Human immunodeficiency virus (HIV) infection is a major public health concern globally, especially in sub-Saharan African countries. Even though determining the incidence of treatment failure and its predictor is a crucial step to reduce the problem, there is limited information indicating the incidence and predictors of treatment failure among children in Ethiopia. Therefore, this study was conducted to assess the incidence and predictors of treatment failure among children on first-line antiretroviral therapy (ART) in Amhara Region referral hospitals, Northwest Ethiopia.An institution-based retrospective follow-up study was conducted from January 30, 2011, to January 30, 2018. A total of 402 children on first-line antiretroviral therapy were selected with a simple random sampling method in Amhara Region Referral Hospitals, Northwest Ethiopia. Data were extracted by reviewing patients’ ART intake and follow-up forms using pretested and structured checklists. The collected data were entered into Epidata Version 4.2 and analysis was done using STATA Version 13. Bivariable and multivariable Cox proportional hazards regression models were fitted to identify predictors of treatment failure.A total of 402 records of children on antiretroviral therapy (ART) were reviewed and treatment failures rate within the follow-up period were 12.19% . This study also found that the overall incidence density rate was 3.77% per 100 person-years observation. Virologic failure accounts 48.98% followed by immunologic (28.57%) and mixed failures (22.44%). Poor ART adherence , drug regimens, AZT-3TC-NVP , and AZT-3TC-EFV , Children whose both parent were died  and world health organization (WHO) clinical stage-4  were found to be predictors for treatment failure among children.The proportion of treatment failure among children on first-line ART in Amhara Region referral hospitals, Northwest Ethiopia was found to be high. Nearly half of the children experienced Virologic failure. Poor ART adherence, children whose parents`died without parents, WHO clinical stage-4 at baseline and type of regimen patients took were found to be predictors of first-line ART treatment failure. Therefore, expanding access to routine viral load, CD4 and clinical monitoring is mandatory to detect and early intervene of treatment failures’ to improve outcomes for children on ART. Patient caregivers or parents should strictly support children on medication adherence. Training to health professionals should be given time-based on revised guidelines, and follow up of treatment outcome should be monitored nationally to take the appropriate intervention. Acquired immune deficiency syndrome (AIDS) is a viral infection caused by the human immunodeficiency virus (HIV) that weakens the immune system and makes the body susceptible to secondary and opportunistic infections . The disAIDS develops very rapidly among Infants and young children living with HIV, have a high risk of poor outcomes, with up to 52% of children born with HIV dying before the age of 2 years and one third before year one in the absence of any intervention . There wEvidences showed that different predictors affect treatment failure. The most common predictors are age , 10–14, To reduce the problem of HIV/AIDS, ART was endorsed in 2003 and in January 2005, free ART was launched in Ethiopia , 23. HowWith the existence of this intervention, the burden of HIV/AIDS still continued to be high, and the national HIV prevalence is 1.16% in [specify the year here] . TherefoAn institution-based retrospective study was conducted to assess treatment failure and its predictors at Debre Markos and Felege Hiwot Referral hospitals in Amhara region, from 2011 to 2018. The health facilities were selected from five referral hospitals found in the region. Debre Markos Referral Hospital (DMRH) is found in Debre Markos town, Amhara national regional state Northwest Ethiopia. The town is located 299 Km northwest of Addis Ababa, the capital city of Ethiopia. The hospital provides service for an estimated of 2,397,876 total catchment populations. It started providing ART service with aid from the Regional Health Bureau and the Ministry of Health in April 2005 , 25. FelAll HIV infected children who were under 15 years of age, who initiated ART between January 30, 2011 and January 2018, and who took first line ART for at least six months were included in this study.’ Those children with incomplete medical record for important predictors were excluded from the study.The sample size was calculated by using the formula for survival analysis , E= after 6 months of effective treatment.Immunological failure: Persistent (at least 2 CD4 measurements) CD4 levels below 200cells/mm for children younger than 5 years and, CD4 levels below 100 cells/mm for older than 5 years.Virological failure: Viral load above 1000 copies/mL based on two consecutive viral load measurements in 3 months, with adherence support following the first viral load test.Censored: Individuals other than treatment failure .Adherence: The extent to which a client’s behavior coincides with the prescribed regimen as agreed upon through a shared decision making process between the client and the health care provider[ provider, evidencA structured English version checklist was developed and used for data extraction from the patients’ medical records on the Federal Ministry of Health Antiretroviral therapy (ART) follow up form and pediatrics HIV intake form. These records included; clients’ age, gender, weight, height, duration of follow up, clinical and laboratory data , the ART regimen and prophylaxis (Co-trimoxazole or Isoniazid), and adherence on a structured data retrieval form. Trained health professionals (nurses) working in the ART clinic were recruited as data collectors and supervisors.One day training was given for data collectors on data extraction system, data collection tools and objectives of the study. To assure the quality of data, the data collection checklist was pretested. After the pretest, necessary modification of data collection tool was made. Strict follow-up and supervision were carried out during data collection by principal investigators and feedback were given on a daily bases. The collected data was reviewed and checked for completeness before data entry.The collected data were entered using EpiData version 4.2 statistical software and exported to Stata version 13 statistical software for further analysis. The Kaplan Meier survival curve methods were used to estimate the time to treatment failure. The Log-rank test was used to compare survival curves between different categories of explanatory variables. The Cox proportional-hazards model was used to assess the predictors associated with treatment failure. To see the effect size of predictors on time to treatment failure, bivariable and multivariable Cox-proportional hazards regression model was fitted. Variables having p-value ≤ 0.25 in the bivariable analysis were fitted into multivariable analysis. Hazard Ratios (HR) with 95% confidence intervals was computed and a statistical significance was declared at 5% level (p<0.05). The necessary assumptions for Cox proportional hazard model were checked using Schoenfeld residuals test and graphically with log-log Cox adjusted survival estimate. The model fitness was checked using the Nelson- Aalen cumulative hazard rate relative to Cox-Snell residuals.Ethical clearance was obtained from Ethical review committee of Debre Markos University Health Science College. The ethics committee formally waived to extract patient data to address objectives aimed by this study. Then, permission was also obtained from the administrative bodies of each hospital  to access the ART client’s database and charts. Confidentiality was also maintained by not recording the identity that describes the individual and limiting the access of the data only for concerned bodies like principal investigator and advisors.After excluding 41 HIV positive children records due to incompleteness, we reviewed a total of 402 HIV positive children charts that were registered from January 30, 2011, to January 29, 2018. During the data extraction process, 41 HIV positive children medical record charts were excluded due to incomplete documentation and lack of important variables. Out of the included participants, slightly more than half, 225 (55.97%) were male and the mean age was 6.15 (SD = ± 3.48) years. One hundred ninety (47.26%) and 212 (52.74%) had a follow up at Debre Markos and Felege Hiwot Referral Hospital respectively. The majority, 329 (81.84%) of study participants were from the urban area and the mean age of caregivers was 37.58 (SD = ± 7.17) years. Most, 332(82.59%) of the children had a relationship with their parents as a caregiver. slightly more than half, 219 (54.48%) of both the mother and father of children were alive. Less than one third, 113 (28.83%) of the care giver were a government employee, and 242 (61.89%) were married .Clinically, one third, 129 (32.58%) of the study participants were categorized to WHO clinical stage 3 and the median CD4 cell count was 410cells (IQR 249–780). During ART follow-up, slightly more than two-thirds, 278 (69.33%) of children experienced opportunistic infection and the commonest, 71 (25.72%), was diarrhea. Most, 106 (72.1%) of the under-five children had appropriate developmental status while the majority, 222 (88.10%) of HIV positive children functional status were ambulatory when they start ART .One hundred forty (34.91%) of study participants were found on AZT-3TC-NVP ART regimen during ART initiation, and the majority, 362 (90.05%) of them had good ART adherence during their last follow-up period. Nearly one-third (32.8%) of the study participants had experience of regimen change during their ART follow-up time. Among the study participants, 49 (12.19%) had experienced ART side effects during their follow-up period, and mostly with AZT-3TC-NVP regimen, anemia being the commonest 15 (30.61%). Toxicity or side effects were the commonest 47 (35.6%) reason for regimen change. program shift from D4T to AZT or TDF, stock out of drugs and treatment failure were other reasons. During the follow–up time, majority 374 (93.27%) of the study participants did not take ART-prophylaxis, following birth from HIV infected mothers (PMTCT). Among the children on ART, 381 (94.78%) took Cotrimoxazole and 245 (61.25%) took on their follow-up time Isoniazid prophylaxis. The median ART clinic follow up period of participants was 35 months (IQR 18–62) months with minimum 6 months and maximum 84 months of follow-up, and nearly half, 205 (50.99%) of the participants were followed for less than 36 months. The majority, 334 (83.08%) of study participants were on ART follow-up at the time of the study, 8 (1.99%) were reported as lost, 10 (2.49%) dead, and 50 (12.44%) were transferred out .All study participants 402) who were followed for different periods in 7 years, a minimum of 6 months and a maximum of 84 months. The median survival time was 83 months with 1298.32 person-years follow up time. Forty-nine, 12.19%  of participants experienced treatment failure. Of this, 48.98%, 28.57% and 8.16% of the children have experienced a Virologic, Immunologic and mixed  respectively. In addition, a small proportion, 6.12%, 6.12% and 2.04% of them were mixed  and mixed  respectively. The overall incidence density rate (IDR) was 3.77  per 100 person-years of observations  times at a higher risk of treatment failure as compared with those whose both parents alive. In addition, children who were presented with WHO clinical stage 4 was 2.95  times at a higher risk of experiencing treatment failure than those who were on WHO clinical stage 1 and 2. Moreover, children who had poor ART adherence were 4.6  times at a higher risk of experiencing treatment failure than the counterpart. In addition, children who are on AZT-3TC-NVP treatment regimen were 5.2  times at a higher risk of treatment failure when compared with those who were on d4t-3TC-NVP. Being on AZT-3TC-EFV were also 6.26  times at higher risk of treatment failure as compared with those who were on d4t-3TC-NVP . TreatmeWhile antiretroviral therapy (ART) service is available worldwide, acquired immune deficiency syndrome (AIDS) was continuing to be a major global health priority problems especially in developing countries including Ethiopia , 3, 5. TThis study found that treatment failure among children was 12.19%  with 3.77 per 100 per-year incidence rates. The current finding is consistent with the finding reported at Jimma University Hospital 11.5%)  and at f.5%  and On the other hand, the incidence of treatment failure is found to be lower as compared with study findings reported by Gondar University Hospital (18.2%) , Fiche aThe current study also identified the predictor variable for treatment failure and found that AZT-3TC-NVP and AZT-3TC-EFV regimen, suboptimal adherence, advanced WHO clinical staging and dead parents of children were predictors of treatment failure.The current study found that children whose parents (both) died were predictors well the treatment failure. This is similar to a study conducted at Cambodia in HIV-Positive Children revealed that , 15. In AZT based regimen was found as a predictor for treatment failure by the current which is similar to a study done in Addis Ababa . In contSimilar to the current study advanced WHO clinical stage at baseline was a significant predictor of treatment failure, as studies show at Gondar University Hospital , Jimma UIn most studies both in our country and others show that low CD4 count at ART initiation and occurrence of severe opportunistic infection  were significant predictors of first-line ART treatment failure , 21, 29,Patients who had height for age in the third percentile or less at initiation of ART were found to have a higher probability of ART treatment failure  but in tSince the study was a retrospective study, it had its own limitation associated with poor documentation. Since the design is secondary, the study was unable to exhaustively explore all predictors variable that may have an effect on the treatment failure.In conclusion, the proportions of treatment failure among children on first-line ART at Northwest Amhara referral hospitals was found to be high and nearly half of the children experienced Virologic failure. Poor ART adherence, children without parents or died, WHO clinical stage IV at baseline and type of regimen patients took were found to be predictors of first-line ART treatment failure. Therefore, expanding access to routine viral load, CD4 and clinical monitoring is mandatory to detect and early intervene of treatment failures’ to improve outcomes for children on ART.Clinicians should provide targeted treatment approach with consideration of the risk of treatment failure  and proper documentation is necessary, since this study is depending on secondary data registered at each hospital, and the outcome relays on it. Expanding access to routine viral load, CD4 and clinical monitoring are mandatory to detect and early intervene of treatment failures’ to improve outcomes for children on ART. Patient caregivers or parents should strictly support children on medication adherence. Training to health professionals should be given time-based on revised guidelines, and follow up of treatment outcome should be monitored nationally to take the appropriate intervention.S1 File(SAV)Click here for additional data file.


Study participants were asked about their interest in switching to novel drug delivery systems that reduce the dosing frequency of antiretroviral regimens. Across a diverse, treatment-experienced cohort, we describe greatest interest in switching to an oral regimen taken once weekly, followed by injections taken every other month and twice-annual implants. The past decade has witnessed a dramatic increase in the number of effective, convenient, and well-tolerated HIV combination antiretroviral therapy (cART) regimens, and novel approaches that allow for significantly decreased dosing frequency are in active development , 2. SimpBarriers to access and incomplete adherence remain obstacles for effective cART and are associated with clinical and virologic failure, emergence of drug resistance, and increased risk of HIV transmission . CentersBetween February and August 2017, 263 adult persons living with HIV (PLWH) from the Infectious Diseases clinics at Duke University and the University of South Carolina were surveyed concerning their preferences for HIV treatment. Multiple recruitment methods, including flyers and invitation cards, referrals from providers, patients, and members of the community advisory board, and consecutive recruitment of patients after completion of their clinic appointments, were used to recruit samples that reflect the diversity of the treatment-experienced populations at the 2 clinics. Study activities were approved by the Institutional Review Boards of the Duke University Health System and the University of South Carolina. Written informed consent was obtained from participants at Duke University; oral informed consent was obtained from participants at the University of South Carolina.Trained research assistants administered in-person surveys on tablet devices at the Infectious Diseases clinics or nearby research offices. Participants were presented with the following:Several new HIV medicines are being developed that could be taken less frequently than currently available options. Compared with your current HIV medicines, how interested would you be in switching to a new treatment that is… a single pill once a week, 2 shots in clinic every other month, or implanting and removing 2 small plastic rods about the size of matchsticks in each forearm every 6 months.Responses were recorded on 5-point visual analog scales with 3 labels . Demographic characteristics, characteristics of patients’ current regimens, and patients’ treatment experience were documented.t tests were used to assess the significance of differences between the 2 clinic populations. Multivariate linear regression was used to jointly model patients’ interest in switching to the 3 advanced regimens as a function of sociodemographic characteristics , HIV treatment characteristics , and characteristics of their current regimen . The model was estimated in Stata 15 .Cohort characteristics were analyzed using descriptive statistics; chi-square statistics and Student The mean age of patients surveyed was 47 years, 56% were male, and 81% were of racial or ethnic minority; of those, 95% were African American. More than half of the patient population had a high school education (42%) or less (12%). On average, patients had been on cART for 12 years; the majority of patients (59%) reported that they were currently on a single-pill once-daily regimen. Integrase inhibitor based therapies (66%) were most common, followed by non-nucleoside and protease inhibitor based regimens . The majority (56%) of patients reported cART regimens with food restrictions. Most patients (82%) reported their most recent viral load to be <200 copies/mL; 22% indicated that they had missed at least 1 dose in the past 2 weeks. Current side effects were mentioned by 34% of participants; 39% reported long-term effects.P = .02), injections every other month (P < .001), and implants every 6 months (P = .002). Younger age (P = .036) and the experience of any long-term effects (P = .018) were associated with increased interest in switching to 2 shots every other month. Being on a 1-pill once-daily regimen was associated with lower interest in switching to a 1-pill once-weekly option (P = .046) and implants every 6 months (P = .014). Clinic site, gender, race/ethnicity, time on ART, a prior AIDS diagnosis, recent viral load, missed doses, experiencing current side effects, and current regimens with food restrictions were not associated with interest in switching to novel regimens.Forward-looking, patient-centered HIV care will require a greater understanding of patient preferences for emerging antiretroviral delivery systems. Which patients desire to switch, what the key motivators and perceived benefits are for switching, and ultimately which trade-offs patients are willing to make to effect switches are largely unknown. These data begin to address these issues. Across a diverse group of highly treatment-experienced PLWH, we describe greatest interest in switching to an oral regimen taken once weekly, followed by injections every other month; patients were least interested in 2 implants twice a year. It is plausible that, among the decreased-frequency dosing options presented, patients were more willing to switch to the administration modality that was most familiar, namely by pill. There was evidence of substantial preference heterogeneity; younger patients, those with higher education, those not taking a single-tablet regimen, and those who experienced at least 1 long-term effect expressed greater interest in 1 or more novel drug delivery systems.As patients become more aware of the availability of these treatment options, formation of preferences may change, especially as the attendant risks of the novel ART delivery options are better understood. In addition, Havlir et al. describe clinic-level implementation challenges for long-acting antivirals, namely prioritization of patient populations for preferred use, clinic infrastructure requirements, and provider and patient training programs . There aOur study has inherent limitations. First, the questions presented were part of a larger study that focused on HIV treatment preferences and shared decision-making; they were not designed to explore specific drivers of patients’ interest in switching to novel approaches. Other characteristics that may influence patients’ interest in switching to specific regimens, such as side effect profiles, long-term toxicities, and cost, could not be assessed. Second, our cohort was highly experienced and was not representative of the entire patient populations at the study sites. Although the 2 clinics in metropolitan areas in North and South Carolina are accessed by diverse patient groups, and the results did not differ between clinics, the findings may not be reflective of patient preferences in other geographic areas. More than 80% of study participants were from minority populations, and members of these populations may be particularly averse to newer medical interventions . PreviouInterest in longer-acting cART delivery systems varies widely among PLWH. Understanding preference heterogeneity for these novel treatment modalities may help to inform their development, predict uptake, and inform educational efforts to better engage patients in shared antiretroviral decision-making.Disclaimer. The views expressed here do not necessarily reflect the views of the Foundation.Financial support. This work was supported by the Robert Wood Johnson Foundation (grant #73056). Potential conflicts of interest. The authors declare that there is no conflict of interest regarding the publication of this article. All authors: no reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.


In an Essay, Mark Tenforde and colleagues advocate continued provision of baseline CD4 cell count testing in HIV care in low- and middle-income countries. Antiretroviral therapy (ART) has substantially decreased HIV morbidity and mortality in high-income as well as low- and middle-income countries (LMICs). Several randomized trials have demonstrated benefits from starting ART regardless of CD4 count  1–3]; t–3; t1–3]In the “treat all” era, the challenge is targeting effective prophylaxis against the bacterial, mycobacterial, and fungal pathogens that are the major causes of death in these late presenters. Such prophylaxis saves lives (see below) but also costs money and raises the potential risk of antimicrobial resistance if used indiscriminately. Further, when prophylaxis is provided conditional on a diagnostic test result, unless truly point-of-care, there is the potential for delays in ART initiation, which may substantially increase mortality risk.Specific tests that have evidence supporting their use in reducing mortality and are recommended to target treatment/prophylaxis to patients with HIV and low CD4 cell counts include tuberculosis (TB) smear, Xpert MTB/RIF and urinary lipoarabinomannan (LAM) for TB, and cryptococcal antigen (CrAg) to detect cryptococcal infection . TheoretCD4 has consistently been found to be a key predictor of mortality after ART initiation , strongeGiven the limited ability of clinical assessment alone to identify patients with advanced HIV disease and the evidence for mortality benefit of enhanced OI screening in patients with advanced disease , we suggCD4 testing remains important for staging HIV pre-ART, as it is the only way to identify the population at highest risk of death. However, even where immediate CD4 testing is unavailable, particularly in sub-Saharan Africa (SSA), all algorithms should still begin with intensive case finding for TB. Same-day screening with either smear or Xpert MTB/RIF should ideally be performed, but absence of such testing should not be a barrier to prompt ART initiation with isoniazid and cotrimoxazole (preferably as a fixed-dose-combination with pyridoxine ) when clThe TEMPRANO trial, which had a predominance of patients with early HIV (CD4 > 350 cells/μL), demonstrated the added benefit of 6 months’ isoniazid prophylaxis to ART (35% reduction in death or severe HIV-related illness), as did the REMEMBER trial in participants with advanced HIV disease (CD4 < 50 cells/μL); in both trials, mortality rates in the isoniazid treatment arm were lower than historical controls ,9. Of noSeveral assays (including Alere’s PIMA and Becton Dickinson’s FACSPresto) are commercially available for rapid CD4 testing. If rapid CD4 testing is available and results are <100 cells/μL, where other tests are not available, low-dose (100 mg/kg) fluconazole and a 5-day course of azithromycin (500 mg/day) along with isoniazid/cotrimoxazole/pyridoxine (as a late-presenters “package”) decreased mortality risk by 27% (3% absolute risk reduction) in the REALITY trial [Where rapid CD4 testing is available and additional tests for OIs can be performed, the challenge is balancing the excess mortality risks associated with potential delays to ART while waiting for test results against benefits of targeting prophylaxis more narrowly. In hospitalized patients with advanced HIV, urinary LAM testing followed by 4-drug anti-TB treatment for positives decreased 6-month mortality ,17. LAM Immunosuppressed patients also benefit from CrAg screening and preemptive therapy with fluconazole if positive, when bundled with ART initiation. The REMSTART trial, which included participants with CD4 < 200 cells/μL, found that CrAg screening coupled with community-based ART support decreased mortality by 28% (4% absolute reduction) . The IMMp = 0.03) in patients receiving enhanced prophylaxis including azithromycin; this difference may have been due in part to reductions in bacterial sepsis. Among those dying from unknown causes, baseline CrAg prevalence was very low, suggesting that these were not due to undiagnosed cryptococcosis [Based on the REALITY trial , 5 days coccosis .Baseline CD4 testing is still important to guide the diagnostic evaluation for common OIs and prophylaxis in patients newly diagnosed with HIV. Conventional laboratory-based CD4 testing using flow cytometry requires substantial infrastructure and technical expertise and may lead to significant delays in obtaining actionable results, with a systematic review estimating a mean time of 10.5 days from the time a CD4 test was conducted to the time the result was received, versus 0.1 days with point-of-care CD4 testing . SeveralBaseline CD4 is an essential part of HIV care, and implementation research is needed to better streamline this and other new point-of-care tests for OIs to make them practical to perform in underresourced centers. We present a resource-based public health approach according to diagnostic test availability that could decrease early mortality after ART initiation and would be practical to implement. Our approach does not allow “the best to be the enemy of the good.” Even the most resource-constrained settings can immediately implement interventions that have the potential to save thousands of lives, and further refinement can be offered in settings where rapid screening for common OIs is feasible. An optimal approach requires that pre-ART CD4 (preferably as a simple point-of-care threshold test) continues to be available. We believe this provides a pragmatic algorithm to avoid delaying ART for the most immunosuppressed patients who are at the highest risk of dying.


We described acquired drug resistance mutations in gag, pol, vif, vpr, tat, vpu, and nef genes between paired baseline and month 6 samples.The outcome of tenofovir (TDF)- and zidovudine (ZDV)-based ART was defined in 874 ART naïve patients using the on-treatment (OT) and intention-to-treat (ITT) analyses. Genotypic resistance testing was done in patients failing ART (> 1000 copies/ml) at month 6 and 12. Near full-length genome sequencing (NFLG) was used to assess amino acid changes in HIV-1 High failure rates were found in ITT analysis at month 6 and 12 . Major nucleoside and non-nucleoside reverse transcriptase (NRTI/NNRTI) drug resistance mutations were detected in most failure patients at month 6  and month 12 . A high rate of K65R was identified only in TDF treated patients . No significant difference was found in failure rate or extent of HIVDR between TDF- and ZDV- treated patients. All target regions of interest for HIVDR were described by NFLG in 16 patients tested before initiation of ART and at month 6.In this first Ethiopian national cohort, a high degree of HIVDR was seen among ART failure patients, independent on whether TDF- or ZDV was given. However, the major reason to ART failure was lost-to-follow-up rather than virologic failure. Our NFLG assay covered all relevant target genes for antiretrovirals and is an attractive alternative for HIVDR surveillance.The online version of this article (10.1186/s12879-019-4196-8) contains supplementary material, which is available to authorized users. In Ethiopia, over 700,000 people are currently estimated to live with human immunodeficiency virus (HIV), corresponding to an adult prevalence of 1.15% , 3; the Only a few studies with relatively small number of patients from limited geographical regions in Ethiopia have reported ART failure rates, including acquired HIV drug resistance (HIVDR) –12. Althn = 389), TDF + 3TC + NVP (n = 78), ZDV + 3TC + EFV (n = 104), ZDV + 3TC + NVP (n = 258), stavudine (d4T) + 3TC + EFV (n = 23), d4T + 3TC+ NVP (n = 21), and abacavir (ABC) + 3TC + EFV (n = 1). Altogether TDF was given to 467 (53.4%), ZDV to 362 (41.4%), d4T to 44 (5%) and ABC to one patient  of ART in Ethiopia from seven university hospitals during 2009–2011 as we described elsewhere , 14. As The treatment outcomes at month 6 and 12 were determined by on-treatment (OT) and intention-to-treat (ITT) analyses. Two categories of virological treatment failures were defined as i) > 150 copies/ml (limit of detection of the assay); ii) > 1000 copies/ml (as per WHO definition). For ITT, treatment failure was defined as failure to attain viral suppression (as described for OT) or lost-to-follow-up (LTFU) including confirmed death, moved from study sites or similar reasons.Clinical, routine laboratory and CD4 T-cells were analysed at the study sites [http://www.mbio.ncsu.edu/bioedit/bioedit.html). Acquired DRM were identified by the Stanford HIVdb Program (hivdb.stanford.edu). Genotypic drug resistance defined as the presence of ≥1 major amino acid substitution included in the 2015 International Antiviral Society–USA (IAS) mutation list [PBSS was attempted on patients with viral load ≥1000 copies/ml at month 6 in 51 subjects and month 12 in 33 patients . In brieion list  and the ET sequence (U46016) described by our group was used as a reference [in-house Perl script that recognized the nucleotide changes from the reference sequence and created a corresponding number code as per HXB2 coordinates (790 to 9417). The resulting matrix was plotted using the TraMineR package [NFLG sequencing was performed on plasma from baseline and month 6 of 16 randomly selected patients among virologic failure patients with VL > 1000 copies/ml at month 6 of whom 12 were given TDF and four ZDV  for prediction of tropism throughout the analysis [Using AliView ver 1.17.1 and BioEdit ver 7.2.6.1 softwares, we aligned nucleotides and amino acids generated for each gene from the paired samples and described the specific amino acid mutations, which had appeared at month 6. The protein alignments were manually reviewed to identify changed residues. As European guideline recommended we have used the Geno2pheno tools at FPR 10% cut-off .Descriptive statistics  were used to summarize sociodemographic, clinical and virological parameters. Treatment outcomes were compared between patients with different NRTI regimens by Chi-square or Fisher’s exact test. The prevalence and type of DRM were compared between patients with TDF- or ZDV-based regimens by Chi-square or Fisher’s exact test. Among TDF treated patients, OT analysis identified virologic failure in 52/350 (15.0%) and 34/350 (9.7%) at month 6 using the 150 copies/ml and 1000 copies/ml cut-off values, respectively. ITT analysis identified treatment failure in 129/426 (30.2%) and 111/426 (26.0%) patients, respectively  and 19/219 (8.7%) patients using the 150 copies/ml and 1000 copies/ml cut-off values, respectively. In ITT analysis treatment failure was found in 133/322 (41.3%) and 122/322 (37.9%) patients, respectively (Table n = 24 (66.7%); only NRTI: n = 2 (5.6%); only NNRTI: n = 10 (27.8%) , 15/19 (78.9%), and 4/4 (100%) of the TDF-, ZDV-, d4T-failing patient samples with VL > 1000 copies/ml, respectively (Table n = 16 (80.0%); only NRTI: n = 1 (5.0%); only NNRTI: n = 3 (15.0%). No significant difference was found in viral load at month 6 or month 12 for patients failing TDF with or without DRM and for patients failing ZDV with or without DRM. Thirteen of 16 (81.3%) TDF failure patients had DRM . K65R was found in eight (50%) patients and TAM-2 in five patients. In ZDV patients, 5/10 (50%) had DRM . With regard to the number of DRM in sequences harboring such mutations, no difference was found between patients failing TDF or ZDV at month 6. However, at month 12 sequences harboring mutations from patients failing TDF had more DRM as compared to those failing ZDV (p = 0.017), NNRTI DRM (p = 0.037), and NRTI DRM (p = 0.040).At month 12, a total of 30 sequences were obtained from 16/17 (94.1%), 10/12 (83.3%) and 4/4 (100%) of the TDF-, ZDV-, d4T-failing patients, respectively. One to eight (median: four) major NRTI and/or NNRTI DRM were found in 20 (66.7%) samples: NRTI+NNRTI: gag, pol, vif, vpr, tat, vpu, and nef were successfully generated in all 32 (16 paired) samples, except for the nef gene at month 6. Maximum likelihood phylogenetic analysis revealed proper matching of the paired NFLG sequences with 100% bootstrap support  DRM.NFLG sequences including n all 32 6 paired pol . The predicted sensitivities to NRTI and NNRTI are given on Fig. After six months of ART, eleven (68.8%) patients had acquired one to five (median: three) major DRM in gag, amino acid changes mostly clustered in the p6 and p17 regions. In the Gag-p6 region, A5G/E/P/P5S/A/R5A and K6S/P6A/S6K/E6K/V6E mutations were displayed in seven and five samples, respectively. PYKE tetra-peptide motif was found both at baseline and month 6 in nine (56.3%) of the 16 sample pairs on the C-terminal position of the p6 region. Duplicate tetra-peptide motif PTAP was identified in two samples at baseline, where the respective month 6 samples had single motif (deletion of one of the double motifs). A triple PTAP tetra-peptide motif was found in paired samples of a patient. The remaining 13 patients (81.3%) had only single P(T/S)AP motif in their paired samples. In the C-terminal of p17, 14 (87.5%) of the 16 patients displayed the R4S mutation. On the other hand, only one patient had a mutation (T375 M) in the p2/NC gag cleavage site.For vif  and in the nef gene . The tat, vpu and vpr showed very few amino acid changes . The Geno2PhenoOf the 16 baseline samples 15 (93.8%) displayed three nuclear factor kappa B (NF-kB) each in the enhancer region of the LTR of HIV-1C and the remaining one sample shows two NF-kB (which has short nucleotide sequence). Again 15 of the 16 (93.8%) month 6 samples displayed triple NF-kB, but one sample displayed large insertions in the LTR and showed four NF-kB instead.In this first large countrywide study of ART outcome in Ethiopia, a high failure rate was identified in the ITT-analysis; around one-fourth at month 6 and one-third at month 12, whereas the OT-analysis revealed less than 10% of the participants failed virologically (> 1000 copies/ml) at month 6 and 12. Viral load is a gold standard for monitoring ART response and is a marker of the treatment outcome . HoweverThis study confirms that early death and LTFU are major reasons to poor treatment outcome of ART in Ethiopia, as described from other sSA countries , althougIn the WHO recommendations, TDF has since several years replaced thymidine analogues (ZDV and stavudine) in first-line regimens. As shown in our study, the introduction of TDF without virological monitoring may result in an extensive evolution of the K65R mutation, especially since it is preferentially selected by HIV-1C in ex vivo and in vitro analysis , 33. Impp < 0.05). In contrary to our observation, studies from resource-limited settings revealed that TDF-based regimens performed better than d4T, most notably with a significantly higher rate of LTFU for d4T patients [Because of its well-recognized toxicities and as per the WHO recommendation , 35, Ethpatients .Retention in care was low and undocumented mortality and self-transfer of patients are likely to have contributed. A possible way forward to improve treatment outcome could possibly be the use of long-acting drugs, such as the integrase inhibitor cabotegravir, which inhibits HIV-1C at least as efficient as HIV-1B , and theenv and the LTR in the majority of patients. In addition, non-drug target sites like Gag and gp41 were sequenced which also can affect the drug efficiency [gag region may influence the efficacy of PI which is second line treatment option in Ethiopia. Gag mutations were found by NFLG in the majority of our patients at failure although the identified point mutations are not known to influence the response to PI. A PYKE tetra-peptide, which was found in the ALIX-binding motif of Gag-p6 at baseline in all patients, remained unaffected by the treatment in all subjects. In an earlier study, this tetra-peptide motif was observed among half of treatment naïve Ethiopian patients, but the status was not known among ART experienced patients [Our HIV-NFLG sequencing assay was used to study the amino acid changes between paired samples from baseline and month 6 of virologic failure patients at several genes. It was found to efficiently amplify key HIV-1 drug target sites  in all 32 tested samples, and the ficiency , 41. Thupatients . In contChanges of the motif PTAP in Gag-p6 were also seen. Thus, a duplicate of the motif was found in baseline samples of two patients of which one motif was deleted from the virus of each patient at month 6. Also, a triple PTAP motif was detected in paired samples of one patient, which has not been described earlier. Subtype specific differences have earlier been observed in Gag-p6 with regard to the motif PTAP. In addition, a difference in ART outcome in relation to duplication of the PTAP motifs for HIV-1C has been reported . The dupIn the present study we have used our NFLG assay to analyse HIV drug resistance in an Ethiopian population and all relevant target regions for HIVDR were described. Although HIV resistance testing is presently not used clinically in most low- and middle income countries (LMIC), the method is an alternative for surveillance of HIV drug resistance. Earlier we have shown the negative impact of pretreatment drug resistance mutations on virologic outcome in our Ethiopian cohort . Also, igag gene, no common pattern of amino acid changes was seen in the other genes, with the exception of known RTI DRM in the pol gene. However, in all of the NFLG sequences we identified three NF-kB binding regions in the LTR which is unique for HIV-1C and described by us earlier [It can also be noted that resistance to dolutegravir has been described outside the integrase, mainly in Nef and LTR overlapped region, in vitro and in patients failing monotherapy with dolutegravir . In cont earlier . AltogetIn conclusion, although the retention in care was low in this first countrywide Ethiopian cohort no differences was found between patients given TDF- or ZDV-based regimens with regard to treatment outcome or level of drug resistance. A broad RT-inhibitor resistance was found in three quarters of the patients who were on-treatment with virologic failure. Our NLFG assay was shown to efficiently amplify genes of known or potential relevance for HIV drug resistance and is an attractive alternative for such surveillance in low and middle income countries.Additional file 1:Table S1. Drug resistance mutations (DRM) associated with reverse transcriptase inhibitors in patients failing at month 6 and/or 12 by the population-based Sanger sequencing (PBSS) assay. (DOCX 18 kb)


Sub-Saharan Africa (SSA) alone has nine out of every 10 children living with HIV globally and monitoring in this setting remains suboptimal, even as these children grow older. With scalability of antiretroviral therapy (ART), several HIV-infected children are growing towards adolescence (over 2.1 million), with the potentials to reach adulthood. However, despite an overall reduction in HIV-related mortality, there are increasing deaths among adolescents living with HIV (ADLHIV), with limited evidence for improved policy-making. Of note, strategies for adolescent transition from pediatrics to adult-healthcare are critical to ensure successful treatment response and longer life expectancy. Interestingly, with uptakes in prevention of mother-to-child transmission, challenges in ART programs, and high viremia among children in SSA, the success rate of paediatric ART might be quickly jeopardised, with possible HIV-1 drug-resistance (HIVDR) emergence, especially after years of paediatric ART exposure. Therefore, monitoring ART response in adolescents and evaluating HIVDR patterns might limit disease progression and guide on subsequent ART options for SSA ADLHIV.Among Cameroonian ADLHIV receiving ART, we shall evaluate the rate of immunovirologic failure, acquired HIVDR-associated mutations, HIV-1 subtype distribution, genetic variability in circulating (plasma) versus archived (cellular) viral strains, and HIVDR early warning indicators (EWIs) at different time-points.A prospective and observational study will be conducted among 250 ADLHIV (10–19 years old) receiving ART in the centre region of Cameroon, and followed-up at 6 and 12 months after enrollment. Following consecutive sampling at enrolment, plasma viral load and CD4/CD8 count will be measured, and genotypic resistance testing (GRT) will be performed both in plasma and in buffy coat for participants experiencing virological failure (two consecutive viremia > = 1000 copies/ml). Plasma viral load and CD4/CD8 will be monitored for all participants at 6 and 12 months after enrolment. HIVDR-EWIs will be monitored and survival analysis performed during the 12 months follow-up. Primary outcomes are rates of virological failure, acquired-HIVDR, and mortality.Our findings will provide evidence-based recommendations to ensure successful transition from paediatrics to adult ART regimens and highlight further needs of active ART combinations, for reduced morbidity and mortality in populations of ADLHIV within SSA.The online version of this article (10.1186/s12887-019-1599-z) contains supplementary material, which is available to authorized users. Sub-Saharan Africa (SSA) alone carries 69.9% of the 36.9 million people infected worldwide and 65.8% of the 1.2 million the global AIDS-related deaths by end 2014, the majority occurring among adolescents ) after enrolment into the study. At each time-point , epidemiological and clinical data will be collected using the same procedure described above during enrolment (i.e. baseline), including ART-adherence assessment. 6 ml of intravenous blood will be collected per visit (for CD4/CD8 and PVL). Among those experiencing VF at these visits, GRT will be performed as described above , 36, 43,At each study phase , residual samples will be stored at − 80° available in the blood bank of the CIRCB. The entire work plan requires 30-months: 6-months (month1–6) for administration/ethics/field preparedness, 6-months (Month 7–12) for enrolment of participants until complete sampling, 12-months monitoring (Month 7–24) for complete follow-up of one-year, and 6-months (Month 25–30) for closing study sites, data processing, and reporting.At month-0, enrolment will be done at study clinic for all adolescents meeting study eligibility criteria. Participants will then be followed up during 12 months after enrolment, with scheduled visits at the clinic at months 6 (±1 month) and 12 (±2 month), for a total of 3 scheduled visits. Patients with virological failure  at any timepoint will undergo an extra visit in the next 1 month with adherence support and to confirm the virological failure (VF) as aforedescribed. Participants’ clinical management not considered in the study protocol will follow local recommendations for clinical practice at the discretion of the responsible physician.Procedures at Baseline visit (month 0). Mothers will be required to provide data contained in the Baseline Case Report Form . Briefly, this will cover detailed epidemiological and clinical information, including socio-demographic and economic data, HIV history, history of antiretroviral usage by the adolescent, including drugs taken as part of PMTCT (if applicable), adherence to ART, and results from any laboratory test that were performed before study enrolment and available in the patient medical record file. Blood will be collected from each participating adolsecent, for a total of volumen 6 ml per timepoint for the following: determination of: viral load (600 μl plasma); CD4/CD8 levels (100 μl whole blood); HIV-1 Sanger sequencing for protease-reverse transcriptase (1000 μl plasma for RNA extraction and 200 μl buffy coat for DNA extraction); residual plasma samples will be stored at the biobank.Laboratory procedures on sample management plans are provided below. Procedures at follow-up visits at months 6 and 12. Participants attending follow-up and their legal gauardians will be required to provide data contained in the follow-up Case Report Form, for updated information on epidemiological and clinical situation, with special emphasis on antiretroviral treatment information and HIV-related clinical events. Blood samples will be collected as described above. Similarly, for plasma viral load > 1000 copies/ml, adherence support will be provide then GRT performed if and only if virological failure is confirmed one-month later.All laboratory analysis will all be performed at the International Reference Centre Chantal Biya (CIRCB) in Yaounde, Cameroon, among which: Viral load measurementsWill be performed using Abbott m2000rt, as per the manufacturer’s instructions. Briefly, 600 μl of plasma collected from EDTA tube is used for viral RNA extraction. Based on the principle of reverse transcription polymerization (RT-PCR), amplicons are generated from the RNA genome of HIV-1 in clinical specimens. An RNA sequence that is unrelated to the HIV-1 target sequence is introduced into each sample at the beginning of sample PCR mix preparation. This unrelated RNA sequence is simultaneously amplified by RT-PCR, and serves as an internal control (IC) to demonstrate that the process has proceeded correctly for each sample. The amount of HIV-1 target sequence that is present at each amplification cycle is measured using fluorescent-labeled oligonucleotide probes on the Abbott m2000rt instrument. The amplification cycle at which an increase in fluorescent signal is detected by the Abbott m2000rt is proportional to the log of the HIV-1 RNA concentration present in the original patient sample.CD4/CD8 will be performed using FACSCalibur, as per the manufacturer’s instructions. Briefly, determination of lymphocyte subsets  will be performed on the flow cytometer  using a single-platform technology. A volume of 50 μl whole blood will be stained with 10 μl of Multitest reagent in Trucount tubes for 15 min , and red blood cells will be lysed by using fluorescence-activated cell sorting lysing solution (BD Biosciences). Data acquisition will be performed using CellQuest-Pro software (BD Biosciences) creating up to nine multicolor acquisition dot plots that use eight multicolor gates. Data will be then analyzed with FlowJo software version 7.5.5 for Microsoft . Using Multitest reagents with Trucount tubes in the laboratory procedure, absolute subset count will be calculated with the following formula: (number of events in the region containing the cell population/number of events in the absolute count bead region) × (the number of beads per tube/test volume). If ncessary, a FACSCanto II  could be used for the same CD4/CD8 measurement purposes. Briefly, these analyses will be performed using the FACS Diva software , by reading information from the BD FACSTM 7-color setup beads label into BD FACSCanto clinical software (2D barcode symbology), and also by reading coded patient information into a worklist. Sanger Sequencing for HIV-1 drug resistance testing will be performed for adolescent experiencing virological faiure, using ABI 3130xl (Genetic Anlyser with 16 capillaries), HIV genotyping will be performed using plasma samples and an in-house reverse transcriptase polymerase chain reaction (PCR) protocol, as previously described . Briefly, RNA will be extracted from 140 μl of concentrated plasma using the QIAamp Viral RNA minikit , according to the manufacturer’s protocol. RNA extracts will then be retrotranscribed and amplified using the kit One-Step Invitrogen  (SuperScript One-Step for long templates RT-PCR) and 2 different sequence-specific primers for 40 cycles. For insufficiently amplified samples after the first round PCR, a second round PCR (semi-nested PCR) will be performed. A direct sequencing reaction will be done using 7–8 overlapping primers. For a full plate run, the sequencer of 16 capillaries will analyse 2 different patient samples simultaneously for ~ 2.5 h per run, and assembly program (Seqscape v2.7) will be used for sequence analysis, and mutations will be interpreted following the Stanford HIVdb algorithm. Therefore, 12 samples will run for 15 h in total. Study timeline is provided as Additional file 1 (on-time pill pick-up), EWI2 (retention in care), EWI3 (pharmacy stock-outs), EWI4 (dispensing practices), EWI5 . Among patients not retained in care during the 12-month follow-up period, we shall investigate to the underpinning reason that might be deliberate ART interruption, transfer out or mortality. Cases of mortality will then serve to determine the survival rate among the study participants.Data on HIVDR EWIs will also be collected at each study phase, consisting of five variables among which EWIVF will be defined as two consecutive PVL ≥ 1000 copies/ml within 1 month under intensive adherence counselling and support [Immunological failure will be defined as persistent CD4 < 100 cells/mm  during fClinical failure will be defined as WHO clinical stage 3 or 4 events or death. Clinical management outside the study protocol will be done according to local national guidelines.Stanford HIVdb algorithm [HIVDR will be defined as the presence of any mutation, in the PR-RT regions of HIV-1, known to reduce the efficacy of NRTI, NNRTI or protease inhibitor, as per the lgorithm , 36, 43.Good adherence will be defined as ≥95% compliance to ART (i.e. only one missing dose), measured by pill count and by self-reported, during the past 30 days.EWIs will be defined and their respective performance targets are provided below: , 3, 8For each of the 3 study phases , quality assurance will be performed by proficiency testing for PVL, CD4/CD8 (cell/mm [Lost to follow-up (LTFU) is a potential risk in reducing the power of this study for evidence-based recommendations. To limit LTFU, adherence support will be intensified and the site community relay agents will undertake active search of defaulters.Investigators’ compliance may be concerning at certain points of the study . To handle this potential challenge, site visit will be conducted weekly and site compliance to study regulations will be assessed by monthly supervision. The lead applicant will regularly coordinate activities with study-site clinicians and with guidance from mentor.Data protection and safety might be concerning. This will be handle by procurement of a new computer device and a specific database dedicated exclusively for the project, with encrypted password and a protected working station. Potential bias in laboratory data will be handled with quality assurance. To ensure functionality of equipment, routine preventive maintenance will be ensured. If for any unforeseen reason an equipment breakdown, the backup laboratory (Centre Pasteur du Cameroun) will covered the required period, under current agreement. For staff protection, biosafety will be ensured by use of personal protective equipment as per international standards, including use of for a biosafety level II cabinet for project related-laboratory activities.Data will be collected per site and stored in a password-protected database at the CIRCB, Yaoundé-Cameroon, whereby the project statistician will coordinate data management and analyses using SPSS software. Study data will consist of sociodemographic, clinical and laboratory information. Data Management will be under the responsibility of the project Biostatistician of the CIRCB in Younde-Cameroon, in coordination with the principal investigator. Data management will therefore entail procedures for sociodemographic, clinical and laboratory data collection, data protection, data quality assurance and data analysis. This procedure will be similar at enrolment (M0), midpoint (M6) and endpoint (M12). Standardized Case Report Forms (CRFs) will be generated and provided to the four study sites for on-site collection of data. In addition to basic demographic information, CRFs will include all clinical information, as well as information on type of sample collected, including date, number of aliquots, volume of aliquots and sample identification numbers to reassure further tracing of laboratory results. CRFs will be translated and provided in French and in English as per national regulations in Cameroon. However, mainly the French version will be used as the study sites are all within the French-speaking sector of the country. CRFs will be completed by the project site collaborator, and double-checked by a research fellow for consistency, if necessary, the CRF will be correctly completed and signed by the site principal investigator or other authorised person. At the end of each study visit, double-checked and signed CRFs will be sent to data entry. Laboratory CRFs will be also generated and provided in paper to the corresponding local laboratories to introduce laboratory results, together with identification number of samples. At the completion of laboratory procedures, laboratory project staff will be responsible for completion of laboratory CRFs, which will be double-checked by the co-investigator for laboratory analyses and, if necessary, the CRF will be correctly completed and signed by the site principal investigator or other authorised person. At the end of this process, double-checked and signed laboratory CRFs will be sent to data entry.Data collected through clinical and laboratory CRFs will be entered on site through double entry, by designated data entry personnel. A specific database will be created for the protected entry of clinical data and laboratory results. Data storage will be done in numerical codes generated consecutively per site. Samples for plasma viral load, CD4/CD8, and eventually for genotypic resistance testing (in case of treatment failure) to be daily centralised for analysis at the CIRCB laboratory, using participant anonymous ID code for their respective sample, alongside the corresponding sample transportation form. The laboratory co-investigator at CIRCB or other authorized person will have full responsibility for the completion of any procedures.At enrolment, midpoint and endpoint, prevalence of VF and HIVDR  will be described globally, by geographical settings and by site. Associated factors will be evaluated using multivariate logistic regression, with an estimative approach for the unbiased effect of ART regimens (first versus second-line) and HIV-1 diversity (CRF02_AG versus non-CRF02-AG) . In ordeOur research objectives will help in generating evidence for optimal monitoring and management of adolescents treated with antiretroviral therapy (ART) in a sub-Saharan African (SSA) context. Specifically, study objective 1 will help in understanding the significance of treatment failure among adolescents living with HIV (ADLHIV) in a resource-limited setting (RLS), will provide relative time-estimates for treatment failure, and propose locally affordable measures to limit ART failure in ADLHIV; study objective 2 will provide the burden of drug resistance according to the national treatment guidelines, and would inform on the adequacy between detected mutations and the most potentially active second- or third-line drugs for ADLHIV in such RLS. This study objective is inform if the development of HIVDR are specific to the different local HIV-1 clades, for possible clinical considerations for a specific management of adolescents in the country; study objective 3 will inform on the potential compartmentalisation of HIVDR among adolescents, in order to design drug combinations that provide maximal efficacy in a long run; study objective 4 will identify both programmatic factors favouring HIVDR emergence among ADLHIV, among which events of ARV stock outs and the quality of dispensing practices. This same objective will also identify the level of adherence of ADLHIV to ART, among which non-observance, delay drug pick-up and lost to follow-up. Importantly, with the exception of settings where HIV-1 subtype C prevails (found mainly in Southern Africa), our study impacts could be generalizable to the population of ADLHIV in several RLS, giving potential endorsements by national and international health regulatory bodies such the World Health Organisation (WHO) and UNAIDS.Additional file 1:Supplementary digital contents (SDC). SDC 1: Overall summary of major timeline and activities. (DOCX 29 kb)


However, up to date, there are no genetic profiling data for the resistance-causing mutations in HIV-1 virus in patients on antiretroviral drugs therapy. This paper presents an initial report and a profiling survey of drug resistance-associated mutations of This is a retrospective study on Patients treated at KFSH&RC since 2003 up to 2016. The analysis was done on the drug resistance mutations profiles of 103 patients who were undergoing highly active antiretroviral therapy protocols.Our analysis shows that the drug resistance mutations reported in our treatment cohort of HIV-infected adults patients is similar what is internationally reported to some extent. Additionally, we have identified novel drug resistance causing mutations. Furthermore, different profile of drug resistance causing mutations was also observed.Patients showed both similar and new drug resistant causing mutations, early identification of these mutations is crucial to guide and avoid failure future therapy. Antiretroviral drugs (ARV) are the group of drugs used for patients diagnosed with HIV infection. These antiretroviral are usually administered in combinations to reduce the emergence of HIV-1 drug resistance. Nonetheless, the acquisition and evolution of secondary resistance, from non-compliance is one of the major obstacles to a successful ARV therapy. Furthermore, Patients who acquire HIV-1 drug resistant viruses have fewer treatment options and are at a higher risk of morbidity [The US Food and drug administration since 2012 approved 28 ARV drugs . These AIn general, the World Health Organization (WHO) recommended a protocol for the initial treatment of HIV-1 infection that generally includes the combined use of three antiretroviral agents. In most cases, the application of two NRTI together with a NNRTI, a PI or an II is recommended . To insuThe development of resistance to ARV therapy remains a major cause of treatment failure among patients living with HIV-1. Additionally, it’s also as important to measure the rates of these mutations in our populations. This will improve our understanding of resistance and lead to more refined treatment strategies and, in some cases, improves the outcome –9.103 HIV-1 patients seen at KFSH&RC. Identifying these resistance causing mutations will guide us to establish an in house diagnostic assay for screening of these possible DRM as a diagnostic tool for the HIV clinic for better patient care and treatment.Antiretroviral drugs have been available at KFSH&RC, a referral hospital, in Riyadh, the capital of Saudi Arabia, for the treatment of adults and children infected with HIV-1 since the early 90’s. However, no genetic profiling of the resistance-causing mutations in the HIV-1 virus obtained from patients on ARV therapy protocols have been done nor analyzed. This paper presents an initial report and a profiling survey of drug resistance-associated mutations (DRMs) of A retrospective analysis was done on the drug resistance mutations profiles of 103 patients at KFSH&RC, who were undergoing highly active antiretroviral therapy (HAART) protocols. Our selection criteria were all HIV-infected adults on HAART regimens and are experiencing virological failure (> 500 copies/mL), with resistance mutations detected by consensus sequencing. Up to date there are no commercially available genotyping test kits to monitor such mutation. Since 2003, these tests were done at our reference laboratory in Mayo clinic. Testing was performed by RT-PCR and DNA sequencing method . Results were based on manufacturer’s most recent FDA-approved interpretive guidelines. HIV-1 genotype was analyzed by TRUGENE HIV-1 Genotyping Kit , and interpretive results were based on manufacturer’s Guidelines v8.0. The evaluation for DRM was at the earliest available testing done and diagnosed within 1 year of their estimated date of infection at KFSH&RC. The mutations reported and their profile for each ARV group of drugs was analyzed. The comparison was done for single drugs in addition to known combination therapy. Additionally, the percentage of primary drug resistance causing mutations reported annually was also analyzed.. Their age ranged from 9 to 78 with a mean of 45 years. The number of males was 70 (68%) and females 33 (32%), with a ratio of male to female 2:1.This study includes 103 HIV-1-infected Saudi patients enrolled between the years 1988–2016The median of CD4 count at diagnosis was 16.8 cells/μl, with an average viral load of 5.3 log10/ml. The average time (years) since diagnosis of these patients to initiation of HAART protocol 1.78 years, with an average therapy time of 14 .1 years.All these patients had drug resistance analysis done on their viral samples, with an average of 5.9 years after the initiation of ART. The drug resistance testing for these patients was performed more than once, ranging from 1 to 12 times with a mean of 3 times in all their therapy years. Table Mutations were analyzed in relation to demographic, clinical, and laboratory variables at time of genotyping. The drug resistance is evaluated through genotyping tests. One hundred and three subjects were analyzed for mutation codon analysis.The highest frequency for drug resistance in our cohort studies was reported to the NRTI’s (68%) followed by (47%) for both NNRTI’s and PRI’s, as seen in Fig. Overall the K103 N was the main and most common mutation detected within the NNRTIs group of drugs. Other Mutations at positions; 98, 100, 101, 103, 106, 108, 138, 179, 181, 188, 190, 225, 227 and 230 have also been detected. Interestingly, mutation I88H, K101Q, E135G, V179E and F227 L are novel mutations reported with our patients’ cohort which were never reported previously. Furthermore, mutations V106A, V108I, Y181C, Y88L, G190A and P225H have been reported in our patients. Figure Resistance to NRTI’s especially AZT was found to be induced by a range of mutations in RT including M41 L, K65R/E/N, D67N, K70R, I74V, I84V, M184 L, L210 W, T215Y/F, and K219E which were all present in our patients. Additionally, a number of resistance causing novel mutations were also observed, including, T215C/D/S/V. Figure The use of PI’s alone also yielded a number of mutations, similar to what is reported internationally, as seen in Fig. Generally the K103 N was the main mutation detected within the NNRTIs group of drugs. K103 N is the most common resistance mutation that emerges during failure of an NNRTI-containing regimen –13. OtheResistance to NRTI’s, specially AZT was found to be induced by a range of mutations in RT including M41 L, K65R/E/N, D67N, K70R, I74V, I84V, M184 L, L210 W, T215Y/F, and K219E as previously reported in previous studies and annual reports , 19. FurMutations in the protease gene which is mediated by amino acid substitutions results in reducing the binding of the inhibitor to the protease leading to drug resistance to PI’s –22. The Additionally, when combinations of PI’s are used in therapy protocol, another set of mutations appear, both novel and reported. For example, mutation at codon 82, V82A/T/F/S, is reported in patients receiving treatment with indinavir and ritonavir  and alsoIt has been shown that mutations at codon 10, 63, or 71 compensated for a reduced viability of HIV-1 variants with a primary resistance mutation at codon 82 . High-leWith the expansion and the availability of different classes of ARV, virological suppression has been mostly achievable. Yet, with the continuous administration of HAART therapy, the development of ARV resistance is the major complication and challenge, in addition to other challenges such as patient compliance and development of ARV therapy toxicity –33. CurrDespite these efforts, drug-resistant viruses are increasing in the recent years in the HIV-infected population and this is caused by the rapid replication rate of the virus and its characteristic genetic adaptation capabilities –45. BaseAntiretroviral therapy failure is well documented and is characterized by the increase in viral replication and load. There are various causes leading to this failure, which is mainly caused by sub-therapeutic drug levels due to drug interactions, poor compliance, or poor absorption, or not following the proper protocol , 33. Thu, and India [Additionally, different backgrounds of the patients and their location all contribute to the developing of resistance causing mutations . Periodind India , in addiThe prevalence of primary or transmitted HIV drug resistance to all of the drugs and drug classes that were evaluated in this study was 66% and 34% respectively. These findings provide a useful background for antiretroviral therapy in KSA and contribute reference data for the surveillance of HIV drug resistance around the world. Additionally, the frequency of the reported mutations and the novel ones, as well as the outcome will be an interesting future study, additionally, reporting the patient behavior towards therapy and their compliance and its correlation with the frequency of development of both types of mutations would also be interesting. There are limited HIV drug resistance surveillance reports done, especially in KSA, this paper is a preliminary retrospective report and analysis of drug resistance mutations among our HIV patients. Identifying these resistance causing mutations will guide us to establish an in house diagnostic assay for screening of these possible DRM as a diagnostic tool for the HIV clinic and therefore to carefully monitor these resistant strains.


In Zimbabwe, viral load (VL) testing for people living with HIV on antiretroviral therapy is performed at the National Microbiology Reference Laboratory using a NucliSens machine. Anecdotal evidence has shown that invalid graphs for “Target Not Detectable (TND)” will upon repeat VL testing produce a valid result for virus not detected, therefore removing the need to repeat the test. This needs formal assessment.To determine i) intra- and inter-rater agreement of the visual interpretation of NucliSens graphs  between two laboratory scientists and ii) sensitivity, specificity and predictive values of the NucliSens graphs compared with repeat VL results.Cross sectional study using secondary data. Two laboratory scientists independently rated graphs one week apart for intra-rater agreement and compared final ratings with each other for inter-rater agreement. Consensus interpretations of graphs were compared with repeat VL results. Kappa coefficients were used to obtain measures of agreement.There were 562 patients with NucliSens graphs and repeat VL. Kappa scores were: 0.98 (Scientist A); 0.99 (Scientist B); 0.96 (Scientist A versus Scientist B); and 0.65 (NucliSens graphs versus VL). Sensitivity, specificity, positive predictive value and negative predictive value for graphs compared with VL were 71%, 92%, 79% and 89% respectively.Intra-and inter-rater agreements were almost perfect. The negative predictive value translates to a false negative rate of 11%. If repeat VL testing is not done, the clinical consequences need to be balanced against cost savings and the risks outweigh the benefits. The scale up of antiretroviral therapy (ART) worldwide has been a major public health success story, with 21.7million people accessing ART by the end of 2016[7 These targets specify that by 2020, 90% of people living with HIV will know their HIV status, 90% of people diagnosed as HIV-positive will receive ART and 90% of those on treatment will be virally suppressed.In the first few years of ART scale up, the World Health Organization (WHO) recommended that the response to therapy be monitored by clinical assessment and by CD4 cell count. This prZimbabwe has adopted the recent WHO Guidelines and the UNAIDS 90-90-90 targets,7. SinceFirst, invalid results and repeat testing often increase the laboratory turnaround time from receipt of specimens to production of results to beyond the expected standard of 14 working days. Second, human and material resources are expended in performing the repeat tests and this has cost implications for the HIV/AIDS programme. Third, in some cases the remaining plasma or Dried Blood Sample (DBS) may be insufficient for a repeat test, and the patient may be asked to attend the health facility and submit another sample. Repeat testing for VL therefore comes at a cost to the programme and to the patient.An invalid VL result is associated with three output graphs, all of which show a non-sigmoidal internal control line and which are stored in the computer of the NucliSens machine. Anecdotal evidence based on visual interpretation of the NucliSens graphs has shown that invalid graphs with “Target Not Detectable (TND)” will upon repeat VL testing almost always produce a valid result for virus not detected. If this anecdotal evidence was proven in a formal study, the laboratory could accept the visual interpretation of an invalid graph for TND as meaning no virus detected and there would be no need to repeat the VL test, saving time, energy and money. According to a PUBMED search, there have been no published studies on this particular subject.We therefore conducted this study in Zimbabwe to assess whether the visual interpretation of invalid graphs for TND was consistently associated with no virus detected on repeat VL testing. To do this, we obtained records of patients tested for VL between 2013 and 2017. We focused our study on those that initially had invalid results which showed, “Target Detectable (TD)”, TND or no line at all (NL) on the NucliSens graph on the initial test and were subjected to repeat VL testing. Specific objectives were to i) document demographic and laboratory characteristics of HIV patients whose records were included in the study, (ii) assess intra- and inter-rater agreement of the visual interpretation of graphs for TD, TND and NL, and to iii) calculate sensitivity, specificity and predictive values of visual interpretation of graphs compared with repeat VL test results.This was a cross-sectional study using routinely collected secondary data.Zimbabwe is a land-locked country in southern Africa with a population of approximately 13 million and a gross domestic product (GDP) of $924 per capita, which compared with $1,589 per capita for the sub-Saharan Africa region[This study was carried out at the NMRL, Harare, Zimbabwe. The NMRL has conducted all VL testing for the country up to February 2017, at which point VL testing was decentralized to different provinces. All VL testing in NMRL was done using the NucliSens platform  up to February 2017 when Abbott M2000sp and M2000rt machines  were also procured for the laboratory. The majority of samples sent from ART sites from around the country are dried blood spots (DBS) with five spots on each card. The only exception is Harare Central Hospital which sends plasma for VL rather than DBS. This is because the NMRL is located within Harare Central Hospital Complex. The procedures for collecting DBS samples and VL testing are described in the NMRL handbook and standard operating procedures manual,11. The Once the VL test has been completed, the machine provides a graphic representation and a printed results summary. Both of these are stored within the machine’s computer system. The printed results summary gives one of the following results: i) invalid result, ii) target virus not detected, or iii) target virus detected with a numeric count of the viral load in copies per ml. As described earlier, all invalid VL results are repeat tested using the remaining samples on the DBS or plasma card.The NucliSens machine produces one of five output graphs: two are valid graphs and three are invalid graphs as shown in Figures A-E in For a valid graph, the screen always shows a sigmoid line for the internal control. For the tested specimen, any visible graph line means virus is detected (TD) Fig A in  and no gFor an invalid graph, the screen shows no line(NL) at all for the internal control (which indicates no amplification) Fig C in , non-sigHIV-infected people on ART who had an initial invalid NucliSens graph and a repeat VL test in Zimbabwe between 2013 and 2017 were included in the study.A line list was made of all the initial invalid VL results that had a repeat VL test. From these, the principal investigator made a list of patients who had graphs . For every TND graph, controls were selected that either had a TD or an NL graph. The graphs were then mixed up. Two similarly experienced medical laboratory scientists (A and B) were trained to interpret the graphs for TND, TD or NL.Intra-rater agreement: each scientist independently decided on whether the graphs were TD, TND or NL at time T1. The same scientist looked again at the graphs one week later, blinded to what they had decided previously, and decided whether the graphs were TD, TND or NL (T2). The results from T1 and T2 were compared for intra-rater agreement.Inter-rater agreement: the final decisions on graphs were obtained for each scientist and these were compared between the two to enable inter-rater agreement to be assessed. Each scientist’s final decision was based on whether they agreed on each graph between T1 and T2 –if they had not agreed, they were asked to look for a third time at the graph and make a decision which was accepted as the final decision.Comparison of graphs with the result of repeat VL testing: When there had been disagreement between the two scientists, a consensus was reached between them on whether the graph was either TD or TND. The concordant results of the two scientists were then compared with the repeat VL printed results–virus detected or virus not detected. The data appears in Data variables for the study were collected into a structured proforma and included: sex; age; reason for viral load testing; graphs selected by the principal investigator as showing TD, TND and NL; repeat VL printed results that showed virus detected and virus not detected. The data codebook appears in “Data were double entered from the paper-based form into EpiData . These data were exported to STATA .The kappa coefficient  of agreement and its interpretation was used to determine intra- and inter-rater agreement on the visual interpretation of NucliSens graphs  by the two scientists (A and B). A two-bEthics approval was obtained from the Medical Research Council of Zimbabwe and the Ethics Advisory Group, International Union Against Tuberculosis and Lung Disease (The Union) Paris, France. As secondary data were used, a waiver for informed patient consent was obtained from the ethics committees.There were 562 patients whose characteristics are shown in Table A in Intra-rater agreement between scientist A and scientist B in the visual interpretation of invalid NucliSens graphs for TD, TND and NL are shown in Table B in Inter-rater agreement between Scientist A and B in the visual interpretation of invalid NucliSens graphs for TD, TND and NL are shown in Table C in Agreement in the visual interpretation of the NucliSens graphs showing TD and TND and the repeat viral load results showing virus detected and virus not detected are shown in Table D in This study confirms that based on the kappa score there was substantial agreement between visual interpretation of the NucliSens graphs showing TND and repeat viral load test results showing virus not detectable. Visual interpretation of the graphs was also straight forward and replicable as indicated by the almost perfect intra-rater and inter-rater agreements when two trained medical laboratory scientists conducted this exercise independently one week apart.From the point of view of laboratory policy, the key question that needed to be asked was whether the visual interpretation of invalid graphs for TND was consistently associated with no virus detectable and could the laboratory therefore dispense with repeat viral load testing. The high specificity in our study means that 92% of patients with no virus detectable had an invalid graph showing TND. The high negative predictive value means that 89% of invalid graphs showing TND had no virus detectable on repeat VL testing. However, this translates to a false negative rate of 11%; namely that when a medical laboratory scientist concludes that the NucliSens graph of an invalid test shows TND there is in fact detectable virus in the specimen.Does a false negative rate of 11% matter in terms of laboratory, clinical and programmatic management? In clinical practice, it would mean that 11% of patients on ART would be assessed as having viral suppression when this is not the case and virus is indeed present. The most recent WHO guidelines specify that if virus is present in a patient sample, then a precise VL must be obtained. If the Is it safe to leave a patient on ART with detectable virus for another 12 months before the next repeat VL monitoring test? Probably not! It is well established that detectable virus leads to immunosuppression which increases the risks of further morbidity and mortality. ART with viral suppression effectively prevents the transmission of HIV to other non-infected persons,14, and What are the cost savings of this potential approach of accepting invalid TND NucliSens graphs as indicating no virus detected and therefore not repeating the VL test? Working on 10,000 VL tests per month (the approximate workload of the NMRL) with 2.5% being invalid, there would be 250 invalid tests of which half (125) would be TND. Each VL test costs about USD$80 and with none of these TND tests being repeated, there would be a cost savings of USD$10,000 per month or USD$120,000 per annum. For a resource-poor country such as Zimbabwe, this is a considerable cost saving for the NMRL.The main strengths of the study were the large sample size, the NMRL being the major reference laboratory for the whole country and the rigorous methodology for ascertaining intra- and inter-rater agreements and sensitivity, specificity and predictive values. The NMRL also participates in an external quality assurance programme with excellent results, and therefore the results of VL testing are probably accurate and consistently reproducible. The main limitations were a failure to collect more detailed baseline clinical information on the patients which may have been useful for further analysis and we did not collect precise measurements of VL in copies/mL which would have helped in deciding who had VL above and below 1000 copies/mL on the 11%.We are unaware of any previous published studies on this subject and therefore cannot compare our findings with those of others. The way forward is for us now to present this data and information to programmatic, clinical and laboratory stakeholders, including those who are responsible for policy decisions within the National AIDS Programme, and decide on what further studies should be done to clarify the situation (such as measuring VL in copies/mL) and on whether or not to dispense with repeat VL testing in those patients for whom an invalid NucliSens graph shows TND. Although there may be considerable cost savings from this approach, there are clinical and programmatic risks, and we feel that we cannot recommend a policy of not repeating the VL if an invalid initial test shows a TND graph.In conclusion, we evaluated whether the visual interpretation of invalid graphs for TND were consistently associated with no virus detected on repeat VL testing. While we observed excellent intra- and inter-agreement between the two trained medical laboratory scientists in visual interpretation of the graphs, the specificity and negative predictive values of TD and TND graphs against repeat VL test results were 92% and 89% respectively. The false negative rate of 11% with its programmatic and clinical consequences has to be balanced against the cost savings of dispensing with the repeat VL test. On balance the risks outweigh the benefits and we cannot recommend a policy of no repeat VL testing.S1 Fig(DOCX)Click here for additional data file.S1 Tables(DOCX)Click here for additional data file.S1 Dataset(ZIP)Click here for additional data file.S2 Dataset(ZIP)Click here for additional data file.


Globally, anemia, among people living with HIV/AIDS, is a major public health problem. It has a significant effect on the progression of HIV/AIDS to advanced stages and there are a number of factors that often affect anemia. However, there is little insight regarding factors affecting anemia among HIV/AIDS patients in developing countries, including Ethiopia. This study aimed at investigating factors affecting anemia among people living with HIV/AIDS taking ART drug at Tikur Anbessa Specialized Hospital, Addis Ababa, Ethiopia. A hospital based cross-sectional study design was used to assess factors affecting anemia among people living with HIV/AIDS. Structured checklist was used to gather information from charts of patients selected by simple random sampling method. We analyzed the data to identify factors associated with anemia among people with HIV/AIDS using logistic regression models. A total of 301 selected charts were reviewed. The median age was 38 ± 10.38. The majority (62.5%) of the patients were taking ZDV-containing ART drug (ZDV/3TC/NVP). The overall anemia prevalence was 34.6%, while about 5%, 15.6%, and 14% of the patients had severe, moderate, and mild prevalence of anemia, respectively. Factors that were found to affect anemia among these patients include gender , occupation , WBC count , platelet count , nutritional status , and WHO clinical stage of HIV/AIDS . About one in three patients was found to be anemic. Intervention aimed at diagnosing and treating anemia among people living with HIV/AIDS should be considered. Anemia is a major public health problem, especially in developing countries –3. It haUndeniably, anemia is the most important clinical problem seen in people living with HIV/AIDS ; its sevHIV leads to declining the production of cytokine, which ultimately influences hematopoiesis process of the cells, specifically erythropoietin concentration . This, iμL, CD4+T-lymphocyte count < 200 cells/μL, and platelets count < 200,000 cells/μL , whereas, for men, it was defined as hemoglobin <13.0:  , 25.μL), WBC count (cells/μL), platelet count (cells/μL), nutritional status (BMI) (kg/m2), WHO clinical stage of HIV/AIDS, ART regimen, and anti-TB treatment history. The CD4 T-lymphocyte was determined based on WHO  recommendation of 2007 to commence ART for HIV positive peoples [The sociodemographic characteristics identified from the patients' chart include age, gender, residence, religion, ethnic group, marital status, educational status, occupation, and monthly income. Moreover, the clinical and biochemical test characteristics of patients gathered from charts include CD4 count (cells/ peoples . The WBC peoples . In orde peoples . The cli peoples . The ARTThe questionnaire (checklist) was developed after reviewing literatures and WHO guidelines. The checklist comprised sociodemographic and clinical characteristics: age, gender, residence, religion, ethnic group, marital status, educational status, occupation, monthly income, and clinical characteristics: CD4 T-lymphocyte, hemoglobin, WBC, platelet count level, BMI level, WHO clinical stage of HIV/AIDS, ART regimen, and history to treatment for TB. Before the actual data collection date, the questionnaire was pretested on 5% of charts of the study population. Then, necessary modification was done based on its analysis. The study questionnaire was prepared in English. A graduate class nurse student collected the data. Principal orientation was given for data collector prior to the data collection date on how to collect the data and make them familiar with the data collection questionnaire. The completeness of each questionnaire was checked on daily basis.Data was entered using EpiData version 3.1 statistical package software. After thorough cleaning, the data was exported to SPSS version 22 statistical package software for further analysis. Descriptive statistics such as mean, standard deviations (SD), frequency, and percentage were used to describe the variables of the study. In order to describe factors associated with anemia among people living with HIV/AIDS, unadjusted and adjusted odds ratios with their 95% CI were calculated using logistic regression models.The ethical clearance letter was obtained from ethical clearance committee of Addis Ababa University College of Health Sciences. After we offered the letter to the head department of ART clinic, the purpose of the study was clearly stated to the healthcare professionals working in the ART clinic of the Hospital.A total of 301 charts were reviewed. The median age was 38±10.38. The majority (62.1%) of the study participants were female. About half (50.5%) of the study participants were unemployed. More than three-fourths (90%) of the study participants were from urban residential area. About half (49.8%) of the study participants were married.  ± 195.67, 5490 ± 4015, and 287000 ± 97499.53, respectively. The majority  of the study participant's CD4, WBC, and platelet count were ≥ 200, ≥ 4500, and ≥ 150,000 cells/μL, respectively. Regarding the study participant's nutritional status, 61.5%, 20.6%, 12%, and 6% of the study participants were found to be of normal weight, overweight, underweight, and obese in their nutritional status, respectively. Further, almost three-fourths of the study participants were taking ZDV-containing ART drug (ZDV/3TC/NVP) (The medians (±SD) of CD4, WBC, and platelet counts were 3323TC/NVP) .μL, respectively. A noteworthy number (40) of anemic study participants were found in stage IV of WHO clinical stage of HIV/AIDS. Regarding ART regimen and anti-TB drug exposure history, a considerable number (60 and 36) of anemic study participants were taking ZDV-containing ART drug (ZDV/3TC/NVP) and had history of anti-TB drug treatment exposure, respectively , a considerable number (60) were anemic, while the remaining 128 were not  of study participants were from the age group of 30-39. A momentous number (75 and 60) of anemic study participants had CD4 T-lymphocyte and WBC count ≥ 200 and ≥ 4500 cells/ectively . Among awere not .χ2 = 6.89, p < 0.010) and occupation , were significantly associated with anemia , platelet count , nutritional status , and WHO clinical HIV/AIDS stage  were significantly associated with anemia ( P < 0.009), WBC count , platelet count , nutritional status , and WHO clinical HIV stage  were among the different factors associated with anemia among people living with HIV/AIDS  Monitor hemoglobin level on each of the follow-up periods by focusing on those people taking ART drug regimen containing ZDV and take appropriate action to combat the anemia if anemia is detected. This is because the pathogenesis of anemia is worrisome and ZDV by itself has an effect on reducing proliferation of red blood cells. (2) Monitor adherence level of the people living with HIV/AIDS taking ART drug to their therapy and motivate, encourage, and advise them to adhere to the ART drug therapy consistently to gain its therapeutic effect. This is because ART drug has an effect on combating anemia among people living with HIV/AIDS taking ART drug. (3) Monitor and take quick appropriate action for drug-drug interaction and magnified side effects of each drug which may occur in people living with HIV/AIDS taking ART drug. This is because drug-drug interaction and side effects of some drugs by themselves interfere with reduced proliferation of granulocytes and contribute to raising immature destruction of cells. (4) Increase awareness of people living with HIV/AIDS taking ART drug about the benefit of adhering to consumption of nutritional diet consistently. This is because consuming appropriate diet helps the body in proliferating enough amounts of red blood cells and other granulocytes. This study has limitations. Firstly, it is a cross-sectional study that has chicken egg dilemma in identifying causal relationship. Secondly, the study excludes people living with HIV/AIDS who are naïve to ART drug. Thus, it is recommended that future study considers longitudinal study by considering ART-naïve people living with HIV/AIDS by determining the severity of anemia.Bearing in mind the effect of anemia and its significant prevalence, it is a great disquiet that more than one-third of the study participants were anemic, while appropriate treatment options are available and the study participants were on ART drug for at least three months, which have an effect to combat the pathogenesis of the disease. Therefore, based on the findings of this study, the following interventions are suggested to detect and combat anemia among people living with HIV/AIDS taking ART drug.In conclusion, it is important to alleviate anemia among people living with HIV/AID taking ART drug, as the presence of anemia among people living with HIV/AIDS fastens the progression of the disease to the advanced stages. The suggested intervention by this study focused at improving nutritional status of people living with HIV/AIDS taking ART drug through enhancing awareness of benefit of consuming balanced diet and monitoring and managing granulocytes level appropriately, which increases survival rate; it is best to monitor granulocytes like CD4 count, platelet and WBC count, and hemoglobin level more frequently. It is also good to focus on sociodemographic characteristics like gender and occupation of the client, which have an ultimate effect in affecting level of anemia among people living with HIV/AIDS.