Violet N. Chihota

Clonal Expansion of a Globally Disseminated Lineage of Mycobacterium tuberculosis with Low IS6110 Copy Numbers

ABSTRACT Knowledge of the clonal expansion of Mycobacterium tuberculosis and accurate identification of predominant evolutionary lineages in this species remain limited, especially with regard to low-IS6110-copy-number strains. In this study, 170 M. tuberculosis isolates with ≤6 IS6110 insertions identified in Cape Town, South Africa, were characterized by principal genetic grouping, restriction fragment length polymorphism analysis, spoligotyping, IS6110 insertion site mapping, and variable-number tandem repeat (VNTR) typing. These analyses indicated that all but one of the isolates analyzed were members of principal genetic group 2 and of the same low-IS6110-copy-number lineage. The remaining isolate was a member of principal genetic group 1 and a different low-IS6110-copy-number lineage. Phylogenetic reconstruction suggests clonal expansion through sequential acquisition of additional IS6110 copies, expansion and contraction of VNTR sequences, and the deletion of specific direct-variable-repeat sequences. Furthermore, comparison of the genotypic data of 91 representative low-IS6110-copy-number isolates from Cape Town, other southern African regions, Europe, and the United States suggests that certain low-IS6110-copy-number strain spoligotypes and IS6110 fingerprints were acquired in the distant past. These clones have subsequently become widely disseminated and now play an important role in the global tuberculosis epidemic.

Population Structure of Multi- and Extensively Drug-Resistant Mycobacterium tuberculosis Strains in South Africa

ABSTRACT Genotyping of multidrug-resistant (MDR) Mycobacterium tuberculosis strains isolated from tuberculosis (TB) patients in four South African provinces (Western Cape, Eastern Cape, KwaZulu-Natal, and Gauteng) revealed a distinct population structure of the MDR strains in all four regions, despite the evidence of substantial human migration between these settings. In all analyzed provinces, a negative correlation between strain diversity and an increasing level of drug resistance (from MDR-TB to extensively drug-resistant TB [XDR-TB]) was observed. Strains predominating in XDR-TB in the Western and Eastern Cape and KwaZulu-Natal Provinces were strongly associated with harboring an inhA promoter mutation, potentially suggesting a role of these mutations in XDR-TB development in South Africa. Approximately 50% of XDR-TB cases detected in the Western Cape were due to strains probably originating from the Eastern Cape. This situation may illustrate how failure of efficient health care delivery in one setting can burden health clinics in other areas.

Emergence and treatment of multidrug resistant (MDR) and extensively drug-resistant (XDR) tuberculosis in South Africa

Drug resistant tuberculosis (TB) has reached alarming proportions in South Africa, draining valuable resources that are needed to fight drug susceptible TB. It is currently estimated that 9.6% of all TB cases have multi-drug resistant (MDR)-TB, thereby ranking South Africa as one of the highest MDR-TB burden countries in the world. Molecular epidemiological studies have demonstrated the complexity of the epidemic and have clearly shown that the epidemic is driven by transmission as a consequence of low cases detection and diagnostic delay. The latter has in turn fueled the amplification of drug resistance, ultimately leading to the emergence of extensively drug resistant (XDR)-TB. Despite the introduction of new drugs to combat this scourge, culture conversion rates for XDR-TB remain below 20%. Failure to achieve cure may be explained from DNA sequencing results which have demonstrated mutations in 7 genes encoding resistance to at least 8 anti-TB drugs. This review shows how molecular epidemiology has provided novel insights into the MDR-TB epidemic in South Africa and thereby has highlighted the challenges that need to be addressed regarding the diagnosis and treatment of MDR-TB. An important step towards for curbing this epidemic will be collaboration between clinicians, laboratories and researchers to establish scientific knowledge and medical expertise to more efficiently guide public health policy.

Tuberculosis outcomes and drug susceptibility in individuals exposed to isoniazid preventive therapy in a high HIV prevalence setting.

Objective:Despite World Health Organization recommendations, concerns about promoting resistance have impeded implementation of isoniazid preventive therapy (IPT) for tuberculosis (TB). We describe characteristics of TB in individuals previously exposed to IPT as part of ‘Thibela TB’, a cluster-randomized trial of community-wide IPT in gold miners in South Africa. Design:Case series including participants who were dispensed IPT, attended at least one follow-up visit and were subsequently treated for TB. Methods:TB episodes were detected through surveillance and through follow-up if IPT was stopped early. Drug susceptibility data were compared with TB episodes detected through surveillance in control clusters (where IPT use was minimal) and a laboratory substudy of mycobacterial sputum culture from TB suspects in control clusters. Results:Among 126 eligible individuals (125 men, median age 43 years), median time from starting IPT to TB treatment was 316 days (interquartile range 174–491). Ninety-four of the 126 (75%) were first episodes. Eighty-nine of 103 (86%) tested HIV-infected, with the median CD4 cell count of 196 cells/μl (n = 51). Sixty-four of 108 (59%) with known treatment outcomes were cured or completed treatment. Among 71 isolates with drug susceptibility results available, 12.1% [95% confidence interval (CI) 5.0–23.3] and 7.7% (95% CI 0.2–36.0) from first and retreatment episodes, respectively, had isoniazid resistance, compared with 6.0% (95% CI 3.1–10.2) and 18.7% (95% CI 10.6–29.3) in control clusters and 11.8% (95% CI 8.2–16.3) among first TB episodes in the laboratory substudy. Conclusion:TB after recent IPT has prevalence of drug resistance similar to background and treatment outcomes typical of this setting. These data support wider implementation of IPT.

Performance Characteristics of the Cepheid Xpert MTB/RIF Test in a Tuberculosis Prevalence Survey

Background Xpert MTB/RIF (“Xpert”) is a molecular test for detection of Mycobacterium tuberculosis (MTB) in sputum. Performance characteristics have been established for its use during passive tuberculosis (TB) case detection in symptomatic TB suspects, but Xpert performance has not been assessed in other settings. Objectives were to determine Xpert performance and costs in the context of a TB prevalence survey. Methodology/Principal Findings This was a diagnostic sub-study of a TB prevalence survey conducted in gold mining companies in South Africa. Sputa (one per participant) were tested using smear microscopy, liquid culture (reference comparator), and Xpert. Costs were collected using an ingredients approach and analyzed using a public health program perspective. 6893 participants provided a sputum specimen. 187/6893 (2.7%) were positive for MTB in culture, 144/6893 (2.1%) were positive for MTB by Xpert, and 91/6893 (1.3%) were positive for acid fast bacilli by microsocopy. Sensitivity, specificity, positive predictive value, and negative predictive value for detection of MTB by Xpert were 62.6% (95% confidence interval [CI] 55.2, 69.5), 99.6% (99.4, 99.7), 81.3% (73.9, 87.3), and 98.9 (98.6, 98.8); agreement between Xpert and culture was 98.5% (98.2, 98.8). Sensitivity of microscopy was 17.6% (12.5, 23.9). When individuals with a history of TB treatment were excluded from the analysis, Xpert specificity was 99.8 (99.7, 99.9) and PPV was 90.6 (83.3, 95.4) for detection of MTB. For the testing scenario of 7000 specimens with 2.7% of specimens culture positive for MTB, costs were

Comparison of laboratory costs of rapid molecular tests and conventional diagnostics for detection of tuberculosis and drug-resistant tuberculosis in South Africa

165,690 for Xpert and

Programmatically Selected Multidrug-Resistant Strains Drive the Emergence of Extensively Drug-Resistant Tuberculosis in South Africa

115,360 for the package of microscopy plus culture. Conclusion In the context of a TB prevalence survey, the Xpert diagnostic yield was substantially higher than that of microscopy yet lower than that of liquid culture. Xpert may be useful as a sole test for TB case detection in prevalence surveys, particularly in settings lacking capacity for liquid culture.

Genotype MTBDRplus for Direct Detection of Mycobacterium tuberculosis and Drug Resistance in Strains from Gold Miners in South Africa

BackgroundThe World Health Organization has endorsed the use of molecular methods for the detection of TB and drug-resistant TB as a rapid alternative to culture-based systems. In South Africa, the Xpert MTB/Rif assay and the GenoType MTBDRplus have been implemented into reference laboratories for diagnosis of TB and drug-resistance, but their costs have not been fully elucidated.MethodsWe conducted a detailed reference laboratory cost analysis of new rapid molecular assays (Xpert and MTBDRplus) for tuberculosis testing and drug-resistance testing in South Africa, and compared with the costs of conventional approaches involving sputum microscopy, liquid mycobacterial culture, and phenotypic drug sensitivity testing.ResultsFrom a laboratory perspective, Xpert MTB/RIF cost

Symptom and chest radiographic screening for infectious tuberculosis prior to starting isoniazid preventive therapy: yield and proportion missed at screening.

14.93/sample and the MTBDRplus line probe assay cost

“Proof-Of-Concept” Evaluation of an Automated Sputum Smear Microscopy System for Tuberculosis Diagnosis

23.46/sample, compared to