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Tuberculosis drug resistance testing by molecular methods: opportunities and challenges in resource limited settings
One of the greatest threats to global tuberculosis (TB) control is the growing prevalence of drug-resistant bacilli. Correctly diagnosing drug-resistant TB patients is more problematic in resource-limited settings as there is no or limited infrastructure for drug susceptibility testing (DST) of TB bacilli. The conventional phenotypic DST method for TB takes weeks before declaring the results and initiating proper anti-TB treatment. Molecular DST offers advantages over the phenotypic methods mainly because of the short turn around time. This review summarizes the different molecular DST methods for TB and discusses challenges and opportunities in implementing them in resource-limited settings
Longlasting insecticidal nets for prevention of Leishmania donovani infection in India and Nepal: paired cluster randomised trial
OBJECTIVE: To test the effectiveness of large scale distribution of longlasting nets treated with insecticide in reducing the incidence of visceral leishmaniasis in India and Nepal. DESIGN: Paired cluster randomised controlled trial designed to detect a 50% reduction in incidence of Leishmania donovani infection. SETTING: Villages in Muzaffarpur district in India and Saptari, Sunsari, and Morang districts in Nepal. PARTICIPANTS: 13 intervention and 13 control clusters. 12 691 people were included in the analysis of the main outcome (infection), and 19 810 were enrolled for the secondary (disease) end point. INTERVENTION: Longlasting insecticidal nets (treated with deltamethrin) were distributed in the intervention clusters in December 2006. MAIN OUTCOME MEASURES: Infection was determined by direct agglutination test at 12 and 24 months after the intervention in those who had negative results (titre <1:1600) at baseline. The effect estimate was computed as the geometric mean of the risk ratios for seroconversion for each cluster pair (net/no net), with its 95% confidence interval. Formal tests of effect of no intervention were obtained with a paired t test. RESULTS: There was no significant difference in the risk of seroconversion over 24 months in intervention (5.4%; 347/6372) compared with control (5.5%; 345/6319 people) clusters (risk ratio 0.90, 95% confidence interval 0.49 to 1.65) nor in the risk of clinical visceral leishmaniasis (0.99, 0.46 to 1.40). Adjustment for covariates did not alter these conclusions. CONCLUSIONS: There is no evidence that large scale distribution of longlasting insecticidal nets provides additional protection against visceral leishmaniasis compared with existing control practice in the Indian subcontinent. The observed effect was small and not significant, though the confidence intervals did not exclude a 50% change in either direction. Trial registration Clinical Trials NCT 2005-015374
Outcomes of antiretroviral treatment program in Ethiopia: retention of patients in care is a major challenge and varies across health facilities
BACKGROUND: Many resource-limited countries are scaling up antiretroviral treatment (ART) towards universal access. However, there are few studies which evaluated outcomes of ART programs in these countries. In addition, these studies generally include a limited number of facilities and patients creating a clear need for studies with a wide range of facilities and large numbers of patients. In this study, we intended to evaluate the outcomes of the ART services in 55 health facilities in Ethiopia. METHODS: A retrospective longitudinal study was conducted to determine levels of patient retention in care, CD4 count and shift to second-line ART regimen in 30 hospitals and 25 health centers selected as sentinel sites for monitoring the outcomes of ART program in the country. The outcomes were determined at baseline, after 6, 12 and 24 months on ART. Data was collected from routine patient registers and charts, and entered and analyzed using EPI-Info statistical software. RESULTS: Health facilities were able to retain 29,893 (80%), 20,079 (74%) and 5,069 (68%) of their patients after 6, 12 and 24 months on ART, respectively. Retention rates vary across health facilities, ranging from 51% to 85% after 24 months on ART. Mortality was 5%, 6% and 8% after 6, 12 and 24 months on ART. More than 79% of patients with available CD4-cell counts had a baseline CD4-cell counts less than 200 cells per micro-liter of blood. The median CD4-cell counts (based on patients who were retained after 24 months on ART) increased from 125 (inter-quartile (IQ), 68-189) at baseline to 242 (IQ, 161-343), 269 (IQ, 185-380) and 316 (IQ, 226-445) cells per micro-liter after 6, 12, and 24 months on ART, respectively. The transition to second-line ART remained very low, 0.33%, 0.58% and 2.13% after 6, 12 and 24 months on ART. Discussion and conclusion: The outcomes of the ART services in the 55 health facilities in Ethiopia are similar to those in other countries. Retention of patients in care is a major challenge and varies across health facilities with high, medium and low retention rates. We therefore recommend further studies to understand the organization of care in health facilities with high, medium and low retention rates. It is also imperative that early initiation of patients on ART is taken seriously as more than 79% of the patients had baseline CD4-cell counts less than 200 cells per micro-liter of blood. Finally, we recommend that the shift to second-line ART might be too low and warrants close monitoring
Validation of a clinical-radiographic score to assess the probability of pulmonary tuberculosis in suspect patients with negative sputum smears
An alternative, sensitive method to detect Helicobacter pylori DNA in feces
Background: Despite the high sensitivity and specificity of PCR, detection of Helicobacter pylori DNA in feces is still challenging. Fecal samples contain inhibitory molecules that can prevent amplification of the target DNA. Even by using specific DNA extraction kits for stools, monitoring of infection by analyzing stool samples remains problematic and endorses the need for improved diagnostic methods. Materials and Methods: The newly proposed method uses selective hybridization of target DNA with biotin-labeled probes, followed by DNA isolation with streptavidin-coated magnetic beads. After three washing steps, the purified DNA can be amplified immediately using conventional or quantitative PCR. In order to test this technique on biological samples, Mongolian gerbils were infected with H. pylori ATCC 43504 and fecal samples were analyzed on days 1, 4, and 10 post infection. Results: A detection limit of one bacterial cell per 100 mg stool sample was established, but only after removal of the magnetic beads from the target DNA by heating. This resulted in a 10-fold increase of sensitivity compared to a commercially available stool DNA extraction kit. Analysis of fecal samples from infected gerbils demonstrated the presence of H. pylori DNA on each time point, while the uninfected animal remained negative. Conclusions: The proposed technique allows detection of very low quantities of H. pylori DNA in biological samples. In laboratory animal models, detailed monitoring of infection and complete clearance of infection can be demonstrated thanks to the low detection limit
Comparative gene expression analysis throughout the life cycle of Leishmania braziliensis: diversity of expression profiles among clinical isolates
BACKGROUND: Most of the Leishmania genome is reported to be constitutively expressed during the life cycle of the parasite, with a few regulated genes. Inter-species comparative transcriptomics evidenced a low number of species-specific differences related to differentially distributed genes or the differential regulation of conserved genes. It is of uppermost importance to ensure that the observed differences are indeed species-specific and not simply specific of the strains selected for representing the species. The relevance of this concern is illustrated by current study. METHODOLOGY/PRINCIPAL FINDINGS: We selected 5 clinical isolates of L. braziliensis characterized by their diversity of clinical and in vitro phenotypes. Real-time quantitative PCR was performed on promastigote and amastigote life stages to assess gene expression profiles at seven time points covering the whole life cycle. We tested 12 genes encoding proteins with roles in transport, thiol-based redox metabolism, cellular reduction, RNA poly(A)-tail metabolism, cytoskeleton function and ribosomal function. The general trend of expression profiles showed that regulation of gene expression essentially occurs around the stationary phase of promastigotes. However, the genes involved in this phenomenon appeared to vary significantly among the isolates considered. CONCLUSION/SIGNIFICANCE: Our results clearly illustrate the unique character of each isolate in terms of gene expression dynamics. Results obtained on an individual strain are not necessarily representative of a given species. Therefore, extreme care should be taken when comparing the profiles of different species and extrapolating functional differences between them
Evaluation of the TB Ag MPT64 rapid test for the identification of Mycobacterium tuberculosis complex
Tuberculosis knowledge, attitudes and health-seeking behaviour in rural Uganda
OBJECTIVES: To assess tuberculosis (TB) knowledge, attitudes and health-seeking behaviour to inform the design of communication and social mobilisation interventions. SETTING: Iganga/Mayuge Demographic Surveillance Site, Uganda. DESIGN: Between June and July 2008, 18 focus group discussions and 12 key informant interviews were conducted, including parents of infants and adolescents and key informant interviews with community leaders, traditional healers and patients with TB. RESULTS: People viewed TB as contagious, but not necessarily an airborne pathogen. Popular TB aetiologies included sharing utensils, heavy labour, smoking, bewitchment and hereditary transmission. TB patients were perceived to seek care late or to avoid care. Combining care from traditional healers and the biomedical system was common. Poverty, drug stock-outs, fear of human immunodeficiency virus (HIV) testing and length of TB treatment negatively affect health-seeking behaviour. Stigma and avoidance of persons with TB often reflects an assumption of HIV co-infection. CONCLUSION: The community's concerns about pill burden, quality of care, financial barriers, TB aetiology, stigma and preference for pluralistic care need to be addressed to improve early detection. Health education messages should emphasise the curability of TB, the feasibility of treatment and the engagement of traditional healers as partners in identifying cases and facilitating adherence to treatment