Plasma cytokines quantification among Trypanosoma brucei rhodesiense sleeping sickness cases and controls in Rumphi, Malawi.

Introduction: Trypanosoma brucei (T.b.) rhodesiense is the cause of the acute form of human African trypanosomiasis (HAT) in eastern and southern African countries, including Malawi. For a long time, untreated HAT infections were believed to be 100% fatal. However, recent studies show that infection by T.b. rhodesiense can result in a wide range of clinical outcomes in its human host. Apart from other factors such as parasite diversity, cytokines have been strongly implicated to play a major role in the outcome of T.b. rhodesiense infections.In this study, we quantify the levels of three cytokines Interleukin-8 (IL-8), Tumor Necrotic Factor alpha (TNF-α) and Interleukin -10 (IL-10) in plasma amongst HAT cases (treated and untreated) and controls recruited during medical survey. Methods: Two-hundred and thirty-three plasma samples (HAT cases and controls) from Rumphi, one of the endemic areas in Malawi were used. Blood collected was centrifuged, plasma extracted and stored in cryovials at -80°C until processing. Plasma cytokine concentration was measured using ELISA. Results: Plasma samples for 233 individuals, 76 HAT cases and 157 controls were quantified. Among the cases, nine had their plasma collected before treatment (untreated) and the rest were treated before blood for plasma analysis was collected. Controls had significantly higher mean plasmatic levels of TNF-α (94.5 ±474.12 pg/ml) and IL-8 (2258.6 ±5227.4 pg/ml) than cases TNF-α (29.35±181.58 pg/ml) and IL-8 (1191.3±4236.09 pg/ml). Controls and cases had similar mean levels of IL-10 in plasma. Only IL-8 had statistically significant higher median levels in the untreated than treated HAT cases P=0.006. Conclusion: Our data suggest that cytokines could be considered as biomarkers of HAT infection and treatment. Further studies with a larger cohort of cases and additional cytokines which are known to be associated with HAT infection outcomes will be required to evaluate these cytokines further.

High Levels of Genetic Diversity within Nilo-Saharan Populations: Implications for Human Adaptation.

Africa contains more human genetic variation than any other continent, but the majority of the population-scale analyses of the African peoples have focused on just two of the four major linguistic groups, the Niger-Congo and Afro-Asiatic, leaving the Nilo-Saharan and Khoisan populations under-represented. In order to assess genetic variation and signatures of selection within a Nilo-Saharan population and between the Nilo-Saharan and Niger-Congo and Afro-Asiatic, we sequenced 50 genomes from the Nilo-Saharan Lugbara population of North-West Uganda and 250 genomes from 6 previously unsequenced Niger-Congo populations. We compared these data to data from a further 16 Eurasian and African populations including the Gumuz, another putative Nilo-Saharan population from Ethiopia. Of the 21 million variants identified in the Nilo-Saharan population, 3.57 million (17%) were not represented in dbSNP and included predicted non-synonymous mutations with possible phenotypic effects. We found greater genetic differentiation between the Nilo-Saharan Lugbara and Gumuz populations than between any two Afro-Asiatic or Niger-Congo populations. F3 tests showed that Gumuz contributed a genetic component to most Niger-Congo B populations whereas Lugabara did not. We scanned the genomes of the Lugbara for evidence of selective sweeps. We found selective sweeps at four loci (SLC24A5, SNX13, TYRP1, and UVRAG) associated with skin pigmentation, three of which already have been reported to be under selection. These selective sweeps point toward adaptations to the intense UV radiation of the Sahel.

Copy number variation in human genomes from three major ethno-linguistic groups in Africa.

BACKGROUND: Copy number variation is an important class of genomic variation that has been reported in 75% of the human genome. However, it is underreported in African populations. Copy number variants (CNVs) could have important impacts on disease susceptibility and environmental adaptation. To describe CNVs and their possible impacts in Africans, we sequenced genomes of 232 individuals from three major African ethno-linguistic groups: (1) Niger Congo A from Guinea and Côte d'Ivoire, (2) Niger Congo B from Uganda and the Democratic Republic of Congo and (3) Nilo-Saharans from Uganda. We used GenomeSTRiP and cn.MOPS to identify copy number variant regions (CNVRs). RESULTS: We detected 7608 CNVRs, of which 2172 were only deletions, 2384 were only insertions and 3052 had both. We detected 224 previously un-described CNVRs. The majority of novel CNVRs were present at low frequency and were not shared between populations. We tested for evidence of selection associated with CNVs and also for population structure. Signatures of selection identified previously, using SNPs from the same populations, were overrepresented in CNVRs. When CNVs were tagged with SNP haplotypes to identify SNPs that could predict the presence of CNVs, we identified haplotypes tagging 3096 CNVRs, 372 CNVRs had SNPs with evidence of selection (iHS > 3) and 222 CNVRs had both. This was more than expected (p < 0.0001) and included loci where CNVs have previously been associated with HIV, Rhesus D and preeclampsia. When integrated with 1000 Genomes CNV data, we replicated their observation of population stratification by continent but no clustering by populations within Africa, despite inclusion of Nilo-Saharans and Niger-Congo populations within our dataset. CONCLUSIONS: Novel CNVRs in the current study increase representation of African diversity in the database of genomic variants. Over-representation of CNVRs in SNP signatures of selection and an excess of SNPs that both tag CNVs and are subject to selection show that CNVs may be the actual targets of selection at some loci. However, unlike SNPs, CNVs alone do not resolve African ethno-linguistic groups. Tag haplotypes for CNVs identified may be useful in predicting African CNVs in future studies where only SNP data is available.

Association of APOL1 renal disease risk alleles with Trypanosoma brucei rhodesiense infection outcomes in the northern part of Malawi.

Trypanosoma brucei (T.b.) rhodesiense is the cause of the acute form of human African trypanosomiasis (HAT) in eastern and southern African countries. There is some evidence that there is diversity in the disease progression of T.b. rhodesiense in different countries. HAT in Malawi is associated with a chronic haemo-lymphatic stage infection compared to other countries, such as Uganda, where the disease is acute with more marked neurological impairment. This has raised the question of the role of host genetic factors in infection outcomes. A candidate gene association study was conducted in the northern region of Malawi. This was a case-control study involving 202 subjects, 70 cases and 132 controls. All individuals were from one area; born in the area and had been exposed to the risk of infection since birth. Ninety-six markers were genotyped from 17 genes: IL10, IL8, IL4, HLA-G, TNFA, IL6, IFNG, MIF, APOL, HLA-A, IL1B, IL4R, IL12B, IL12R, HP, HPR, and CFH. There was a strong significant association with APOL1 G2 allele (p = 0.0000105, OR = 0.14, CI95 = [0.05-0.41], BONF = 0.00068) indicating that carriers of the G2 allele were protected against T.b. rhodesiense HAT. SNP rs2069845 in IL6 had raw p < 0.05, but did not remain significant after Bonferroni correction. There were no associations found with the other 15 candidate genes. Our finding confirms results from other studies that the G2 variant of APOL1 is associated with protection against T.b. rhodesiense HAT.

Comparative evaluation of dry and liquid RIME LAMP in detecting trypanosomes in dead tsetse flies.

Xenomonitoring is an important approach in assessing the progress of trypanosomiasis control as well as in estimating the endemicity of trypanosomes in affected areas. One of the major challenges in this approach is the unavailability of sensitive and easy to use xenomonitoring tools that can be used in the remote areas where the disease occurs. One tool that has been used successfully in detecting the parasites in tsetse flies is the repetitive insertion mobile element loop-mediated isothermal amplification (RIME LAMP). This tool has recently been modified from the liquid form to dry form for use in remote areas; however, uptake for use in the field has been slow. Field-collected tsetse flies were used to evaluate the performance of dry RIME LAMP over the conventional liquid RIME LAMP. All the samples were also subjected to internal transcribed spacer 1 (ITS1) ribosomal deoxyribonucleic acid (DNA) polymerase chain reaction (PCR) as a standard. ITS1-PCR-positive samples were further sequenced for confirmation of the species. A total of 86 wild tsetse flies were left to dry at room temperature for 3 months and DNA was extracted subsequently. All 86 flies were Glossina morsitans morsitans. From these, dry RIME LAMP detected 16.3% while liquid RIME LAMP detected 11.6% as infected with trypanosomes. Ten positive samples on ITS1-PCR were sequenced and all were shown to be trypanosomes. The use of dry RIME LAMP in the field for xenomonitoring of trypanosomes in tsetse flies will greatly contribute towards control of this neglected tropical disease as it provides the cheapest, fastest and simplest way to estimate possible human infective trypanosome infection rates in the tsetse fly vectors.

How operational research influenced the scale up of antiretroviral therapy in Malawi.

The national scale up of antiretroviral therapy in Malawi is based on a public health approach, with principles and practices borrowed from the successful World Health Organization "DOTS" tuberculosis control framework. The scale up of antiretroviral therapy was under-pinned by a very strong monitoring and evaluation system, which was used to audit the scale up approach and conduct operational research to answer relevant questions. Examples of research included:- i) access to antiretroviral therapy, populations and social groups served, and how the different groups fared with regard to outcomes; ii) determining whether the quality of data at antiretroviral therapy sites was adequate and whether external supervision was needed; iii) finding feasible ways of reducing the high early mortality in patients starting treatment in both Malawi and the sub-Saharan African region; iv) the causes of loss-to-follow-up, what happened to patients who transferred out of sites and whether transfer-out patients had outcomes comparable to those who did not transfer; and v) the important question of whether antiretroviral therapy scale up reduced population mortality. The answers to these questions had an important influence on how treatment was delivered in the country, and show the value of this work within a programme setting. Key generic lessons include the importance of i) research questions being relevant to programme needs, ii) studies being coordinated, designed and undertaken within a programme, iii) study findings being disseminated at national stakeholder meetings and through publications in peer-reviewed journals and iv) research being used to influence policy and practice, improve programme performance and ultimately patient treatment outcomes.

Mortality reduction associated with HIV/AIDS care and antiretroviral treatment in rural Malawi: evidence from registers, coffin sales and funerals.

BACKGROUND: To report on the trend in all-cause mortality in a rural district of Malawi that has successfully scaled-up HIV/AIDS care including antiretroviral treatment (ART) to its population, through corroborative evidence from a) registered deaths at traditional authorities (TAs), b) coffin sales and c) church funerals. METHODS AND FINDINGS: Retrospective study in 5 of 12 TAs (covering approximately 50% of the population) during the period 2000-2007. A total of 210 villages, 24 coffin workshops and 23 churches were included. There were a total of 18,473 registered deaths at TAs, 15781 coffins sold, and 2762 church funerals. Between 2000 and 2007, there was a highly significant linear downward trend in death rates, sale of coffins and church funerals (X(2) for linear trend: 338.4 P<0.0001, 989 P<0.0001 and 197, P<0.0001 respectively). Using data from TAs as the most reliable source of data on deaths, overall death rate reduction was 37% (95% CI:33-40) for the period. The mean annual incremental death rate reduction was 0.52/1000/year. Death rates decreased over time as the percentage of people living with HIV/AIDS enrolled into care and ART increased. Extrapolating these data to the entire district population, an estimated 10,156 (95% CI: 9786-10259) deaths would have been averted during the 8-year period. CONCLUSIONS: Registered deaths at traditional authorities, the sale of coffins and church funerals showed a significant downward trend over a 8-year period which we believe was associated with the scaling up HIV/AIDS care and ART.

Diagnosis and management of antiretroviral-therapy failure in resource-limited settings in sub-Saharan Africa: challenges and perspectives.

Despite the enormous progress made in scaling up antiretroviral therapy (ART) in sub-Saharan Africa, many challenges remain, not least of which are the identification and management of patients who have failed first-line therapy. Less than 3% of patients are receiving second-line treatment at present, whereas 15-25% of patients have detectable viral loads 12 months or more into treatment, of whom a substantial proportion might have virological failure. We discuss the reasons why virological ART failure is likely to be under-diagnosed in the routine health system, and address the current difficulties with standard recommended second-line ART regimens. The development of new diagnostic tools for ART failure, in particular a point-of-care HIV viral-load test, combined with simple and inexpensive second-line therapy, such as boosted protease-inhibitor monotherapy, could revolutionise the management of ART failure in resource-limited settings.

Scaling up antiretroviral therapy in Malawi-implications for managing other chronic diseases in resource-limited countries.

The national scale-up of antiretroviral therapy (ART) in Malawi is based on the public health approach, with principles and practices borrowed from the successful DOTS (directly observed treatment, short course) tuberculosis control framework. The key principles include political commitment, free care, and standardized systems for case finding, treatment, recording and reporting, and drug procurement. Scale-up of ART started in June 2004, and by December 2008, 223,437 patients were registered for treatment within a health system that is severely underresourced. The Malawi model for delivering lifelong ART can be adapted and used for managing patients with chronic noncommunicable diseases, the burden of which is already high and continues to grow in low-income and middle-income countries. This article discusses how the principles behind the successful Malawi model of ART delivery can be applied to the management of other chronic diseases in resource-limited settings and how this paradigm can be used for health systems strengthening.

Knowledge of HIV status, sexual risk behaviors and contraceptive need among people living with HIV in Kenya and Malawi.

BACKGROUND: Several studies support the need for effective interventions to reduce HIV transmission risk behaviors among people living with HIV/AIDS (PLWHAs). DESIGN: Cross-sectional nationally representative demographic health survey of Kenya (2003) and Malawi (2004-2005) that included HIV testing for consenting adults. METHODS: We analyzed demographic health survey data for awareness of HIV status and sexual behaviors of PLWHAs (Kenya: 412; Malawi: 664). The analysis was adjusted (weighted) for the design of the survey and the results are nationally representative. FINDINGS: Eighty-four percent of PLWHAs in Kenya and 86% in Malawi had sex in the past 12 months and in each country, 10% reported using condoms at last intercourse. Among sexually active PLWHAs, 86% in Kenya and 96% in Malawi reported their spouse or cohabiting partner as their most recent partner. In multivariate logistic regression models, married or cohabiting PLWHAs were significantly more likely to be sexually active and less likely to use condoms. Over 80% of PLWHAs were unaware of their HIV status. Of HIV-infected women, nearly three-quarters did not want more children either within the next 2 years or ever, but 32% in Kenya and 20% in Malawi were using contraception. INTERPRETATION: In 2003-2005, majority of PLWHAs in Kenya and Malawi were unaware of their HIV status and were sexually active, especially married or cohabiting PLWHAs. Of HIV-infected women not wanting more children, few used contraception. HIV testing should be expanded, prevention programs should target married or cohabiting couples and family planning services should be integrated with HIV services.

Burden of cryptococcal meningitis in Malawi.

There is little information about the national burden of cryptococcal meningitis (CM) in African countries affected by the HIV/AIDS epidemic. From April 2005 onwards, we used national supervision visits of all health facilities that provided antiretroviral therapy to collect data on the number of new patients diagnosed and treated for CM in the previous quarters - using mainly fluconazole registers. For two 12-month reporting periods, there were 2125 and 2464 patients suffering from CM, giving an estimated annual incidence of 2.2% and 2.6%, respectively, of those infected with HIV in Malawi. Between 40-50% of all patients with CM were diagnosed and treated at central hospitals; no more than 1% were diagnosed and treated at smaller antiretroviral therapy sites. These data are useful for quantifying the need for better diagnostic reagents and antifungal drugs.

A public health approach to rapid scale-up of antiretroviral treatment in Malawi during 2004-2006.

BACKGROUND: Approximately 1 million people are infected with HIV in Malawi, where AIDS is the leading cause of death in adults. By December 31, 2007, more than 141,000 patients were initiated on antiretroviral treatment (ART) by use of a public health approach to scale up HIV services. METHODS: We analyzed national quarterly and longitudinal cohort data from October 2004 to December 2006 to examine trends in characteristics of patients initiating ART, end-of-quarter clinical outcomes, and 6- and 12-month survival probability. FINDINGS: During a 27-month period, 72,666 patients were initiated on ART, of whom about two-thirds were women. The percentage of patients initiated on ART who were children and farmers increased from 5.5% to 9.0% and 23% to 32%, respectively (P < 0.001 for trends). Estimated survival probability ranged from 85% to 88% at 6 months and 81% to 84% at 12 months on ART. INTERPRETATION: In Malawi, a public health approach to ART increased treatment access and maintained high 6- and 12-month survival. Resource-limited countries scaling up ART programs may benefit from this approach of simplified clinical decision making, standardized ART regimens, nonphysician care, limited laboratory support, and centralized monitoring and evaluation.

World Health Organization Clinical Stage 3 disease conditions in HIV-infected patients who start antiretroviral therapy in Malawi.

There is little information about disease conditions that are diagnosed in patients diagnosed as having World Health Organization Clinical Stage 3 HIV who are started on antiretroviral therapy (ART) in Africa. We therefore conducted an audit in the central region of Malawi of patients registered for ART between January and September 2006. There were 4299 patients in Stage 3 of whom 4154 had data about their disease conditions. Only one condition was listed for 3880 patients. Of these, 1892 (48.8%) had unexplained weight loss, chronic fever or chronic diarrhoea, 822 (21.2%) had active/previous tuberculosis (TB) and 671 (17.3%) had a severe presumed bacterial infection. No patient was diagnosed as having haematological abnormalities. Nearly half the patients started on ART had a symptomatic, unspecified disease, (which may be obscuring important pathologies such as TB) and almost no laboratory assessment had taken place before the commencement of ART. These two areas need to be addressed in order to improve the management of patients starting on ART.

A national survey of prisoners on antiretroviral therapy in Malawi: access to treatment and outcomes on therapy.

BACKGROUND: Malawi is making good progress scaling up antiretroviral therapy (ART), but we do not know the levels of access of high-risk, disadvantaged groups such as prisoners. The aim of this study was to measure access and treatment outcomes of prisoners on ART at the national level. METHODOLOGY: A retrospective cohort study was conducted examining patient follow-up records from all 103 public sector ART clinics in Malawi, and observations were censored on 31 December, 2006. RESULTS: By 31 December, 2006, a total of 81,821 patients had been started on ART. Of these, 103 (0.13%) were prisoners. At ART initiation, 93% of prisoners were in World Health Organization (WHO) clinical stage 3 or 4 while 7% started in stage 1 or 2 with a CD4-lymphocyte count of < or =250/mm3. Treatment outcomes by the end of December 2006 were as follows: 66 (64%) alive and on ART at their registration facility; 9 (9%) dead; 8 (8%) lost to follow-up; and 20 (19%) transferred out to another facility. The probability of being alive and on ART at 6 and 12 months was 82.5% and 77.7%. CONCLUSIONS: In spite of the rapid scale-up of ART, only a small number of HIV-positive prisoners had accessed ART by the end of 2006. Treatment outcomes were good. Initiatives are now needed to improve access to HIV testing and ART in Malawi's prisons.

Early warning indicators for HIV drug resistance in Malawi.

BACKGROUND: Malawi started rapid scale-up of antiretroviral therapy (ART) in 2004 and by December 2006 had initiated over 85,000 patients on treatment. Early warning indicator (EWI) reports can help to minimize the risk of emerging drug resistance. METHODS: Data collected during the routine quarterly supervision of 103 public sector sites was used to compile the first EWI report for HIV drug resistance (HIVDR) in Malawi, reflecting outcomes for October to December 2006. RESULTS: All sites reach the World Health Organization (WHO) targets for prescribing practices and drug supply continuity. The target for adherence was achieved by 85% of sites and 84% achieved the target for minimizing treatment defaults; however, less than half of all sites reach the WHO target for patient retention. CONCLUSIONS: These results emphasize the importance of defaulter tracing and initiating treatment earlier in the course of HIV infection. As part of a comprehensive HIVDR monitoring programme, the Ministry of Health plans for on-going tracking of these indicators, as well as special data collection from the private sector. Plans are also underway to gather information on other recommended indicators that are not collected during routine supervision.

Surveillance of transmitted HIV drug resistance with the World Health Organization threshold survey method in Lilongwe, Malawi.

BACKGROUND: Malawi started rapid scale-up of antiretroviral therapy (ART) in 2004 and by December 2006 had initiated 81,821 patients on treatment in the public sector. Owing to capacity constraints, standard patient care, treatment initiation and follow up are based on World Health Organization (WHO) clinical staging and do not provide laboratory monitoring to assess treatment failure. METHODS: To monitor possible transmission of HIV drug resistance (HIVDR) an HIVDR threshold surveillance based on the WHO guidelines was implemented in Malawi. Anonymous dried blood specimens were collected from routine blood samples of HIV-positive women attending primagravida antenatal care and aged <25 years. RESULTS: Of 59 samples tested, 54 were successfully amplified indicating good specimen quality and processing. The WHO protocol algorithm to classify the prevalence of transmitted drug resistance in the site sample only required the genotyping of 34 of the samples. None of the major drug resistance mutations on the WHO surveillance list were found in these 34 specimens. CONCLUSIONS: Malawi HIVDR transmission can be classified as <5% for all relevant drugs and drug classes in this population. On the basis of the very positive experience of this survey, an expanded HIVDR surveillance system will be implemented to inform the ART program as it continues to scale-up.

What happens to patients on antiretroviral therapy who transfer out to another facility?

BACKGROUND: Long term retention of patients on antiretroviral therapy (ART) in Africa's rapidly expanding programmes is said to be 60% at 2 years. Many reports from African ART programmes make little mention of patients who are transferred out to another facility, yet Malawi's national figures show a transfer out of 9%. There is no published information about what happens to patients who transfer-out, but this is important because if they transfer-in and stay alive in these other facilities then national retention figures will be better than previously reported. METHODOLOGY/PRINCIPAL FINDINGS: Of all patients started on ART over a three year period in Mzuzu Central Hospital, North Region, Malawi, those who transferred out were identified from the ART register and master cards. Clinic staff attempted to trace these patients to determine whether they had transferred in to a new ART facility and their outcome status. There were 805 patients (19% of the total cohort) who transferred out, of whom 737 (92%) were traced as having transferred in to a new ART facility, with a median time of 1.3 months between transferring-out and transferring-in. Survival probability was superior and deaths were lower in the transfer-out patients compared with those who did not transfer. CONCLUSION/SIGNIFICANCE: In Mzuzu Central Hospital, patients who transfer-out constitute a large proportion of patients not retained on ART at their original clinic of registration. Good documentation of transfer-outs and transfer-ins are needed to keep track of national outcomes. Furthermore, the current practice of regarding transfer-outs as being double counted in national cohorts and subtracting this number from the total national registrations to get the number of new patients started on ART is correct.

Assessing the quality of data aggregated by antiretroviral treatment clinics in Malawi.

PROBLEM: As national antiretroviral treatment (ART) programmes scale-up, it is essential that information is complete, timely and accurate for site monitoring and national planning. The accuracy and completeness of reports independently compiled by ART facilities, however, is often not known. APPROACH: This study assessed the quality of quarterly aggregate summary data for April to June 2006 compiled and reported by ART facilities ("site report") as compared to the "gold standard" facility summary data compiled independently by the Ministry of Health supervision team ("supervision report"). Completeness and accuracy of key case registration and outcome variables were compared. Data were considered inaccurate if variables from the site reports were missing or differed by more than 5% from the supervision reports. Additionally, we compared the national summaries obtained from the two data sources. LOCAL SETTING: Monitoring and evaluation of Malawi's national ART programme is based on WHO's recommended tools for ART monitoring. It includes one master card for each ART patient and one patient register at each ART facility. Each quarter, sites complete cumulative cohort analyses and teams from the Ministry of Health conduct supervisory visits to all public sector ART sites to ensure the quality of reported data. RELEVANT CHANGES: Most sites had complete case registration and outcome data; however many sites did not report accurate data for several critical data fields, including reason for starting, outcome and regimen. The national summary using the site reports resulted in a 12% undercount in the national total number of persons on first-line treatment. Several facility-level characteristics were associated with data quality. LESSONS LEARNED: While many sites are able to generate complete data summaries, the accuracy of facility reports is not yet adequate for national monitoring. The Ministry of Health and its partners should continue to identify and support interventions such as supportive supervision to build sites' capacity to maintain and compile quality data to ensure that accurate information is available for site monitoring and national planning.

Antiretroviral therapy in the Malawi defence force: access, treatment outcomes and impact on mortality.

BACKGROUND: HIV/AIDS affects all sectors of the population and the defence forces are not exempt. A national survey was conducted in all public and private sectors in Malawi that provide antiretroviral therapy (ART) to determine the uptake of ART by army personnel, their outcomes while on treatment, and the impact of ART on mortality in the Malawi Defence Force. METHODOLOGY/PRINCIPAL FINDINGS: A retrospective cohort analysis was carried out, collecting data on access and retention on treatment from all 103 public and 38 private sector ART clinics in Malawi, using standardised patient master cards and clinic registers. Observations were censored on December 31(st) 2006. Independent data on mortality trends in army personnel from all causes between 2002 and 2006 were available from army records. By December 31(st) 2006, there were 85,168 patients ever started on ART in both public and private sectors, of whom 547 (0.7%) were army personnel. Of these, 22% started ART in WHO clinical stage 1 or 2 with a CD4-lymphocyte count of