patients before 2, 3, 4 and 6months, respectively, with median time of sputum smear and culture conversion at 3months. The 44 MDR-TB isolates with MIRU-VNTR result, showed high genetic diversity with low clustering rate (9.09%) and Uganda II being the most prevalent sub-family lineage detected in 68.2% of isolates. Beijing family was the least common genotype detected (2.3%, 1 isolate). The high success rates for MDR-TB treatment achieved in Rwanda were comparable to outcomes observed in resource-rich settings with HIV being an independent risk factor for poor treatment outcome. High genetic diversity and low clustering rate reported here suggest that reactivation of previous disease plays an important role in the transmission of MDR-TB in Rwanda.

Universal genotyping reveals province-level differences in the molecular epidemiology of tuberculosis.

ObjectivesCompare the molecular epidemiology of tuberculosis (TB) between two large Canadian provinces–Ontario and British Columbia (BC)–to identify genotypic clusters within and across both provinces, allowing for an improved understanding of genotype data and providing context to more accurately identify clusters representing local transmission.DesignWe compared 24-locus Mycobacterial Interspersed Repetitive Units-Variable Number of Tandem Repeats (MIRU-VNTR) genotyping for 3,314 Ontario and 1,602 BC clinical Mycobacterium tuberculosis isolates collected from 2008 through 2014. Laboratory data for each isolate was linked to case-level records to obtain clinical and demographic data.ResultsThe demographic characteristics of persons with TB varied between provinces, most notably in the proportion of persons born outside Canada, which was reflected in the large number of unique genotypes (n = 3,461). The proportion of clustered isolates was significantly higher in BC. Substantial clustering amongst non-Lineage 4 TB strains was observed within and across the provinces. Only two large clusters (≥10 cases/cluster) representing within province transmission had interprovincial genotype matches.ConclusionWe recommend expanding analysis of shared genotypes to include neighbouring jurisdictions, and implementing whole genome sequencing to improve identification of TB transmission, recognize outbreaks, and monitor changing trends in TB epidemiology.

Whole genome sequencing for improved understanding of Mycobacterium tuberculosis transmission in a remote circumpolar region.

Few studies have used genomic epidemiology to understand tuberculosis (TB) transmission in rural and remote settings - regions often unique in history, geography and demographics. To improve our understanding of TB transmission dynamics in Yukon Territory (YT), a circumpolar Canadian territory, we conducted a retrospective analysis in which we combined epidemiological data collected through routine contact investigations with clinical and laboratory results. Mycobacterium tuberculosis isolates from all culture-confirmed TB cases in YT (2005-2014) were genotyped using 24-locus Mycobacterial Interspersed Repetitive Units-Variable Number of Tandem Repeats (MIRU-VNTR) and compared to each other and to those from the neighbouring province of British Columbia (BC). Whole genome sequencing (WGS) of genotypically clustered isolates revealed three sustained transmission networks within YT, two of which also involved BC isolates. While each network had distinct characteristics, all had at least one individual acting as the probable source of three or more culture-positive cases. Overall, WGS revealed that TB transmission dynamics in YT are distinct from patterns of spread in other, more remote Northern Canadian regions, and that the combination of WGS and epidemiological data can provide actionable information to local public health teams.

Set-up and validation of mycobacterial interspersed repetitive unit-variable number of tandem repeat (MIRU-VNTR) analysis of Mycobacterium tuberculosis using BioNumerics software.

The objective was to describe and validate a new and alternative software procedure for 24-locus mycobacterial interspersed repetitive unit-variable number-tandem repeat (MIRU-VNTR) typing of Mycobacterium tuberculosis (Mtb) based on the multipurpose BioNumerics software. DNA from randomly selected isolates of Mtb from two European laboratories, including external control samples for MIRU-VNTR typing, were analysed. Samples were genotyped using the commercial 24-locus VNTR typing kit from GenoScreen. The PCR amplified fragments were separated by capillary electrophoresis. For the subsequent analyses, the currently used software GeneMapper was compared with BioNumerics. The endpoint was the level of concordance when comparing genotyping results obtained from BioNumerics with results obtained from GeneMapper and the ECDC proficiency study reference results. Also, the number of necessary manual standard size corrections and allele assignments in the two different software methods were compared. In total, 272 DNA samples, including the ECDC proficiency panel, were analysed. For all samples, there were 100% concordance of results. For a randomly selected set of 96 samples the numbers of manual corrections needed for size standards were 199 with GeneMapper versus zero for BioNumerics. The numbers of manual corrections for allele assignments were 122 with GeneMapper versus 16 with BioNumerics. In conclusion, we have validated the multipurpose software BioNumerics for standard 24-locus MIRU-VNTR typing and the software shows promising benefits in terms of simplification and minimization of hand-on time.

Analysis of potential household transmission events of tuberculosis in the city of Belem, Brazil

Tuberculosis (TB) is an infectious disease with a higher risk for infection and disease among household contacts (HHC). Here, we report a molecular epidemiology-based approach to study disease transmission and the genetic characteristics of Mycobacterium tuberculosis (Mtb) strains among HHC in the city of Belem, the capital of the state of Para in north Brazil. The study included 63 TB patients belonging to 26 HHC groups (HHC1 to HHC26). Spoligotyping and 24-loci Mycobacterial Interspersed Repetitive Unit - Variable Number of Tandem Repeat (MIRU-VNTR) revealed indistinguishable bacterial genotypes among 26 patients in 14 (53.8%) HHC groups. Drug susceptibility testing (DST) revealed that 45 (71.4%) of the Mtb isolates were multidrug resistant. The major cluster composed of isolates from five HHCs and on three of these, whole genome sequencing (WGS) was performed confirming their high genetic similarity. These results pinpoint the need for improved vigilance for TB control in households in the city of Belém. When comparing WGS versus phenotypic resistance detection methods as DST and Minimum Inhibitory Concentration (MIC) our data suggest that depending on the colonies selection, results may present variation.

Strain diversity and relative transmission of Mycobacterium tuberculosis in south coastal Karnataka, India.

There are no published reports on the strain diversity and relative transmission of Mycobacterium tuberculosis isolates circulating in Karnataka State, India. To explore the strain diversity of M. tuberculosis isolates and their relative transmission in south coastal Karnataka using spoligotyping and mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTR) typing. A total of 108 clinical isolates of M. tuberculosis were processed for spoligotyping, and 12-locus MIRU-VNTR typing and cluster analysis was performed. Spoligotyping data of 108 isolates revealed 63 spoligotype patterns: 36 (80 isolates, 74.1%) patterns corresponded to spoligotype international types (SITs), whereas 27 (28 isolates, 25.9%) patterns were orphans. A further 57 (52.8%) isolates were clustered into 12 clusters; 51 (47.2%) isolates were unique. The largest spoligotype cluster comprised SIT 48 (L1.2.2), followed by SIT 1942 (L3) and SIT 11 (L1.1.2). Combined MIRU-VNTR typing and spoligotyping analysis further differentiated these 108 isolates into five clusters of two isolates each and 98 individual patterns. Combined use of spoligotyping and MIRU-VNTR typing is best suited for genotyping studies in this region. Very high genetic diversity was observed among the clinical isolates. Further elaborate studies are required for a better understanding of the genetic diversity and transmission dynamics of the strains circulating in this region.

The Epidemiological Significance and Temporal Stability of Mycobacterial Interspersed Repetitive Units-Variable Number of Tandem Repeats-Based Method Applied to Mycobacterium tuberculosis in China.

This study aimed to validate the epidemiological significance and temporal stability of Mycobacterial Interspersed Repetitive Units-Variable Number of Tandem Repeats (MIRU-VNTR) typing in a genetically and geographically diverse set of clinical isolates from patients diagnosed with pulmonary tuberculosis in China. Between 2010 and 2013, a total of 982 Mycobacterium tuberculosis isolates were collected from four population-based investigations in China. Apart from the currently applied 24-locus MIRU-VNTR, six additional hypervariable loci were analyzed in order to validate the MIRU-VNTR combinations in terms of their epidemiological links, clustering time span, and paired geographic distance. In vitro temporal stability was analyzed for both individual MIRU-VNTR loci, and for several combinations of loci. In the present study, four MIRU-VNTR combinations, including the hypervariable loci 3820, 3232, 2163a, and 4120, were evaluated. All of these combinations obtained a Hunter-Gaston discriminatory index (HGDI) value over 0.9900 with a reduced clustering proportion (from 32.0% to 25.6%). By comparing epidemiological links, clustering time span, and paired geographic distance, we found that the performances of the four MIRU-VNTR combinations were comparable to the insertion sequence 6110 restriction fragment length polymorphism (IS6110-RFLP), and significantly better than that of 24-locus MIRU-VNTR genotyping alone. The proportion of temporally stable loci ranged from 90.5% to 92.5% within the combined MIRU-VNTR genotyping, which is higher than IS6110-RFLP (85.4%). By adding four hypervariable loci to the standard 24-locus MIRU-VNTR genotyping, we obtained a high discriminatory power, stability and epidemiological significance. This algorithm could therefore be used to improve tuberculosis transmission surveillance and outbreak investigation in China.

Investigating a tuberculosis cluster among Filipino health care workers in a low-incidence country.

Nearly 8% of adult tuberculosis (TB) cases in England, Wales and Northern Ireland (EW&NI) occur among health care workers (HCWs), the majority of whom are from high TB incidence countries. To determine if a TB cluster containing multiple HCWs was due to nosocomial transmission. A cluster of TB cases notified in EW&NI from 2009 to 2014, with indistinguishable 24-locus mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTR) profiles, was identified through routine national cluster review. Cases were investigated to identify epidemiological links, and occupational health (OH) information was collected for HCW cases. To further discriminate strains, typing of eight additional loci was conducted. Of the 53 cases identified, 22 were HCWs. The majority (n = 43), including 21 HCWs, were born in the Philippines. Additional typing split the cluster into three subclusters and seven unique strains. No epidemiological links were identified beyond one household and a common residential area. HCWs in this cluster received no or inadequate OH assessment. The MIRU-VNTR profile of this cluster probably reflects common endemic strains circulating in the Philippines, with reactivation occurring in the UK. Furthermore, 32-locus typing showed that 24-locus MIRU-VNTR failed to distinguish strain diversity. The lack of OH assessment indicates that latent tuberculous infection could have been identified and treated, thereby preventing active cases from occurring.

High genetic diversity among Mycobacterium tuberculosis strains in Tehran, Iran

IntroductionTuberculosis (TB) still remains an important public health problem in Iran. The genotyping of Mycobacterium tuberculosis isolates is expected to lead to a better understanding of M. tuberculosis transmission in Tehran, the most populated city of Iran. Materials and MethodsA total of 2300 clinical specimens were obtained from TB suspected patients who were referred to a TB center in Tehran from Jan 2014 to Dec 2016. Identification was performed using both conventional and molecular methods. The presence of resistance to rifampicin was examined by the GeneXpert MTB/RIF. The standard 15-locus mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) typing method was applied to genotype of clinical isolates. ResultsOf 2300 specimens, 80 isolates were identified as M. tuberculosis by using biochemical and molecular tests. Of 80 M. tuberculosis isolates, 76 (95%) had unique genotypic profiles and 4 (5%) shared a profile with one or more other strains. Based on single loci variation (SLV) 4 clonal complexes were observed. NEW-1 was found to be the most predominant lineage (22.5%) followed by West African (1.25%), Central Asian (CAS)/Delhi (1.25%), Bovis (1.25%), H37Rv (1.25%) and multiple matches (1.25%). Loci MIRU10, MIRU26, MTUB21 and QUB26 were found as highly discriminative. No mutation was detected in the hotspot region of rifampicin by using GeneXpert MTB/RIF. ConclusionsOur study findings show that there was considerable genotypic diversity among M. tuberculosis isolates in Tehran. The 15-locus MIRU-VNTR showed high HGDI and could be used as a first-line genotyping method for epidemiological studies.

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.

Molecular epidemiology of Mycobacterium tuberculosis complex strains isolated from livestock and wild animals in Italy suggests the need for a different eradication strategy for bovine tuberculosis.

Bovine tuberculosis (bTB) is an important zoonosis, which has been re-emerging in different ecological scenarios. In Sicily, Italy, from 2004 to 2014, an anatomopathological survey for tuberculosis-like lesions both in farmed and wild animals was performed. The isolates were genotyped using spoligotyping and Mycobacterial Interspersed Repetitive Units-Variable Number of Tandem Repeats (MIRU-VNTR) techniques. High prevalence of lesions was observed for cattle (4%), pigs (4.9%) and wild boars (6.8%), and a total of 625 Mycobacterium bovis isolates were identified. Genotyping analysis showed the presence of 37 different spoligotypes including fifteen spoligotypes not present in other Italian regions and 266 MIRU-VNTR profiles. Spoligotype SB0120 exhibited the highest prevalence in cattle (50%) and pigs (56%) and the highest genetic variety with 126 different MIRU-VNTR profiles. The isolation of M.bovis in a farmer underlines the importance of M.bovis identification during the human TB diagnostic processes. This study supported the use of the genotyping analysis as a valuable tool for the evaluation of the epidemiological role of pigs and other domestic reservoirs such as goats and the role of wildlife in the maintenance of bTB infection.

Trends and differences in tuberculosis incidences and clustering among natives in Denmark, Sweden and Finland: comparison of native incidences and molecular epidemiology among three low-incidence countries

ObjectivesTo compare the epidemiology of tuberculosis (TB) in Denmark, Sweden and Finland, by focusing on the native population in order to identify epidemiologic differences and thus indirectly possible differences in TB control. MethodsTB incidence trends from 1990 through 2015 were compared among the countries. In addition, for the periods 2012–2013 and 2014–2015, genotyping data were compared. Genotyping was performed using the 24-locus mycobacterial interspersed repetitive unit–variable number of tandem repeat (MIRU-VNTR) method in Denmark and Sweden. For Finland, spoligotyping in conjunction with the 15-locus MIRU-VNTR method was used for 2012–2013 and translated into the 24-locus MIRU-VNTR when feasible, and for 2014–2015 only MIRU-VNTR was used. Both incidence trends and molecular epidemiology were assessed for native cases. ResultsThe average annual rate of change in TB incidence for native Danes was −2.4% vs. −6.1% and −6.9% for native Swedes and Finns respectively. In 2012–2013 Denmark had 52 native cases in the largest transmission chain vs. three cases in Sweden and ten in Finland, and during the same period the clustering rate for native Danes was 48.8% vs. 6.5% and 18.2% for native Swedes and Finns respectively. For 2014–2015, a similar pattern was seen. ConclusionsThe decline of TB among natives in Denmark is slower than for Sweden and Finland, and it seems Denmark has more active transmission among natives. The focused assessment on basic native TB epidemiology reveals striking differences in TB transmission among otherwise similar low-TB-incidence countries.

Molecular clustering of patients with diabetes and pulmonary tuberculosis: A systematic review and meta-analysis.

IntroductionMany studies have explored the relationship between diabetes mellitus (DM) and tuberculosis (TB) demonstrating increased risk of TB among patients with DM and poor prognosis of patients suffering from the association of DM/TB. Owing to a paucity of studies addressing this question, it remains unclear whether patients with DM and TB are more likely than TB patients without DM to be grouped into molecular clusters defined according to the genotype of the infecting Mycobacterium tuberculosis bacillus. That is, whether there is convincing molecular epidemiological evidence for TB transmission among DM patients. Objective: We performed a systematic review and meta-analysis to quantitatively evaluate the propensity for patients with DM and pulmonary TB (PTB) to cluster according to the genotype of the infecting M. tuberculosis bacillus.Materials and methodsWe conducted a systematic search in MEDLINE and LILACS from 1990 to June, 2016 with the following combinations of key words “tuberculosis AND transmission” OR “tuberculosis diabetes mellitus” OR “Mycobacterium tuberculosis molecular epidemiology” OR “RFLP-IS6110” OR “Spoligotyping” OR “MIRU-VNTR”. Studies were included if they met the following criteria: (i) studies based on populations from defined geographical areas; (ii) use of genotyping by IS6110- restriction fragment length polymorphism (RFLP) analysis and spoligotyping or mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTR) or other amplification methods to identify molecular clustering; (iii) genotyping and analysis of 50 or more cases of PTB; (iv) study duration of 11 months or more; (v) identification of quantitative risk factors for molecular clustering including DM; (vi) > 60% coverage of the study population; and (vii) patients with PTB confirmed bacteriologically. The exclusion criteria were: (i) Extrapulmonary TB; (ii) TB caused by nontuberculous mycobacteria; (iii) patients with PTB and HIV; (iv) pediatric PTB patients; (v) TB in closed environments (e.g. prisons, elderly homes, etc.); (vi) diabetes insipidus and (vii) outbreak reports. Hartung-Knapp-Sidik-Jonkman method was used to estimate the odds ratio (OR) of the association between DM with molecular clustering of cases with TB. In order to evaluate the degree of heterogeneity a statistical Q test was done. The publication bias was examined with Begg and Egger tests. Review Manager 5.3.5 CMA v.3 and Biostat and Software package R were used.ResultsSelection criteria were met by six articles which included 4076 patients with PTB of which 13% had DM. Twenty seven percent of the cases were clustered. The majority of cases (48%) were reported in a study in China with 31% clustering. The highest incidence of TB occurred in two studies from China. The global OR for molecular clustering was 0.84 (IC 95% 0.40–1.72). The heterogeneity between studies was moderate (I2 = 55%, p = 0.05), although there was no publication bias (Beggs test p = 0.353 and Eggers p = 0.429).ConclusionThere were very few studies meeting our selection criteria. The wide confidence interval indicates that there is not enough evidence to draw conclusions about the association. Clustering of patients with DM in TB transmission chains should be investigated in areas where both diseases are prevalent and focus on specific contexts.

Genetic diversity of Mycobacterium tuberculosis and transmission associated with first-line drug resistance: a first analysis in Jalisco, Mexico

The objectives of this study were to analyse the frequency of gene mutations associated with antitubercular drug resistance in clinical samples from the population of Jalisco State (Mexico) and to evaluate the genetic variability of Mycobacterium tuberculosis and multidrug-resistant (MDR) M. tuberculosis strains to describe the frequency of various families. Clinical isolates of M. tuberculosis obtained from Jalisco State were analysed. Isolates were subjected to drug susceptibility testing, and mutations were characterised by sequencing, followed by genotyping using spoligotyping and mycobacterial interspersed repetitive units-variable-number of tandem repeats (MIRU-VNTR). Moreover, the prevalence of mutations was analysed by phylogenetic lineages. Resistant strains were analysed by sequencing of katG, inhA and rpoB genes to determine the presence of mutations associated with isoniazid and rifampicin resistance. In MDR, monoresistant and polyresistant isolates, mutations were found in 17 (54.84%) of 31 strains. Spoligotyping identified six different strain lineages [T1 (25.40%), H3 (7.94%), MANU (4.76%), X1 (3.17%), EAI5 (1.59%) and LAM1 (1.59%)], with the remaining strains identified as orphans. In additional tree-based identification, a dendrogram of spoligotype patterns generated five different similarity clusters. When combining 24-loci MIRU-VNTR and spoligotyping approaches, the results shows that there is no cluster formation, indicating low transmission of the samples. This study using spoligotyping and MIRU-VNTR showed that the analysed strains were not related to each other since no two identical strains were found. Families with the highest prevalence in the study were orphans followed by T family.

Evaluation of 24-locus MIRU-VNTR genotyping in Mycobacterium tuberculosis cluster investigations in four jurisdictions in the United States, 2006–2010

The U.S. Centers for Disease Control and Prevention (CDC) uses a combination of spacer oligonucleotide typing (spoligotyping) and mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) analyses as part of the National TB Genotyping Service (NTGS). The NTGS expansion from 12-locus MIRU-VNTR (MIRU12) to 24-locus MIRU-VNTR (MIRU24) in 2009 enhanced the ability to discriminate Mycobacterium tuberculosis strains. In the current study, we investigated the MIRU24 concordance among epidemiologic-linked tuberculosis (TB) patients in four U.S. health jurisdictions. We also evaluated the programmatic benefits of combining MIRU24 and spoligotyping with epidemiologic evidence in identifying potential recent TB transmission. We examined 342 TB patients in 42 spoligotype/MIRU12 (PCRType) clusters (equivalent to 46 spoligotype/MIRU24 [GENType] clusters) to identify epidemiologic links among cases. GENType clusters, when compared to PCRType clusters, had 12 times higher odds of epidemiologic links being identified if patients were younger than 25 years and 3 times higher odds if patients resided in the same zip code, or had HIV infection. Sixty (18%) fewer PCRType-clustered patients would need investigations if clusters are defined using GENType instead of PCRType. An important advantage of defining clusters by MIRU24 is resource savings related to the reduced number of clustered cases needing investigation.

Pathology and genetic findings in a rare case of Mycobacterium caprae infection in a sow

Bovine tuberculosis, a reemerging zoonosis in diverse ecological scenarios, has been reported in the autochthonous Nebrodi black pig breed population used for meat production in Italy. During a routine abattoir inspection in 2013, 24 of 299 carcasses (8%) of Nebrodi black pigs presented tuberculosis-like lesions at pathologic examination. Mycobacterium bovis was isolated from 23 animals and M. caprae from a 3-year-old sow. The sow showed severe diffuse lesions involving the visceral organs, right coxofemoral joint, and mammary glands. Isolation of M. caprae from mammary glands is uncommon, with only one other case involving a sow reported so far; however, Mycobacteria infection of the mammary glands may be transmitted from lactating sows to piglets, contributing to the spread and maintenance of bovine tuberculosis in swine. Genotyping analysis showed M. caprae spoligotype SB0866 and profile 4,1,5,4,4,11,4,2,4,3,8,7 MIRU-VNTR (mycobacterial interspersed repetitive units-variable number of tandem repeats). The worldwide prevalence of this spoligotype is very low. The finding of severe, diffuse tuberculous lesions strongly suggests that Nebrodi black pigs are susceptible for Mycobacterium spp. and that they might act as a distributor for these microorganisms. Since natural ecosystems with multiple contacts among different livestock species and wild animals are very common in Mediterranean regions, current surveillance and eradication plans for bovine tuberculosis will need to be extended to other potential reservoir species in regions where extensive and traditional breeding systems are operated.

Recurrence of tuberculosis in a low-incidence setting without directly observed treatment: Victoria, Australia, 2002-2014.

Victoria, Australia, is an industrialised setting with low tuberculosis (TB) incidence and universal health care. Individually tailored adherence support for self-administered daily anti-tuberculosis treatment is provided. Directly observed treatment (DOT) is very rarely used. To review the rate of recurrent TB in Victoria between 2002 and 2014. This was a retrospective cohort study. All recurrent episodes of TB were reviewed and 24-locus MIRU-VNTR (mycobacterial interspersed repetitive units-variable number of tandem repeats) molecular typing was used where possible to determine the likelihood of relapse or reinfection. Of 4766 notifications, 32 (0.7%) were recurrent episodes. Of 20 episodes that occurred in patients who had previously completed treatment, 11 were culture-positive (0.4% of 3012 culture-positive episodes): 9 were likely relapses (distinguishable at no more than one of 24 loci) and two were likely reinfections, giving a TB relapse rate among culture-positive episodes of 52.5/100 000 person-years (mean time to study end per patient of 5.7 years). The median time until relapse was 18 months (interquartile range 12-30). The low rate of relapse in our setting demonstrates that individually tailored adherence support for self-administered anti-tuberculosis treatment can achieve excellent treatment outcomes.

Comparative study of genotypes of Mycobacterium tuberculosis from a Northern Indian setting with strains reported from other parts of India and neighboring countries

This study was carried out to characterize Mycobacterium tuberculosis population in Ghatampur, Kanpur, North India, by spoligotyping and Mycobacterial Interspersed Repetitive Units-Variable Number of Tandem Repeats (MIRU-VNTRs) typing. A total of 335 isolates were genotyped by spoligotyping and Central Asian (CAS) sub-lineage was the most prevalent, comprising 59.1% of all isolates. Other lineages were: East-African Indian (EAI) (19.10%), T (5.07%), Beijing (3.28%), Manu (2.98%), X (2.68%), S (0.89%), H3 (0.59%), Ural (0.59%), LAM 9 (0.29%) and unknown (5.37%). This data was compared with 8444 clinical isolates from other parts of India and neighboring countries. Thanks to interrogation of the SITVIT2 database, which shows that China is unique in having a predominance of Beijing lineage; Iran in having an almost equal proportion of Ural and CAS lineages; while the rest of the Middle-East and Indian subcontinent shows a gradient of CAS lineage predominating in the north of tropic of cancer, and the ancestral EAI lineage in South India and South-East Asia. Additionally, 12 loci MIRU-VNTR typing efficiently discriminated 13 spoligotype-defined clusters into 92 patterns; 53 isolates showed >70% homology. It was observed that Beijing lineage strains were more frequently associated with MDR strains (p-value = 0.001). A multi-step application of combination of spoligotyping and MIRU-VNTR typing for analyzing the molecular epidemiology of TB may provide a better means of fingerprinting and studying transmission dynamics.

Molecular analysis of pyrazinamide resistance in Mycobacterium tuberculosis in Vietnam highlights the high rate of pyrazinamide resistance-associated mutations in clinical isolates

Pyrazinamide (PZA) is a key antibiotic in current anti-tuberculosis regimens. Although the WHO has stressed the urgent need to obtain data on PZA resistance, in high tuberculosis burden countries, little is known about the level of PZA resistance, the genetic basis of such resistance or its link with Mycobacterium tuberculosis families. In this context, this study assessed PZA resistance through the molecular analysis of 260 Vietnamese M. tuberculosis isolates. First-line drug susceptibility testing, pncA gene sequencing, spoligotyping and mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) typing were performed. Overall, the pncA mutation frequency was 38.1% (99 out of 260 isolates) but was higher than 72% (89 out of 123 isolates) in multidrug and quadruple-drug resistant isolates. Many different pncA mutations (71 types) were detected, of which 55 have been previously described and 50 were linked to PZA resistance. Among the 16 novel mutations, 14 are likely to be linked to PZA resistance because of their mutation types or codon positions. Genotype analysis revealed that PZA resistance can emerge in any M. tuberculosis cluster or family, although the mutation frequency was the highest in Beijing family isolates (47.7%, 62 out of 130 isolates). These data highlight the high rate of PZA resistance-associated mutations in M. tuberculosis clinical isolates in Vietnam and bring into question the use of PZA for current and future treatment regimens of multidrug-resistant tuberculosis without PZA resistance testing.Emerging Microbes & Infections (2017) 6, e86; doi:10.1038/emi.2017.73; published online 11 October 2017