Diagnostic accuracy of a new Leishmania PCR for clinical visceral leishmaniasis in Nepal and its role in diagnosis of disease

Objective To develop a new PCR for Leishmania detection and to estimate its diagnostic accuracy in a visceral leishmaniasis (VL) endemic area. Methods After providing the proof‐of‐concept, the diagnostic accuracy was estimated on blood from 247 non‐endemic control persons and on blood and bone marrow from 173 confirmed VL, 39 probable VL and 87 non‐VL patients from south‐eastern Nepal. Results The PCR showed a specificity of 99.64% [95% confidence interval (CI): 98.93–100%) on non‐endemic controls and a sensitivity of 92.1% (95% CI: 87.6–96.6%) on blood and 92.9% (95% CI: 89–96.8%) on bone marrow from the confirmed VL patients. Leishmania DNA was detected in blood and bone marrow of 67.6% (95% CI: 50.8–80.9%) and 71.8% (95% CI: 56.2–83.5%) of the probable VL patients, respectively, and of 38.2% (95% CI: 28–49.4%) and 29.9% (95% CI: 21.3–40.2%) of the non‐VL patients, respectively. The PCR showed 97% concordance with a positive DAT status while for a negative DAT status this was only 41.3% (kappa‐index 0.416, 95% CI: 0.30–0.53). Conclusions Our findings indicate that PCR alone rather provides a marker for infection than a marker for disease and its role in VL diagnosis in endemic regions is discussed.     

Accordance and concordance of PCR and NASBA followed by oligochromatography for the molecular diagnosis of Trypanosoma brucei and Leishmania

Objective To evaluate the repeatability and reproducibility of four simplified molecular assays for the diagnosis of Trypanosoma brucei spp. or Leishmania ssp. in a multicentre ring trial with seven participating laboratories. Methods The tests are based on PCR or NASBA amplification of the parasites nucleic acids followed by rapid read‐out by oligochromatographic dipstick (PCR‐OC and NASBA‐OC). Results On purified nucleic acid specimens, the repeatability and reproducibility of the tests were Tryp‐PRC‐OC, 91.7% and 95.5%; Tryp‐NASBA‐OC, 95.8% and 100%; Leish‐PCR‐OC, 95.9% and 98.1%; Leish‐NASBA‐OC, 92.3% and 98.2%. On blood specimens spiked with parasites, the repeatability and reproducibility of the tests were Tryp‐PRC‐OC, 78.4% and 86.6%; Tryp‐NASBA‐OC, 81.5% and 89.0%; Leish‐PCR‐OC, 87.1% and 91.7%; Leish‐NASBA‐OC, 74.8% and 86.2%. Conclusion As repeatability and reproducibility of the tests were satisfactory, further phase II and III evaluations in clinical and population specimens from disease endemic countries are justified.     

Latent class analysis of diagnostic tests for visceral leishmaniasis in Brazil

Objective To estimate the sensitivities and specificities of different diagnostic tests for visceral leishmaniasis (VL) using latent class analysis (LCA). Methods This study was performed using data from a prospective study conducted in four Brazilian states from May 2004 to May 2007. Five diagnostic tests for VL were evaluated in 285 VL cases and 119 non‐cases: microscopy, indirect fluorescence antibody test (IFAT), enzyme‐linked immunosorbent assay using recombinant K39 antigen (rK39‐ELISA), direct agglutination test (DAT) and the rK39 rapid test. Results Microscopy showed sensitivity of 77.0% (CI: 71.5–81.5) and specificity of 99.0% (CI: 94.0–99.7). The IFAT and the DAT showed similar sensitivities, 88.3% (CI: 84.0–92.0) and 88.5% (CI: 84.1–92.0), respectively, but the DAT had a higher specificity (95.4%, CI: 89.2–98.1) than did the IFAT (83.0%, CI: 75.0–88.2). The rK39‐ELISA and the rK39 rapid test showed sensitivities of 99.0% (CI: 96.3–99.6) and 94.0% (CI: 90.1–96.3), and specificities of 82.5% (CI: 75.0–88.3) and 100% (CI: 97.0–100.0%), respectively. Conclusions Considering the lack of an adequate reference standard, LCA proved to be a useful tool in validating diagnostic methods for VL. The DAT and the rK39 rapid test showed better performance. Thus, clinically suspected cases of VL in a Brazilian endemic area could be treated based on the positivity of one of these tests.     

Performance of Latex Agglutination Test (KAtex) In Diagnosis of Visceral Leishmaniasis in Iran

Background: Visceral leishmaniasis (VL) is an endemic disease in some parts of Iran. Many techniques have been used for diagnosis of VL, among which the urine based la-tex agglutination test (KAtex) is a promising one. Objective: To compare three diag-nostic tests of VL including KAtex, ELISA and Direct Agglutination Test (DAT) in VL patients and healthy controls in the south west of Iran. Methods: Serum (n = 29) and urine samples (n = 31) were collected from parasitologically confirmed VL patients. Control samples were obtained from healthy individuals (n = 61) and also from patients with infectious diseases other than VL. The collected serum samples were tested by DAT and ELISA using crude antigen from promastigotes of Leishmania infantum and the urine samples were tested by KAtex. Results: Sensitivity and specificity of KAtex for diagnosis of VL was found to be 83.9% and 100%, respectively. Sensitivities of DAT and ELISA were 93.1% and 86.2% and their specificities were 100% and 90.5%, respectively. Conclusion: KAtex yielded a satisfactory sensitivity and specificity in di-agnosis of VL in Iran and can be recommended as a rapid, field applicable and reliable test for diagnosis of VL in this region.     

Visceral leishmaniasis in the early post‐transplant period after kidney transplantation: clinical features and therapeutic management

M. Veroux, D. Corona, G. Giuffrida, B. Cacopardo, N. Sinagra, T. Tallarita, A. Giaquinta, D. Zerbo, P.F. Veroux. Visceral leishmaniasis in the early post‐transplant period after kidney transplantation: clinical features and therapeutic management.
Transpl Infect Dis 2010: 12: 387–391. All rights reserved Abstract: Visceral leishmaniasis (VL) is a rare complication of kidney transplantation, with <100 cases reported in the literature. It is a life‐threatening condition and usually occurs as a late complication after transplantation, with a median delay of 18 months between transplantation and onset of disease. We report the clinical features and management of 5 kidney transplant recipients who presented with VL in the early post‐transplant period. All patients were successfully treated with liposomal amphotericin B (L‐AMB), but 2 patients experienced graft loss. VL should be considered in the differential diagnosis in kidney transplant recipients living in endemic areas, who present with unexplained fever and pancytopenia in the early post‐transplant period. Leishmania serology should be included in the screening of all transplant recipients, in order to identify a group of patients who could benefit from preemptive anti‐Leishmania therapy. Therapy with L‐AMB is highly effective and well tolerated in kidney transplant recipients with VL.     

Clinical characteristics and treatment outcome of patients with visceral leishmaniasis and HIV co‐infection in northwest Ethiopia

Objectives To describe the clinical presentation of patients with visceral leishmaniasis (VL) with and without human immunodeficiency virus (HIV) co‐infection and factors associated with poor outcome in northwest Ethiopia. Method Retrospective review of 241 patients with VL (92 with and 149 without HIV co‐infection). Results HIV co‐infection was present in 92 (38%) of the patients. Clinical presentation of VL was indistinguishable between patients with and without HIV co‐infection. Co‐infected patients had a poorer outcome i.e. either death or treatment failure (31.5%vs. 5.6%, P < 0.001). The presence of tuberculosis or sepsis syndrome among patients with VL and HIV co‐infected independently predicted death or treatment failure [odds ratio 4.5 (95% CI 1.47–13.92, P = 0.009) and 9.1 (95% CI 2.16–37.97, P = 0.003), respectively]. Despite having similar clinical presentation at the time of diagnosis, VL and HIV co‐infected patients had a higher mortality and treatment failure than immunocompetent patients. Conclusion The frequency of HIV co‐infection among patients with VL is high in the study area, and this co‐infection was associated with death or treatment failure. The clinical management of VL in HIV co‐infected patients is a major challenge that requires new treatment approaches to improve its outcome.     

Importance of L. Infantum H2B Recombinant Antigen for Serodiagnosis of Visceral Leishmaniasis

Background: Visceral leishmaniasis (VL) can lead to death in more than 95% of cases if left untreated. Accurate and early diagnosis has an important role in reducing mortality rate of this disease. Objective: To express recombinant H2B antigen from an Iranian isolate of Leishmania Infantum and evaluate its efficacy in the diagnosis of VL. Methods: The recombinant H2B antigen was produced in a prokaryotic system, and its efficacy for VL diagnosis was evaluated by ELISA. The serum samples from 80 VL patients, 100 individuals from endemic and non-endemic regions of VL, and 58 non-VL patients were collected. VL cases were confirmed based on the clinical sign, positive IFAT (>64), real time PCR, and response to treatment. Results: The H2B gene sequence of the Iranian L. infantum isolate had about 4% diversity in comparison with the H2B gene of the L. infantum counterpart. ELISA, using the produced H2B recombinant antigen, showed sensitivity of 71.25% (95% CI: 60.05%-80.82%) and specificity of 69.62% (95% CI: 61.81%-76.68%) regarding VL diagnosis. Conclusion: Recombinant H2B antigen expressed in the prokaryotic system had suboptimal performance for the serological diagnosis of VL. It seems that the production and expression of recombinant H2B antigen in a eukaryotic system may enhance the performance of this antigen in the diagnosis of VL in Iran.     

Visceral leishmaniasis after renal transplantation: report of 4 cases in northeastern Brazil

Abstract: Visceral leishmaniasis (VL) is a well recognized opportunistic infection in immunosuppressed patients, which may cause febrile illness. We describe 4 renal transplant patients with VL in an endemic area in Brazil and their response to therapy. In 3 cases the diagnosis was confirmed by bone marrow aspirate that revealed the presence of Leishmania. In 1 case the bone marrow aspirate was inconclusive and the diagnosis was made through spleen biopsy that showed the presence of the parasite. VL needs to be considered as a cause of febrile illness in transplanted patients living in endemic areas.     

Enhanced expression of Toll‐like receptors 2 and 4, but not 9, in spleen tissue from patients with visceral leishmaniasis

Toll‐like receptor (TLR) signalling is involved in first‐line defence against Leishmania parasites by triggering NF‐κB activation and downstream production of proinflammatory cytokines. Experimental models of visceral leishmaniasis (VL) support a protective role for TLRs 2, 4 and 9 in host immune responses to Leishmania infection. There are limited data available on expression of these TLRs in human VL, particularly in sites of infection, such as the spleen. This study aimed to determine whether the expression of mRNA encoding the expression of TLRs 2, 4 and 9 was altered in VL and compare expression patterns in splenic biopsies and peripheral blood mononuclear cells.     

Post‐kala‐azar dermal leishmaniasis in visceral leishmaniasis‐endemic communities in Bihar,India

We assessed the prevalence of post‐kala‐azar dermal leishmaniasis (PKDL), a late cutaneous manifestation of visceral leishmaniasis (VL), in 16 VL‐endemic communities in Bihar, India. The prevalence of confirmed PKDL cases was 4.4 per 10 000 individuals and 7.8 if probable cases were also considered. The clinical history and treatment of the post‐kala‐azar dermal leishmaniasis cases are discussed.     

Field validity, reproducibility and feasibility of diagnostic tests for visceral leishmaniasis in rural Nepal

OBJECTIVES: To assess the field accuracy, reproducibility and feasibility of the formol gel test (FGT), the urine latex agglutination test (KAtex) and a rK39 antigen-based dipstick for the diagnosis of visceral leishmaniasis (VL) in rural Nepal. METHOD: Patients with clinical suspicion of VL were recruited at Rangeli District Hospital (DH), a 15-bed government hospital located in south-eastern Nepal. FGT, KAtex and rK39 dipstick tests were performed on site and later repeated at a reference kala-azar diagnostic laboratory to assess reproducibility. Diagnosis of VL was confirmed by either a positive bone marrow aspirate examination or a positive direct agglutination test (DAT titre > or = 1:3200) in patients who later responded to anti-leishmanial therapy. RESULTS: Of 155 patients initially recruited, 142 (85 with VL and 57 with another diagnosis) were included in the study. The sensitivity of the rK39 dipstick [89%; 95% confidence interval (CI): 81-94] was significantly higher than that of the KAtex (57%; 95% CI: 46-67) and the FGT (52%; 95% CI: 41-62). All three tests had a specificity of at least 90%. Agreement was higher for the rK39 dipstick (kappa = 0.87) than for the FGT (0.68) and the KAtex (0.43). All tests required < or = 20 min of actual work and < or = 40 min to obtain the results. CONCLUSION: The rK39 dipstick was easy to do, more accurate and reproducible than other rapid diagnostic tests for VL in a DH of rural Nepal. It should be integrated into the field diagnostic algorithm of VL in this region and mechanisms to secure its availability should be found.     

Recombinant K28 antigen in ELISA in the diagnosis of canine visceral leishmaniosis

Crude total antigen (CTA) from Leishmania infantum and recombinant antigen K39 (rK39) and recombinant antigen K28 (rK28) were compared using an ELISA for the diagnosis of canine visceral leishmaniosis (CVL). Forty‐two blood samples from healthy dogs from a nonendemic area and 80 blood samples from an endemic area for dogs with visceral leishmaniosis (VL), confirmed with positive parasitological tests for Leishmania spp., were used in an ELISA. The parasitological diagnosis was chosen as a gold standard. The ELISA with rK28 antigen showed sensitivity of 100% and specificity of 100%, high agreement with CTA and rK39, indicating that the rK28 antigen is useful for ELISA serological diagnosis of CVL.     

Comparative evaluation of parasitology and serological tests in the diagnosis of visceral leishmaniasis in India: a phase III diagnostic accuracy study

In this phase III trial for diagnostics for visceral leishmaniasis (VL) in India, we compared parasitological diagnosis with several serological tests: direct agglutination test (freeze dried; DAT-FD), rK-39 strip test, rK-26 strip test and a latex agglutination test for antigen detection in urine (KAtex) in 452 subjects from the endemic regions of Bihar, India. The subjects were segregated into four categories: 230 confirmed patients, 52 probable cases, 70 non-cases and 100 healthy endemic controls. The first two groups were used for estimating sensitivity, the latter two for specificity. Sensitivity of DAT-FD was 98.9%, rK-39: 98.9%, KAtex: 67.0% and rK-26: 21.3%. Sensitivity of DAT-FD on blood taken on filter paper (DAT-FDF) was 99.3%, which was comparable with that using serum. Specificity of serological tests was comparable and high (DAT-FD and DAT-FDF: 94%, rK-39 strip test: 97%, KAtex: 99% and rK-26 strip test: 100%). The classical 'gold standard' parasitological demonstration in splenic smear performed poorly as it missed 18.4% of cases that benefited from VL treatment. Reproducibility of the serological tests between field and central laboratories was excellent (kappa = 1.0, 0.99, 0.96 and 0.94 respectively for microscopy, DAT-FD, rK-39 strip test and rK-26 strip test). A high degree of agreement was observed between DAT-FD and rK-39 strip test (kappa = 0.986). Although DAT-FD and rK-39 strip test were highly sensitive with excellent specificity, the ease of use of the latter makes it most suitable for the diagnosis of VL in the field conditions.     

A simplified and standardized polymerase chain reaction format for the diagnosis of leishmaniasis

BACKGROUND: Definite diagnosis of Leishmania infections is based on demonstration of the parasite by microscopic analysis of tissue biopsy specimens or aspirate samples. However, microscopy generally shows low sensitivity and requires invasive sampling. METHODS: We describe here the development of a simple and rapid test for the detection of polymerase chain reaction-amplified Leishmania DNA. A phase 1 evaluation of the text was conducted in clinical samples from 60 nonendemic and 45 endemic control subjects and from 44 patients with confirmed cutaneous leishmaniasis (CL), 12 with mucocutaneous leishmaniasis (MCL), and 43 with visceral leishmaniasis (VL) from Peru, Kenya, and Sudan. RESULTS: The lower detection limits of the assay are 10 fg of Leishmania DNA and 1 parasite in 180 microL of blood. The specificity was 98.3% (95% confidence interval [CI], 91.1%-99.7%) and 95.6% (95% CI, 85.2%-98.8%) for nonendemic and endemic control samples, respectively, and the sensitivity was 93.2% (95% CI, 81.8%-97.7%), 91.7% (95% CI, 64.6%-98.5%), and 86% (95% CI, 72.7%-93.4%) for lesions from patients with CL or MCL and blood from patients with VL, respectively. CONCLUSIONS: The Leishmania OligoC-TesT showed high specificity and sensitivity in clinical samples and was able to detect the parasite in samples obtained by less invasive means, such as blood, lymph, and lesion scrapings. The assay is a promising new tool for simplified and standardized molecular detection of Leishmania parasites.     

Low castes have poor access to visceral leishmaniasis treatment in Bihar,India

Objectives Bihar, the poorest state in India, concentrates most of the visceral leishmaniasis (VL) cases in the country. A large proportion of the poor rural communities where VL is endemic are marginalized by their socio‐economic status, intrinsically related to the caste system. In this study, we evaluated whether people from low socio‐economic strata had difficulties accessing VL treatment in Bihar. As a secondary outcome, we evaluated whether people delaying their VL treatment had poorer clinical indicators at admission. Methods Data on 2187 patients with VL treated by Médecins Sans Frontières (MSF) in Vaishali district from July 2007 to December 2008 were analysed. Patients who reported having onset of symptoms ≥8 weeks before admission were defined as ‘late presenters’. Logistic regression models were used to evaluate whether low castes had higher risk to be ‘late presenters’ compared to the rest of castes and whether ‘late presenters’ had poorer indicators at admission (i.e. haemoglobin level, spleen size). Results After adjusting for age, gender and distance to VL treatment facility, Mushars (the lowest caste in Bihar) had twice the odds to be ‘late presenters’ compared to the rest of castes (OR 2.05, 95% CI: 1.24–2.38). Subjects that had VL symptoms for ≥8 weeks had a larger spleen and lower haemoglobin level than those that were treated earlier. Conclusion Low castes have poor access to VL treatment in Bihar, and late presenters have poorer clinical indicators at admission. These findings have implications at individual and community levels and should stimulate targeted VL control programmes to ensure that marginalized communities in Bihar are properly treated.     

Out‐of‐pocket costs for paediatric admissions in district hospitals in Kenya

Objective To describe out‐of‐pocket costs of inpatient care for children under 5 years of age in district hospitals in Kenya. Methods A total of 256 caretakers of admitted children were interviewed in 2‐week surveys conducted in eight hospitals in four provinces in Kenya. Caretakers were asked to report care seeking behaviour and expenditure related to accessing inpatient care. Family socio‐economic status was assessed through reported expenditure in the previous month. Results Seventy eight percent of caretakers were required to pay user charges to access inpatient care for children. User charges (mean, US$ 8.1; 95% CI, 6.4–9.7) were 59% of total out‐of‐pocket costs, while transport costs (mean, US$ 4.9; 95% CI, 3.9–6.0) and medicine costs (mean, US$ 0.7; 95% CI, 0.5–1.0) were 36% and 5%, respectively. The mean total out‐of‐pocket cost per paediatric admission was US$ 14.1 (95% CI, 11.9–16.2). Out‐of‐pocket expenditures on health were catastrophic for 25.4% (95% CI, 18.4–33.3) of caretakers interviewed. Out‐of‐pocket expenditures were regressive, with a greater burden being experienced by households with lower socio‐economic status. Conclusion Despite a policy of user fee exemption for children under 5 years of age in Kenya, our findings show that high unofficial user fees are still charged in district hospitals. Financing mechanisms that will offer financial risk protection to children seeking care need to be developed to remove barriers to child survival.     

Retention of HIV‐infected and HIV‐exposed children in a comprehensive HIV clinical care programme in Western Kenya

Background To describe incidence rates (IR) and risk factors for loss‐to‐follow‐up (LTFU) among HIV‐infected and HIV‐exposed children in a large HIV treatment programme in Western Kenya. Methods The USAID‐AMPATH Partnership has enrolled >100 000 patients (20% children) at 23 clinic sites throughout western Kenya. LTFU is defined as being absent from the clinic for >3 months if on combination antiretroviral treatment (cART) and >6 months if not. Included in this analysis were children aged <14 years, HIV exposed or infected at enrolment, and enrolled between April 2002 and March 2009. The IR for LTFU are presented per 100 child‐years (CY) of follow‐up. Proportional hazards models with time‐independent and time‐dependent covariates were used to model factors associated with LTFU. Weight for height Z‐scores were calculated using EpiInfo, with severe malnutrition being defined as a Z‐score ≤?3.0. Immune suppression was defined as per WHO age‐specific categories. Results There were 13 510 children eligible for analysis, comprising 3106 children who at enrolment were HIV infected and 10 404 children who were HIV exposed. The overall IR of LTFU was 18.4 (17.8–18.9) per 100 CY. Among HIV‐infected children, 15.2 (13.8–16.7) and 14.1 (13.1–15.8) per 100 CY became LTFU, pre‐ and post‐cART initiation, respectively. The only independent risk factor for becoming LTFU among the HIV‐infected children was severe immune suppression (AHR: 2.17, 95% CI: 1.51–3.12). Among the HIV‐exposed children, 20.1 per 100 (19.4–20.7) became LTFU. Independent risk factors for LTFU among them were being severely low weight for height (AHR: 1.69, 95% CI: 1.25–2.28), being orphaned at enrolment (AHR: 1.57, 95% CI: 1.23–1.64), being CDC Class B or C (AHR: 1.41, 95% CI: 1.14–1.74), and having received cART (AHR: 1.56, 95% CI: 1.23–1.99). Protective against becoming LTFU among the HIV exposed were testing HIV positive (AHR: 0.26, 95% CI: 0.21–0.32), older age (AHR: 0.90, 95% CI: 0.85–0.96), enrolling in later time periods, and receiving food supplementation (AHR: 0.58, 95% CI: 0.32–1.04). Conclusions There is a high rate of LTFU among these highly vulnerable children, particularly among the HIV exposed. These data suggest that HIV‐infected and HIV‐exposed children are at especially high risk for LTFU if they are sick or malnourished.     

Plasma lipoproteins in visceral leishmaniasis and their effect on Leishmania‐infected macrophages

This work aimed at investigating the lipid profile of zoonotic visceral leishmaniasis (VL) patients’ sera and the effect of lipoproteins on the in vitro production of tumour necrosis factor‐α (TNF‐α), interleukin (IL)‐6, IL‐10 and IL‐12 by Leishmania infantum‐infected and uninfected macrophages. Lipids were quantified in 26 VL patients’ sera and 26 healthy controls from a VL endemic area. The patients’ sera had higher triglyceride and very low density lipoprotein (VLDL) levels, and much lower apolipoprotein A1, total cholesterol, low density lipoprotein (LDL) and high density lipoprotein (HDL) levels than the control sera. Lipoprotein fractions were obtained by ultracentrifugation of sera. The addition of LDL and HDL to Leishmania‐infected and uninfected macrophages, in physiological concentrations, enhanced the production of IL‐6 and IL‐10, but not of IL‐12. LDL stimulated the production of TNF‐α only in infected macrophages, whereas HDL stimulated the production of lower amounts of TNF‐α in both infected and uninfected macrophages. VLDL stimulated only the production of IL‐10. It is proposed herein that LDL may influence the development of VL by promoting the production of TNF‐α by infected macrophages. A decrease in plasma LDL in some VL patients (to 20 mg/mL or less); however, would tend to reduce the production of TNF‐α and therefore to limit the development of immune‐mediated pathology, not withstanding the fact that it would perhaps increase the permissiveness of macrophages to Leishmania growth.     

Visceral leishmaniasis in southeastern Nepal: a cross-sectional survey on Leishmania donovani infection and its risk factors

OBJECTIVE: To document the frequency of Leishmania donovani infection at community level in a highly endemic region in southeastern Nepal, and to assess socioeconomic and environmental risk factors. METHODS: A random cross-sectional population survey was held in two visceral leishmaniasis (VL) foci in Morang District in April to May 2003, enrolling individuals 2 years or older and residing in the endemic area for at least 12 months. Leishmania infection was defined as a direct agglutination test (DAT) titre equal to or higher than 1:3200. Risk factors were identified by logistic regression. RESULTS: The direct agglutination test was positive in 7.5% (95% CI: 5.1-10.8) and the leishmanin skin test (LST) in 13.2% (95% CI: 9.9-17.2) of the 373 study participants. No case of current kala-azar was found, but 5.1 % (95% CI: 3.1-7.8) reported having suffered from VL. Independent risk factors for Leishmania infection were proximity of the house to ponds [odds ratio (OR) 3.7, 95% CI: 1.6-8.5], family size (OR 4.4, 95% CI: 1.6-12.6), age > or =15 years (OR 5.5, 95% CI: 1.2-25.0) and house constructed in mud (OR 3.0, 95% CI: 1.1-7.6). Bednets, not impregnated and in poor condition, were used by 95.2% (95% CI: 92.3-97.0) of the population, but did not show any protective effect. CONCLUSION: This study shows that there is a serious problem of transmission of VL in this area of Nepal. The risk factors identified are linked with the socioeconomic level and the environment. The population would benefit from a community intervention to improve the environmental and housing conditions in the villages.     

Early HIV viral load determination after initiating first‐line antiretroviral therapy for indentifying patients with high risk of developing virological failure: data from a cohort study in a resource‐limited setting

Objectives To evaluate the performance of a single determination of HIV viral load (VL) 6–12 months after starting antiretroviral therapy (ART) for identifying patients who will subsequently develop virological failure. Methods We selected HIV‐infected patients with at least two VL determinations after 6 months of ART from an HIV cohort study in India. Patients were divided in two groups depending on whether the first VL was below (Group 1) or above (Group 2) 1000 copies/ml. Cut‐off for virological failure was defined according to World Health Organization recommendation (>5000 copies/ml). Results The study included 584 patients and 560.1 person‐years of follow‐up. Of all virological failures, 83% were diagnosed at the first VL determination. The cumulative incidence of virological failure after 1 and 2 years since the first VL was 0.9% [95% confidence interval (CI), 0.3–2.7] and 1.7% (95% CI, 0.6–5), respectively, for Group 1, and 58.2% (95% CI, 47–69.7) and 63.1% (95% CI, 49.8–76.4), respectively, for Group 2. Compared with Group 1, patients from Group 2 had a hazard ratio for virological failure of 78.3 (95% CI, 27.8–220.2). Conclusions A single VL determination after 6 months of ART was able to identify patients with high risk of virological failure.