A laboratory-based study to identify and speciate non-tuberculous mycobacteria isolated from specimens submitted to a central tuberculosis laboratory from throughout KwaZulu-Natal Province, South Africa

Background. Non-tuberculous mycobacteria (NTM) are important environmental pathogens capable of causing a spectrum of infection. The different species exhibit varied geographical prevalence worldwide. Identification of the infecting organism may be helpful in determining the clinical significance of the isolate.Objective. To describe the spectrum of NTM isolated from clinical specimens received at the National Health Laboratory Service central tuberculosis laboratory in KwaZulu-Natal Province, South Africa.Method. In a laboratory-based prospective study, 200 suspected NTM were randomly selected over a period of 1 year and identified to species level using a commercially available DNA strip assay (GenoType Mycobacterium, CM/AS; Hain Lifescience, Germany).Results. Of the 200 suspected NTM, 133 (66.5%) were confirmed to be NTM by the molecular test. The most frequently isolated NTM species were Mycobacterium intracellulare (45.9%), M. avium subspecies (11.3%), M. gordonae (6.0%) and M. kansasii (4.5%).Conclusion. It is important for laboratories to document the local spectrum of NTM because of the geographical variation in the different NTM species isolated. Although molecular tests for identifying NTM are relatively expensive, they have the advantage of providing rapid and accurate identification of the various NTM species

Diagnostic accuracy of quantitative PCR (Xpert MTB/RIF) for tuberculous meningitis in a high burden setting: a prospective study

Background: Tuberculous meningitis (TBM) is difficult to diagnose promptly. The utility of the Xpert MTB/RIF test for the diagnosis of TBM remains unclear, and the effect of host- and sample-related factors on test performance is unknown. This study sought to evaluate the sensitivity and specificity of Xpert MTB/RIF for the diagnosis of TBM. Methods and Findings: 235 South-African patients with a meningeal-like illness were categorised as having definite (culture or Amplicor PCR positive), probable (anti-TBM treatment initiated but microbiological confirmation lacking), or non-TBM. Xpert MTB/RIF accuracy was evaluated using 1 ml of uncentrifuged and, when available, 3 ml of centrifuged cerebrospinal fluid (CSF). To evaluate the incremental value of MTB/RIF over a clinically based diagnosis, test accuracy was compared to a clinical score (CS) derived using basic clinical and laboratory information. Of 204 evaluable patients (of whom 87% were HIV-infected), 59 had definite TBM, 64 probable TBM, and 81 non-TBM. Overall sensitivity and specificity (95% CI) were 62% (48%–75%) and 95% (87%–99%), respectively. The sensitivity of Xpert MTB/RIF was significantly better than that of smear microscopy (62% versus 12%; p = 0.001) and significantly better than that of the CS (62% versus 30%; p = 0.001; C statistic 85% [79%–92%]). Xpert MTB/RIF sensitivity was higher when centrifuged versus uncentrifuged samples were used (82% [62%–94%] versus 47% [31%–61%]; p = 0.004). The combination of CS and Xpert MTB/RIF (Xpert MTB/RIF performed if CS<8) performed as well as Xpert MTB/RIF alone but with a ∼10% reduction in test usage. This overall pattern of results remained unchanged when the definite and probable TBM groups were combined. Xpert MTB/RIF was not useful in identifying TBM among HIV-uninfected individuals, although the sample was small. There was no evidence of PCR inhibition, and the limit of detection was ∼80 colony forming units per millilitre. Study limitations included a predominantly HIV-infected cohort and the limited number of culture-positive CSF samples. Conclusions: Xpert MTB/RIF may be a good rule-in test for the diagnosis of TBM in HIV-infected individuals from a tuberculosis-endemic setting, particularly when a centrifuged CSF pellet is used. Further studies are required to confirm these findings in different settings

Rifampicin mono-resistance in mycobacterium tuberculosis in KwaZulu-Natal, South Africa : A significant phenomenon in a high prevalence TB-HIV region.

This laboratory based study was conducted at NHLS TB Laboratory, Durban, which is the reference laboratory for culture and susceptibility testing in KwaZulu-Natal. We retrospectively determined, for the period 2007 to 2009, the proportion of RMR amongst Mycobacterium tuberculosis (MTB) isolates, that were tested for both RIF and INH, using the gold standard of culture based phenotypic drug susceptibility testing (DST). Gender and age were also analysed to identify possible risk factors for RMR.Setting: The dual epidemics of HIV-TB including MDR-TB are major contributors to high morbidity and mortality rates in South Africa. Rifampicin (RIF) resistance is regarded as a proxy for MDR-TB. Currently available molecular assays have the advantage of rapidly detecting resistant strains of MTB, but the GeneXpert does not detect isoniazid (INH) resistance and the GenoTypeMTBDRplus(LPA) assay may underestimate resistance to INH. Increasing proportions of rifampicin monoresistance resistance (RMR) have recently been reported from South Africa and other countries. Objective: This laboratory based study was conducted at NHLS TB Laboratory, Durban, which is the reference laboratory for culture and susceptibility testing in KwaZulu-Natal. We retrospectively determined, for the period 2007 to 2009, the proportion of RMR amongst Mycobacterium tuberculosis (MTB) isolates, that were tested for both RIF and INH, using the gold standard of culture based phenotypic drug susceptibility testing (DST). Gender and age were also analysed to identify possible risk factors for RMR. Design: MTB culture positive sputum samples from 16,748 patients were analysed for susceptibility to RIF and INH during the period 2007 to 2009. RMR was defined as MTB resistant to RIF and susceptible to INH. For the purposes of this study, only the first specimen from each patient was included in the analysis. Results: RMR was observed throughout the study period. The proportion of RMR varied from a low of 7.3% to a high of 10.0% [overall 8.8%]. Overall, males had a 42% increased odds of being RMR as compared to females. In comparison to the 50 plus age group, RMR was 37% more likely to occur in the 25–29 year age category. Conclusion: We report higher proportions of RMR ranging from 7.3% to 10% [overall 8.8%] than previously reported in the literature. To avoid misclassification of RMR, detected by the GeneXpert, as MDR-TB, culture based phenotypic DST must be performed on a second specimen, as recommended by the SA NDOH TB guidelines as well as WHO. We suggest that two sputum samples should be obtained at the first visit. The second sputum sample should be stored at 4uC. The latter sample is then readily available for performing additional DST (phenotypic or genotypic) for 2nd lines drugs, resulting in a decreased waiting period for DST results to become available

Blood cultures for the diagnosis of multidrug-resistant and extensively drug-resistant tuberculosis among HIV-infected patients from rural South Africa: a cross-sectional study

<p>Abstract</p> <p>Background</p> <p>The yield of mycobacterial blood cultures for multidrug-resistant (MDR) and extensively drug-resistant tuberculosis (XDR-TB) among drug-resistant TB suspects has not been described.</p> <p>Methods</p> <p>We performed a retrospective, cross-sectional analysis to determine the yield of mycobacterial blood cultures for MDR-TB and XDR-TB among patients suspected of drug-resistant TB from rural South Africa. Secondary outcomes included risk factors of <it>Mycobacterium tuberculosis </it>bacteremia and the additive yield of mycobacterial blood cultures compared to sputum culture.</p> <p>Results</p> <p>From 9/1/2006 to 12/31/2008, 130 patients suspected of drug-resistant TB were evaluated with mycobacterial blood culture. Each patient had a single mycobacterial blood culture with 41 (32%) positive for <it>M. tuberculosis</it>, of which 20 (49%) were XDR-TB and 8 (20%) were MDR-TB. One hundred fourteen (88%) patients were known to be HIV-infected. Patients on antiretroviral therapy were significantly less likely to have a positive blood culture for <it>M. tuberculosis </it>(p = 0.002). The diagnosis of MDR or XDR-TB was made by blood culture alone in 12 patients.</p> <p>Conclusions</p> <p>Mycobacterial blood cultures provided an additive yield for diagnosis of drug-resistant TB in patients with HIV from rural South Africa. The use of mycobacterial blood cultures should be considered in all patients suspected of drug-resistant TB in similar settings.</p

Antimycobacterial activity of aqueous and methanol extracts of nine plants against Mycobacterium bacteria

Purpose: The present study was done to evaluate the antimycobacterial activity of aqueous and methanol extracts of nine plants viz., Buddleja saligna, Carpobrotus dimidiatus, Capparis tomentosa, Dichrostachys cinerea, Ekebergia capensis, Ficus sur, Gunnera perpensa, Leonotis leonurus and Tetradenia riparia in South Africa. Methods: Aqueous and methanol extracts of the leaves the plants were tested in vitro for their activity against Mycobacterium smegmatis, Mycobacterium tuberculosis H37Rv (ATCC 25177) and three well-characterized clinical isolates of MDR and XDR-TB isolates using the agar incorporation method. The minimum inhibitory concentration (MIC) of each of the active plant extract was determined using the broth microdilution method. Cytotoxic effect was evaluated against the mouse BALB/C monocyte macrophage (J774.2) and peripheral blood mononuclear cells (PBMCs) whole the toxicity screening was done using the brine shrimp lethality assay. Composition of each of the plants was determined using thin layer chromatography while qualitative analysis of antimycobacterial compounds was done using TLC-Bioautography Results: The methanol extracts of B. saligna, C. tormentosa and C. dimidiatus; and aqueous extracts of G. perpensa and T. riparia possessed significant activity against M. smegmatis, M. tuberculosis H37Rv (ATCC 25177) and the three well-characterized clinical isolates of MDR and XDR-TB. Except for a high concentration of G. perpensa none of the other plants which possessed antimycobacterial activity showed any toxic or cytotoxic activity. Conclusion: Our findings show that B. saligna, C. tormentosa, C. dimidiatus, G. perpensa, and T. riparia have antimycobacterial activity. Further studies would aim at isolation and identification of the active compounds from the plants extracts which had positive antimycobacterial activity

Adjusted logistic regression of being Rifampicin mono-resistant, assessing the effect of age, overall and stratified by gender.

*<p>Adjusting for gender and quarter.</p>**<p>Adjusting for quarter.</p

Rifampicin mono-resistance over quarters from 2007 to 2009 overall and by gender.

*<p>A total of 431 patients had missing gender data.</p

Cerebrospinal T-Cell Responses Aid in the Diagnosis of Tuberculous Meningitis in a Human Immunodeficiency Virus– and Tuberculosis-Endemic Population

Rationale: Current tools for the rapid diagnosis of tuberculous meningitis (TBM) are suboptimal. We evaluated the clinical utility of a quantitative RD-1 IFN-γ T-cell enzyme-linked immunospot (ELISPOT) assay (T-SPOT.TB), using cerebrospinal fluid cells for the rapid immunodiagnosis of TBM

Extensively Drug-Resistant Mycobacterium tuberculosis from Aspirates, Rural South Africa

The yield from aspirating lymph nodes and pleural fluid for diagnosing extensively drug-resistant (XDR) tuberculosis is unknown. Mycobacterium tuberculosis was cultured from lymph node or pleural fluid aspirates of 21 patients; 7 (33%) cultures grew XDR M. tuberculosis. Additive diagnostic yield for XDR M. tuberculosis was found in parallel culture of sputum and fluid aspirate