North-west Ethiopia faces the highest burden world-wide of visceral leishmaniasis (VL) and HIV co-infection. VL-HIV co-infected patients have higher  parasitological failure and relapse rates than HIV-negative VL patients. Whereas secondary prophylaxis reduces the relapse rate, parasitological failure rates remain high with the available antileishmanial drugs, especially when administered as monotherapy. We aimed to determine the initial effectiveness  of a combination of liposomal amphotericin B (AmBisome) and miltefosine for treatment of VL in HIV co-infected patients.We conducted a retrospective cohort study at a Médecins Sans Frontières—supported health center in north-west Ethiopia. We included VL-HIV co-infected adults, treated for VL between January 2011 and August 2014, with AmBisome infusion  and miltefosine orally for 28 days (100 mg/day). Proportions of initial treatment outcome categories were calculated. Predictors of initial parasitological failure and of death were determined using multivariable logistic regression. Of the 173 patients included, 170 (98.3%) were male and the median age was 32 years. The proportion of patients with primary VL (48.0%) and relapse VL (52.0%) were similar. The majority had advanced HIV disease  and were on antiretroviral therapy prior to VL diagnosis . Initial cure rate was 83.8% ; death rate 12.7%  and parasitological failure rate 3.5% . Tuberculosis co-infection at VL diagnosis was predictive of parasitological failure (adjusted odds ratio (aOR), 8.14; p = 0.02). Predictors of death were age >40 years , hemoglobin ≤6.5 g/dL  and primary VL .Initial parasitological failure rates were very low with AmBisome and miltefosine combination therapy. This regimen seems a suitable treatment option. Knowledge of predictors of poor outcome may facilitate better management. These findings remain to be confirmed in clinical trials. North-west Ethiopia faces the highest burden world-wide of visceral leishmaniasis (VL) and HIV co-infection. VL treatment outcomes in HIV co-infected patients are associated with high initial treatment  failure and recurrence rates after cure (relapse). With secondary chemoprophylaxis, the risk of relapse can be reduced. However, with the current VL treatment regimens, the initial parasitological failure rates remain high. In this study, we aimed to determine the initial effectiveness of a combination of liposomal amphotericin B (AmBisome) and miltefosine for treatment of VL in HIV patients in Ethiopia. We conducted a retrospective study using routine program data from a Médecins Sans Frontières—supported health center in north-west Ethiopia. We included 173 adult VL-HIV co-infected patients treated for VL with a combination of AmBisome and miltefosine. Initial cure rate was 83.8%, death rate 12.7% and parasitological failure rate 3.5%. Tuberculosis co-infection at VL diagnosis was predictive of initial parasitological failure. Predictors of death were age >40 years, hemoglobin ≤6.5 g/dL and primary VL. Initial parasitological failure rates were very low with AmBisome and miltefosine combination therapy. This regimen seems a suitable treatment option. Knowledge of predictors of poor outcome may facilitate better management. These findings remain to be confirmed in other studies. Leishmania donovani species complex , duration of illness (months) and VL treatment history .2], spleen size (centimeters), the level of weakness, ascites, peripheral edema, bleeding and jaundice. The spleen size (centimeters) was measured from the junction of the anterior axillary line and the left coastal margin to the tip of the spleen. Weakness severity was defined according to MSF guidelines ) . Hemogloidelines ,33.The primary outcome was the initial treatment outcome . The proportion of individuals with the different outcome categories (excluding patients that defaulted or were transferred-out), were calculated with 95% Wilson confidence intervals (CI). In secondary analysis, the association of initial treatment outcome with VL treatment history was assessed by the Chi-squared or Fisher’s exact test.p-value was <0.1, the predictor was included in a multivariable logistic regression model. Non-significant variables  were removed step by step until no more variables could be dropped.Predictors of initial parasitological failure and predictors of death were determined. Predictors of parasitological failure were analyzed among patients with parasitological failure or cure, whereas predictors of death were analyzed among patients who died or stayed alive . The choice of variables analyzed as predictors was based on literature review and consideration of variables available in the dataset. To overcome the problem of substantial missing baseline CD4 count results, we created a composite marker for advanced HIV disease, defined as having either additional WHO stage IV disease  or a CD4Leishmania parasites at exit, they were all classified as having parasitological failure. All the statistical methods described above were then repeated. All statistical analyses were performed with Stata version 14.Lastly, a sensitivity analysis similar to an intention to treat analysis was performed. Defaulters and transfer-outs were included in this sensitivity analysis; and considering that they probably had Ethics approval was received from the Institutional Review Board of the Institute of Tropical Medicine, Antwerp, Belgium, and the Ethical Review Committee of the Institute of Public Health, Gondar University, Ethiopia. This research fulfilled the exemption criteria set by the MSF Ethical Review Board (ERB) for a posteriori analyses of routinely collected clinical data, and thus did not require MSF ERB review. It was conducted with permission from the Medical Director of the MSF Operational Centre Amsterdam.Between January 2011 and August 2014, 227 patients were diagnosed with VL-HIV co-infection and treated at the Abdurafi health center. Forty patients were treated with other VL treatment regimens . Two patients were started on AmBisome and miltefosine combination treatment, however miltefosine was later discontinued. The reason for discontinuing miltefosine was because of miltefosine stock-out rather than adverse event. Both of these patients completed AmBisome monotherapy and one was cured and the other died. Four patients defaulted, 5 were transferred out and 3 had unknown treatment outcome. These 54 (23.8%) patients were excluded from the main (per-protocol) analysis. A total of 173 patients were included in the main analysis .A comparison of characteristics of patients excluded or included in the study is shown in Most patients were male , residents  and young . The proportion of patients with primary VL  and relapse VL  were similar. Most patients had advanced HIV disease  and were on ART prior to VL diagnosis  .vs. 75.6%), had been ill for ≥2 months (46.2% vs. 27.9%) and had not started ART prior to the VL episode (57.7% vs. 16.1%). Lastly, primary VL patients had a lower parasite load at admission (median +4 vs. +5) and a lower proportion had cure confirmed by a parasitological test (43.1% vs. 65.0%)  .The outcomes were: cured, 145/173 ; died, 22/173  and parasitological failure, 6/173 . The outcome by VL treatment history was significantly different as shown in Of the 6 patients with initial parasitological failure , 1 was rTuberculosis co-infection at VL diagnosis was predictive of initial parasitological failure (adjusted odds ratio (aOR), 8.14; 95% CI, 1.42–46.72; p = 0.02). There was a statistically non-significant association between high tissue parasite load (parasite grade 6+) at VL diagnosis and initial parasitological failure. In multivariable analysis, VL treatment history was not significantly associated with initial parasitological failure .Independent predictors of death were age >40 years , hemoglobin ≤6.5 g/dL  and primary VL  as shown in 2  was also predictive of initial parasitological failure as shown in The initial treatment outcomes were similar to those from the main analysis as shown in VL-HIV co-infected patients are confronted with high  parasitological failure rates and high relapse rates ,8. Whilevs. 32.8%; p<0.001), cure rates were significantly higher , and death rates were non-significantly higher . None of the deaths are considered treatment-related. In the present study, we had a higher admission rate of late stage VL patients that were referred from other hospitals as compared to the previous AmBisome monotherapy study. We have recently shown that besides HIV serostatus, other important predictors of death were: age >40 years, hemoglobin ≤6.5 g/dL, bleeding, jaundice, edema, ascites and tuberculosis [vs. 39.6%; p<0.001) [Although it remains difficult to compare historical cohorts, the initial treatment outcomes with combination therapy compared with those for AmBisome monotherapy—the previous first line treatment at the MSF treatment site , are as rculosis . We alsorculosis ,36. If wp<0.001) .In vitro studies have demonstrated synergy between liposomal amphotericin B and miltefosine [In contrast to pentavalent antimonials that cause severe adverse events resulting in high case fatality rates ,12–18, Atefosine . A combitefosine . AmBisomtefosine . For co-tefosine .Thanks to an agreement between Gilead and WHO, on a donation programme, WHO is providing AmBisome for treatment of visceral leishmaniasis for free to low income countries in East Africa and South Asia. This donation programme started in 2012, and has been extended in 2017 for another five years, including middle income countries. This access to free AmBisome will enhance affordability of implementing AmBisome-based treatment regimens for VL-HIV –44. Comb2  was predictive of initial parasitological failure. As with tuberculosis, severe malnutrition causes immunosuppression which may inhibit parasite clearance, resulting in parasitological failure.We found that tuberculosis co-infection at VL diagnosis was predictive of initial parasitological failure. This is similar to findings from a study conducted in Sudan . TubercuThe predictors of death identified—age >40 years and hemoglobin level ≤6.5 g/dL—are similar to those reported from other studies ,48,49. PMost of our patients are young adult males who get infected with leishmania while working in the agricultural fields within the VL endemic region. HIV infection is also more common in young adults than children. As shown in e.g. patients with a history of VL) were more likely to get a parasitological test for confirmation of cure at the end of treatment. Furthermore, in a recent study from this setting, we found no difference in long term outcomes (relapse or death) among patients with treatment outcome at discharge of parasitological cure versus those with clinical cure [There are some limitations to this study. Diagnosis and cure were not systematically confirmed by parasitological tests. Fifty-seven 88.0%) of the patients whose cure was assessed clinically had non or barely palpable spleen at the end-of-treatment, inhibiting performing a spleen aspirate, they declined having a bone marrow aspirate because the procedure is painful and they didn’t have palpable lymph nodes. This occurs commonly in settings without non-invasive investigations to assess VL cure. We acknowledge that this could likely lead to some degree of underestimation of the failure rates. However, it is important to note that in this cohort of patients, the more ill patients at admission (for instance with tuberculosis co-infection) and those more likely to fail Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 Table(DOCX)Click here for additional data file.S1 Checklist(DOC)Click here for additional data file.


HIV genetic diversity implicates major challenges for the control of viral infection by the immune system and for the identification of an effective immunotherapeutic strategy. With the present case report we underline as HIV evolution could be effectively halted by early antiretroviral treatment (eART). Few cases supported this evidence due to the difficulty of performing amplification and sequencing analysis in long-term viral suppressed patients. Here, we reported the case of limited HIV-1 viral evolution over time in a successful early treated child.A perinatally HIV-1 infected infant was treated within 7 weeks of age with zidovudine, lamivudine, nevirapine and lopinavir/ritonavir. At antiretroviral treatment (ART) initiation HIV-1 viral load (VL) and CD4 percentage were >500,000 copies/ml and 35%, respectively. Plasma genotypic resistance test showed a wild-type virus. The child reached VL undetectability after 33 weeks of combination antiretroviral therapy (cART) since he maintained a stable VL <40copies/ml. After 116 weeks on ART we were able to perform amplification and sequencing assay on the plasma virus. At this time VL was <40 copies/ml and CD4 percentage was 40%. Again the genotypic resistance test revealed a wild-type virus. The phylogenetic analysis performed on the HIV-1 pol sequences of the mother and the child revealed that sequences clustered with C subtype reference strains and formed a monophyletic cluster distinct from the other C sequences included in the analysis . Any major evolutionary divergence was detected.eART limits the viral evolution avoiding the emergence of new viral variants. This result may have important implications in host immune control and may sustain the challenge search of new personalized immunotherapeutic approaches to achieve a prolonged viral remission.The online version of this article (doi:10.1186/s12879-016-2092-z) contains supplementary material, which is available to authorized users. HIV-1 infection is characterized by broad genetic diversity and rapid evolution that influence the pathogenesis, transmission and clinical management of the infection . SimilarAn infant was born by elective caesarean section, two hours after membrane rupture at 38 weeks of gestation to an HIV-1 infected woman who started a first line ART with lamivudine, zidovudine and lopinavir/ritonavir 1 week before the delivery. Maternal VL at delivery was 203,686 copies/ml. Intrapartum antiretroviral prophylaxis with intravenous zidovudine was administered and the child started postnatal prophylaxis with zidovudine at 6 h of birth for 6 weeks. Breastfeeding was avoided. Polymerase chain reaction (PCR) for genes GAG, POL, and ENV performed at 6 weeks of life was positive and his HIV-RNA VL was >500,000 copies/mL with CD4 lymphocyte percentage 35%.th week from starting therapy despite an adequate maternal compliance with infant’s drugs and on subsequent medical check the baby maintained undetectable HIV-1 RNA and CD4 T count within the range for age. At 48 weeks from starting therapy, ART was simplified by suspending protease inhibitor. Concurrently to the VL undetectability, HIV-1 antibodies were negative in the child at 26, and 31 months of age. After 116 weeks on cART, we were able to perform viral isolation and amplification. At this time VL was stable <40 copies/ml (ABBOTT) and CD4 percentage was 40%. A genotypic resistance test for pol was re-performed and no-drug resistance was found for a second time.The genotypic resistance test from both mother and child didn’t show any transmitted drug resistance for PI, NRTI or NNRTI drug classes. Based on these results, at 7 weeks of age cART was initiated. A four-drug regimen of zidovudine, lamivudine, nevirapine and lopinavir/ritonavir was selected according to the results of genotypic resistance test. Plasma VL remained detectable till the 32pol sequences at the time of birth and 2 years later (corresponding to the 116 week of ART), were available for the child. To define the HIV-1 subtype and the sequence inter-relationships between the mother/child pair a neighbor joining (NJ) tree ).HIV-1 populations in the blood of the newly infected individuals are largely homogenous and evolve in a manner consistent with exponential viral replication . Thus, s


HIV‐infected children. A retrospective cohort study was conducted between January 2006 and December 2015. All pediatric patients of <15 years old and who took at least 6 months of ART in Fiche and Kuyu hospitals of Oromia region, Ethiopia were included in the study. Collected data were entered and analyzed using SPSS version 20. Multivariable logistic regression was employed to identify predictors of treatment failure. Data of 269 children were analyzed and majority (53.9%) of the children were males with a mean age of 9.8 ± 3.64 years. Based on the two WHO criteria, overall ART failure was found to be 51 (18.8%), of which 33 (12.26%) had clinical and 18 (6.69%) had immunologic failures. The mean time to the detection of treatment failure was 41 ± 24.96 months. Children's age between 6 and 9 years  was protective toward treatment failure, while lost to follow‐up , died , transferred out , suboptimal adherence to ART , baseline CD4 count of <50 cells/mm3 , and WHO advanced clinical stage  were found to be predictors of treatment failure. The study revealed that the treatment failure is high and the mean time to develop treatment failure is short. The predictors for treatment failure were suboptimal adherence, lost to follow‐up, transferred out, initial CD4 count <50 cells/mm3, initial WHO stages 3 and 4. On the other hand, being in the age of 6–9 years is protective from developing treatment failure as compared to the other age category.The objective of the study was to assess the magnitude and predictors of first‐line antiretroviral treatment failure among  AIDSacquired immune deficiency syndromeARTantiretroviral treatmentARVantiretroviralCD4cluster of differentiation 4CDCCenter for Disease Control and PreventionDBSdried blood spotHIVhuman immunodeficiency virusIPTisoniazid prophylaxisOIopportunistic infectionPMTCTprevention of mother to child transmissionWHOWorld Health OrganizationAlthough initially acquired immune deficiency syndrome (AIDS) was discovered among a certain vulnerable populations, it later became a worldwide pandemic. In 2015, there were 2.1 million new human immunodeficiency virus (HIV) infections worldwide, adding up to a total of 36.7 million people living with HIV WHO, . In the Even with a lot of efforts in place to control the disease, it remains a major global public health challenge. There are 2.1 million new HIV infections that occurred globally and majority of cases are in sub‐Saharan African countries. HIV infection among children constitutes 7.5% of people living with HIV worldwide and 14.4% of total HIV/AIDS deaths. About 50% of children living with HIV/AIDS die before the age of 2 years as a result of recurrent opportunistic infections such as pneumonia, diarrhea, malnutrition, and malaria WHO, .At the end of 2015, there were 17 million people on ART and the global coverage reached to 46% WHO, . On the Ethiopia is one of the top five African countries having total of 765,500 people living with HIV/AIDS , free ART service provision was started in three hospitals since 2005 and later decentralized to other hospitals and health centers. During the program initiation, general medical practitioners were the sole service providers, but later the government started a task shifting to other health professionals  so as to increase access. In June 2014, the region had a total of 194,370 HIV‐positive patients and 115,334 of them were on ART follow‐up, of these 15,586 were <15 years old  had clinical treatment failure, 79 (6.7%) immunologic failure, and 18 (1.5%) developed both immunologic and clinical failure FMOH, . A studyThere are various risk factors described in different studies; the sociodemographic factors , baseline clinical factors , drug–drug interactions, drug side effects, drug toxicity, or inadequate adherence to treatment are some of the factors associated with treatment failure  were male. It has 14 woreda health offices, and ART service is being provided by 15 ART sites . Besides, there are 15 prevention of mother to child transmission (PMTCT) only sites providing option B+ service in the zone. Since the initiation of the program, a total of 7075 patients started ART, and of them 5174 were on ART, as of December 2015. Fiche and Kuyu hospitals are among high client load ART facilities in Oromia region, and they started provision of free ART services in 2005 , ART regimen, prophylaxis, and adherence level in pediatric patients. Caregivers‐related data were also retrieved from the record.Two diploma level holders of ART clinic data entry clerks were recruited as data collectors. The data collectors attended a 1 day training focused on the aim of the study and detailed review of the tool. In addition to the practical training, adequate supervision and follow‐up were done by the supervisor to maximize quality of the data collected.P < 0.05.The collected data were then coded, entered, cleaned, and analyzed using SPSS version 20. Simple descriptive statistics including mean, percentage, and standard deviations were computed to summarize categorical variables. Moreover, multivariable logistic regression was performed to explore possible associations between the dependent variable (first‐line ART failure) and the independent variables. Association between the outcome and the independent variables was taken as significant at The study used the standard definition of immunologic failure which is defined as developing or returning to the following age‐related immunological thresholds after at least 24 weeks on ART, CD4 count of <200 or <10 for child 2–5 years, and CD4 count <100 for a child 5 years or older. Clinical failure is also considered for a child with appearance or reappearance of WHO clinical stage 3 or 4 disease at least 24 weeks after treatment initiation FMOH, .In Ethiopia viral load testing is not available in routine practice settings; hence, treatment failure in this study is determined based on the two WHO criteria . In this cohort, the mean time to detection of treatment failure indicates the time between ART initiation and detection of failure of first‐line ART. Time to switch to second‐line ART regimens represents the time between detection of first‐line treatment failure and the time of initiation of second‐line drugs. Suboptimal adherence is defined in terms of total missed appointments described by more than 3 days per month FMOH, .Ethical approval was sought from the Ethics Review Committee of Africa Medical College and Oroima Regional Health Bureau. Permission letter was also granted from North Shewa zonal health department and both hospitals. Patient cards were coded and collected data were locked in a lockable cabinet in order to maintain confidentiality of the information obtained.A total of 269 study participants medical charts were reviewed, of them 118 and 151 pediatric patients had ART follow‐up in Kuyu and Fiche hospitals, respectively. More than half (53.9%) of patients were males and majority (56.8%) of the pediatrics were within the age of 10–15 years (Table Record of school enrollment status of study participants indicated that majority (76.2%) of them had history of school enrollment records. Regarding their family status, 148 (55%) of them reported that both of their parents were alive, 71 (26.4%) had one parent alive, and 50 (18.6%) of the pediatrics claim that both parents died. The study also showed that most (68.8%) of the study participants were accompanied by their mother as their primary caregiver, followed by fathers, 33 (12.3%). Concerning the employment status of their primary caregivers, more than two third (60.2) of the caregivers were not employed  of them were on ART follow‐up at the time of study, 19 (7.1%) were reported as lost, 9 (3.3%) dead, and 43 (16.0%) transferred out. The study also revealed that PMTCT coverage was very low, where only 16 (5.9%) of them had documented provision of NVP prophylaxis during delivery and postnatal period. Isoniazid prophylaxis (IPT) was provided to 158 (58.7%) of the pediatrics, but the rest (41.3%) of the participants did not received IPT at least once  were at severe immunological stage with CD4 count <50 cells/mm3. Record of their initial WHO stage also revealed that more than half (53.9%) of the pediatrics were at WHO clinical stage 3. The choice of ART regimen for pediatric patients was compared and accordingly, higher proportion (89.7%) of initial regimen prescribed at baseline included nevirapine as a combination. A d4t‐3TC‐NVP regimen covered 124 (46.1%) and d4t‐3TC‐EFV regimen covered 8 (2.9%) of the total prescribed ARV regimen. Zidovudine (AZT) with NVP was prescribed as initial regimen to 98 (36.4%) of the pediatric patients. However, first‐line ARV regimen substitution was done for 131 (48.7%) to other alternative group first‐line ARV drugs due to side effect, severe toxicity, and opportunistic infection, and most of them taking 4a (d4t+3TC+NVP).From the 269 pediatrics who were eligible for ART initiation, 100 (37.2%) of the ART were initiated by advanced WHO clinical staging criteria, 48 (17.8%) due to low CD4 count, 117 (43.5%) by both immunologic and clinical criteria. During the initiation of ART, 59 (24.0%) of study participant were found to be in advanced immunodeficiency state with CD4 <200 cells/mmAdherence to ART for the last 6 months of follow‐up was assessed and the study showed that majority (84.4%) of the study participants had optimal adherence and 48 (15.6%) of the pediatrics had suboptimal adherence. The number of missed appointment days was also found to be high where more than 9 (93.6%) of the 10 study participants had missed appointments for 8 days or more. The HIV‐positive status of the study participants was not disclosed to majority, 253 (94.1%), of the pediatric patients  standard growth curve. The study revealed that most of them were malnourished, where 93 (44.71%) of them were severely malnourished and 113 (54.33%) of the children were moderately malnourished. Similar anthropometric measurement was sought for the last visits and 57 21.2%) and 128 (47.6%) of the pediatrics were found to be severely and moderately malnourished, respectively. Of the children, 34 (12.64%) developed opportunistic infections after initiation of ART and of these, extrapulmonary tuberculosis (29.4%) accounted the largest proportion followed by diarrhea (14.7%)  failure cases were reported. Majority (12.36%) of the treatment failures were clinical and only 6.6% were immunologic failures. Of total 51 ART failure cases, 12 (23.4%) of them were confirmed by viral load test and switched to second‐line ART regimen. The mean time to develop treatment failure after initiation of first‐line regimen was 41 ± 24.96 months with a range from 16 to 104 months. More than 1 in 10 of the pediatrics developed treatment failure within 24 months. Of those who confirmed to have treatment failure, most (75.0%) of them were switched to second‐line ART in <30 days. After detection of the treatment failure, on average, it took 33.41 ± 11.93 days to initiate the second‐line ARVs regimen  as compared to 10–15 years old children. In other words, pediatrics in the age group of 6–9 years had a 74% reduction in risk of developing treatment failure than their counterparts. Pediatrics' outcome at last visit was positively associated with treatment failure in that those who lost to follow‐up , died , and transferred out  had more risk of acquiring treatment failure compared to those pediatrics who were still on ART follow‐up . Children who were in advanced WHO clinical stage 3 or 4 had 2.56 times more risk of developing treatment failure when compared to those asymptomatic and WHO stage 2 patients  . Children who had baseline CD4 counts <50 cells/mmThis retrospective cohort study was designed to determine the magnitude and predictors of first‐line ART failure among pediatric patients in Fiche and Kuyu hospitals in Oromia region, Ethiopia. The study showed that clinical failure was commonly encountered than immunologic failure with a total ART failure rate of 18.9%. The proportion of patients that failed treatment in the current study is higher than the retrospective cohort study done in Jimma University hospital and four referral hospitals in Addis Ababa  of the children were disclosed about their serology status, while their mean age lies between 10 and 14 years which is near to adolescent age. On the other hand, disclosure is not significantly associated with treatment failure which is against other findings from Ethiopia and Tanzania  of them were clinical failures. The mean time to ART failure was 41 ± 24.96 months and the mean days to initiation of second‐line ARV after detection of first‐line treatment failure was 33.41 ± 11.93 days. Patients' age between 6 and 9 years is protective from treatment failure, while initiation of ART at low CD4 count <50 cells/mmAll authors contributed equally in the design, data collection, and write up of this article. All the authors have read and approved the final manuscript.We confirm that this work is original and has not been published elsewhere nor is it currently under consideration for publication elsewhere. The authors have no support or funding to report. The authors have declared that no conflicts of interest exist.


Clinical trials of PI monotherapy indicate that most participants maintain viral suppression and emergent protease resistance is rare. However, outcomes among patients receiving PI monotherapy for clinical reasons, such as toxicity or adherence issues, are less well studied.An observational study of patients attending an HIV treatment centre in London, UK, who had received PI monotherapy between 2004 and 2013, was conducted using prospectively collected clinical data and genotypic resistance reports. Survival analysis techniques were used to examine the times to virological failure and treatment discontinuation.Ninety-five patients had PI monotherapy treatment for a median duration of 126 weeks. Virological failure occurred during 64% of episodes and 8% of patients developed emergent protease mutations. We estimate failure occurs in half of episodes within 2 years following initiation. Where PI monotherapy was continued following virological failure, 68% of patients achieved viral re-suppression. Despite a high incidence of virological failure, many patients continued PI monotherapy and 79% of episodes were ongoing at the end of the study. The type of PI used, the presence of baseline protease mutations and the plasma HIV RNA at initiation did not have a significant impact on treatment outcomes.There was a higher incidence of virological failure and emerging resistance in our UK clinical setting than described in PI monotherapy clinical trials and other European observational studies. Despite this, many patients continued PI monotherapy and regained viral suppression, indicating this strategy remains a viable option in certain individuals following careful clinical evaluation. Combination regimens may be more difficult to adhere to because of reduced tolerability, increased pill burden or demands such as the timing of doses. Theoretically, modern boosted PIs are good candidates for use as monotherapy as this class is known to have a high genetic barrier to resistance following viral failure, requiring several mutations before phenotypic drug susceptibility is significantly reduced.12 In PIVOT, the largest PI monotherapy trial to date, which included 296 patients in the monotherapy arm, 31% experienced virological failure.5 In the MONARK trial,13 the only PI monotherapy trial of ART-naive patients who were viraemic at baseline, the proportion that did not achieve sustained virological suppression was 33% by week 48 and 54% by week 96. Most trial protocols mandate a switch to cART in the case of virological failure and so there is little information on virological re-suppression on monotherapy. In the PIVOT study, of the 93 patients with virological rebound, 22 had spontaneously re-suppressed on subsequent testing and a further two re-suppressed after changing the PI but remaining on monotherapy.5 There were considerable differences among trials in the definition of emergent mutations, with some reporting all protease mutations and others only the mutations related to the specific PI agent used for monotherapy. The proportion of trial participants that developed emergent protease mutations when PI monotherapy was used as a maintenance strategy was on average <1% (range 0%–2.3%).15 Emergent resistance resulted in the loss of future PI options in 1% of patients in the PIVOT trial.5 There was a higher proportion of emergent protease mutations, 6%, in the MONARK trial, where PI monotherapy was used as first-line therapy.13Randomized controlled trials investigating PI monotherapy as a maintenance strategy indicate that most participants maintain viral suppression and the emergence of resistance-associated protease mutations is rare. Trials showed a wide range of virological failure when a threshold of plasma HIV RNA >50 copies/mL was used, varying from 5% to 53% over 48 or 96 week follow-up periods.16 Furthermore, intensification to cART following virological failure would only be possible if the patient could tolerate cART, which may not be the case in a real-world clinical setting. We conducted a review of patients who had received PI monotherapy at our centre in order to determine the clinical effectiveness of this strategy, the emergence of resistance and factors associated with virological failure. We show that virological failure and emergent protease mutations are more common in our cohort than in randomized controlled trials and other observational studies of European clinical settings.However, the results of these trials may not be generalizable to patients who receive PI monotherapy for clinical reasons. Trial participants had mostly already achieved virological suppression, did not have pre-existing protease resistance mutations and presumably were considered likely to adhere to a study protocol. Virological failure was not usually reported beyond 96 weeks, which limits the ability to extrapolate the results to the long-term success of this approach. PI monotherapy may not be as successful in patients with a history of treatment failure. The EARNEST study evaluated lopinavir monotherapy in patients who had failed first-line cART in resource-limited settings and found that 18% developed intermediate or high-level lopinavir resistance.A retrospective analysis of 10 years of prospectively collected data was conducted at a large HIV treatment centre in central London, UK. The study population was selected from HIV-1-positive patients attending the clinic between 1 January 2004 and 31 December 2013. Patients were included in the study if they were aged 16 years or over and had received at least 30 days of monotherapy with a boosted PI, and were excluded if they had been enrolled in a clinical trial of PI monotherapy. Data on demographics, antiretroviral treatment history, HIV RNA measurements and genotypic resistance testing were gathered from clinic records and the laboratory information management system.17 Archived genotypic resistance reports were available from 2004 onwards. The Stanford HIV Drug Resistance Database2 was used to determine HIV-1 subtype, identify baseline and emergent protease resistance-associated mutations and evaluate the impact of emergent mutations.HIV-1 RNA quantification and genotyping were performed using validated in-house methods.The primary endpoint was time from initiation of PI monotherapy to virological failure, which was defined as either after persistently detectable viraemia throughout the first 6 months of the PI monotherapy episode or virological rebound (two consecutive plasma samples with HIV RNA >50 copies/mL or a single plasma sample with HIV RNA >1000 copies/mL) at any point following viral suppression (at least one sample with HIV RNA <50 copies/mL). Other outcomes of interest were the times to: (i) viral re-suppression on PI monotherapy ; (ii) treatment discontinuation (switch to cART); and (iii) the emergence of protease resistance and loss of future PI therapeutic options (development of intermediate or high-level resistance to any PI was considered to be loss of that agent).Kaplan–Meier survival curves were plotted to describe the distribution of the times to virological failure and treatment discontinuation. The patients were stratified according to the presence of protease mutations at PI monotherapy initiation, baseline HIV RNA and the type of PI agent, to examine the effects of these factors. If patients had received consecutive periods of PI monotherapy with different PIs, i.e. changed PI agent but remained on PI monotherapy, this was considered as one continuous PI monotherapy episode for the main analyses. The individual periods of PI monotherapy with different agents were only considered separately when examining the effect of the type of PI on the virological outcome. The associations between factors and time to an event were tested using the log rank test, except when assessing type of PI, when Cox regression with a frailty term for patient was used to acknowledge the clustering of periods of different PI treatment for some patients. A univariate Cox regression analysis was performed to look for factors associated with virological failure. If any variables were significant then these would be included in a multivariate Cox regression analysis. Stata/IC 14 software  was used for all statistical analyses.In the UK retrospectively obtained anonymized NHS data do not require patient consent.One hundred and twenty-three patients received PI monotherapy at the study centre during the 10 year observation period. Twenty-eight patients were excluded from the analysis as they had participated in a PI monotherapy trial where use of PI monotherapy was not driven by clinical circumstances. A total of 95 patients that had received PI monotherapy for clinical reasons remained for the final analysis. Their characteristics are shown in Table PIs used for monotherapy were darunavir, atazanavir and lopinavir, all boosted with ritonavir. Seventeen patients received two different PIs as monotherapy at separate times during the observation period and one patient received three different agents, giving a total of 114 individual periods of PI monotherapy. The PI agent was changed during an uninterrupted episode of monotherapy in 17 cases, and these were treated as continuous PI monotherapy episodes. The reasons for changing PI were not known; 12 changed PI while virally suppressed and 5 while viraemic. Overall, 97 continuous PI monotherapy episodes were included in the analysis (two patients had two non-consecutive PI monotherapy episodes each). The median episode duration was 126 weeks (IQR = 96–174). At the end of the study period, 77 PI monotherapy episodes were ongoing, and 57 of those had viral suppression at the last HIV RNA measurement of the study period. No patients were lost to follow-up.P = 0.84), the baseline plasma HIV RNA (P = 0.90) and the type of PI (P = 0.47) did not affect the rate of virological failure  of episodes. Four were due to persistent viraemia and 58 had rebounded following a period of viral suppression. The overall time-to-event analysis is shown in Figure /97 of epViral re-suppression on PI monotherapy, without changing therapy, occurred in 38 episodes. This represents 61% (38/62) of all virological failure episodes and 68% (38/56) of those episodes in which PI monotherapy was continued for at least 1 month following virological failure. The median time to viral re-suppression after the initial date of virological failure was 14 weeks (IQR = 9–31), as shown in Figure Treatment was discontinued during follow-up in 21% (20/97) of PI monotherapy episodes. From Figure Genotypic resistance tests performed prior to PI monotherapy were available for 60 patients, of whom 29 had pre-existing protease mutations. Five patients had major mutations at baseline and 28 had minor mutations. Thirty-six patients had tests available from both before and after starting PI monotherapy. Eight patients  had emergent protease mutations in repeat testing that had not been detected before PI monotherapy .23 We did not find an independent association between CD4 count and failure, as did four of the observational studies.23 One study found an association between a nadir CD4 of <200 cells/mm3 and virological failure.19 Two of the trials, PROTEA14 and MODAt,15 found a low CD4 count was related to outcome, while MONET7 did not find an independent association.This is the first study, to our knowledge, to investigate the real-life use of PI monotherapy in a clinical setting where the use of PI-containing cART as first line is uncommon. There have been other observational studies in Spain and France, where PI-containing cART is more frequently used as first-line treatment, and our cohort otherwise had comparable baseline characteristics to those studies, with a preponderance of male patients, white ethnicity, MSM mode of transmission and subtype B infection, all typical of a European setting.23 We only investigated patients prescribed PI monotherapy for clinical reasons but one large multicentre study of clinical practice did not exclude PI monotherapy trial participants.19 When the indications for PI monotherapy were cited they were commonly treatment simplification or patient preference.23 This is in line with European and Spanish guidelines, which since 2009 cite PI monotherapy as an alternative strategy in the management of selected patients.25 French national guidelines have been more conservative, in 2010 deeming PI monotherapy to be inadequate, and in the 2013 iteration indicating that this strategy can be considered on a case-by-case basis.27 However, UK guidelines either did not mention PI monotherapy or expressly recommended against its use throughout the 10 year period applicable to the present study.28 PI monotherapy is not generally considered an acceptable treatment option in the UK unless there are extenuating circumstances that preclude the use of cART.29 It is our experience that such scenarios where PI monotherapy is employed include severely ill patients receiving intensive care who require a regimen with minimal systemic toxicity to control viral replication,30 or a patient struggling to comply with a more complex regimen where PI monotherapy is used as a holding strategy while further support and adherence interventions are attempted. A limitation of this study is the lack of complete data on the reasons for PI monotherapy and subsequent management decisions. However, we believe these reflect practical choices related to adherence and toxicity issues.There were some clear differences between our findings and the other analyses of PI monotherapy in clinical practice. In the other studies, plasma HIV RNA had to be fully suppressed for at least 6 months before PI monotherapy was commenced, whereas ∼40% of our patients were viraemic at PI monotherapy initiation.22 10%,21 14%18 and 31%.20 The cumulative proportion of episodes with virological failure in this study was around 30% by 48 weeks and around 50% by 2 years. A reason for the higher risk of virological failure than in most other studies, particularly in comparison with trial participants, may be that our cohort was less likely to adhere to therapy. Virological re-suppression was common in our study when patients remained on PI monotherapy following failure, suggesting that adherence was the main determinant of loss of viral control, and not lack of drug potency. The HIV-1 RNA level at virological failure did not differ significantly between those who did and did not go on to experience viral re-suppression. The phenomenon of re-suppression has also been observed amongst first-line cART-treated patients in resource-limited settings.31 Continuation of monotherapy despite virological failure at our centre probably reflects a pragmatic clinical decision based on concerns about adherence and dosing/toxicity issues associated with alternative regimens.Previous clinical practice studies reported 48 week virological failure rates of 4%,22 There was considerable variation in the frequency of genotypic resistance testing following virological failure. Between 20% and 66% of virological failure patients in the clinical studies had subsequent resistance testing, compared with 80% in our study. There were also differences in the reporting of mutations across studies. These two factors complicate comparisons of prior estimates of emergent mutations in clinical settings, which ranged from 0.4% to 2.2%, with our data (8%).23 There was no significant difference in study endpoints when the cohort was stratified according to baseline protease mutations, HIV RNA at initiation or PI agent. Two clinical practice studies found that a shorter period of virological suppression prior to PI monotherapy was associated with increased risk of virological failure.20 One of these also found that virological failure and treatment failure were more likely to occur in patients receiving atazanavir monotherapy, compared with those on lopinavir or darunavir.20 Another found lopinavir was associated with a greater risk of treatment discontinuation than darunavir; however, the follow-up period was much longer in this group.18 A further study of similar size to ours did not find any significant predictors of virological failure.21 One limitation of the present study is the relatively small sample size that could have resulted in insufficient statistical power to detect small differences among subgroups. However, our findings are in line with a large Spanish study, which did not find any significant effect of minor pre-existing mutations, PI agent or duration of previous viral suppression on the rates of virological failure in patients without major baseline protease mutations commencing either darunavir or lopinavir monotherapy.23In the three clinical studies reporting baseline genotypes, the prevalence of major protease mutations was between 3% and 7% prior to PI monotherapy, similar to the 5% baseline prevalence in this study.An advantage of the present study is that it investigates a real-world clinical setting with a pragmatic approach to the use of PI monotherapy in patients with less favourable baseline characteristics. However, these same attributes could limit the external validity for other settings in which PI monotherapy is to be used in stable patients with virological suppression and a good history of treatment engagement. We found high rates of virological failure but that if PI monotherapy was continued then virological re-suppression was often achieved. Emergent resistance was also more common than previously reported, as was the loss of future PI options. Despite this, the majority of PI monotherapy episodes were ongoing at the end of this study and over half were ongoing with viral suppression. Some of the discrepancies in outcomes compared with other studies may be a result of a longer observation period, thus capturing more events. It may be that prolonged follow-up of several years is required to understand the true outcomes of PI monotherapy in a clinical setting.Wellcome Trust, fellowships to R. K. G. (WT093722MA) and K. E. B. (170461), and the British Infection Society, fellowship to D. C. (E19115).This work was funded by the National Institutes for Health Research University College London Hospitals Biomedical Research Centre.We also received support from the K. E. B., D. C., E. N. and A. J. C. have no conflicts of interest. R. F. M. has received honoraria from Gilead, Janssen, Merck and ViiV for giving non-promotional lectures on clinical aspects of HIV, is a member of the British HIV Association TB/HIV Guidelines Committee and is a panel member for Guidelines for the Prevention and Treatment of Opportunistic Infections in HIV-Infected Adults and Adolescents National Institutes of Health, Centers for Disease Control and Prevention, HIV Medicine Association of the Infectious Diseases Society of America. R. K. G. has received honoraria from Janssen for giving non-promotional lectures on HIV drug resistance.


High levels of adherence are required to achieve the full benefit of ART. We assess the effectiveness of electronic adherence monitoring devices among patients failing second-line ART, as measured by viral load suppression. Cohort study of Wisepill™ real-time adherence monitoring in addition to intensified adherence counselling over 3 months in adults with a viral load ≥400 copies/ml on second-line ART in Johannesburg, South Africa between August 2013 and January 2014. Patients were sent SMS reminders upon missing a scheduled dose. We compared outcomes to earlier historical cohorts receiving either intensified adherence counselling or adherence counselling alone. Overall, 63 % of the participants (31/49) took >80 % of their prescribed medication; this dropped from 76 to 53 and 49 % at 1, 2 and 3 months post-enrolment respectively. Compared to those with good adherence (>80 %), participants with poor adherence (≤80 %) had a higher risk for a subsequently elevated viral load ≥400 copies/ml (relative risk (RR) 1.47 95 % CI 0.97–2.23). Participants found the intervention “acceptable and useful” but by 6 months after eligibility they were only slightly more likely to be alive, in care and virally suppressed compared to those who received intensified adherence counselling  or adherence counselling alone . In patients with an elevated viral load on second-line ART electronic adherence monitoring was associated with a modest, but not significant, improvement in viral suppression. The advent of antiretroviral therapy (ART) has led to significant reductions in morbidity and mortality –3. DespiSome patients demonstrating poor adherence go on to fail therapy, develop resistance and require more expensive subsequent treatment regimens –12. In mWhile several interventions to improve adherence exist , 15, mosWe conducted a pilot study to assess the effectiveness of EAMD in achieving adherence among adult patients with an elevated viral load (≥400 copies/ml) on second-line ART. Adherence was defined as achieving a suppressed viral load (<400 copies/ml) between 3 and 6 months after an elevated viral load (≥400 copies/ml) on second-line ART. We compare outcomes to two historical cohorts receiving the standard of care; the one cohort received intensified adherence counselling while the other received standard adherence counselling. In addition to generating preliminary estimates of effectiveness, we generate preliminary cost data and evaluate the feasibility of EAMD.This study was conducted among HIV-positive adult (≥18 years) patients receiving treatment at the Themba Lethu Clinic in Johannesburg, South Africa. The Themba Lethu Clinic (TLC) cohort has been described elsewhere . Themba By January 2014, Themba Lethu had initiated over 22,000 adult patients onto ART and of these 2790 have initiated second-line therapy. Of the patients that have initiated second-line ART, close to 20 % received intensified adherence counselling for an elevated viral load while on second-line ART.We conducted an ambi-directional cohort study comparing patients in the intervention cohort (EAMD in addition to intensified adherence counselling) to two historical comparison cohorts. Eligible patients were adult ≥18 years) HIV positive patients at Themba Lethu who were receiving a second-line ART regimen containing lopinavir/ritonavir or atazanavir/ritonavir and experienced a single elevated viral load (≥400 copies/ml) on second-line ART . These patients received routine re-adherence counselling with a counsellor or social worker and the viral load was repeated 3 months later. Genotyping and third-line drugs were not available during this period.The second, called the “intensified adherence cohort” included patients enrolled in the second-line ART clinic between July 2012 and the start of the intervention period (August 2013). Since July 2012 Themba Lethu has operated a second-line ART clinic focused on managing patients who are at high risk of failing second-line therapy . Patients are typically identified using an electronic medical record called TherapyEdge-HIV™. Patients with high viral loads are flagged for enhanced adherence counselling at their next clinic visit (1–2 months). The clinic manages these patients together during normal clinic hours however these patients bypass normal queues and undergo a longer intensified adherence counselling session than normal with an experienced counsellor or social worker. Patients then meet with a senior clinician that same day to address any adherence problems identified during counselling and additional issues pertaining to treatment side effects. Unlike the general clinic, attempts are made for patients to be seen by the same clinician at subsequent visits to maintain continuity and establish a patient-provider relationship. The patient is advised to continue their current regimen and return 3 months later to retest whether their viral load is suppressed. Patients that re-suppress (<400 copies/ml) their viral load at this point return to the normal clinic flow whereas those that fail to re-suppress are either sent for HIV genotype testing if the clinician suspects drug resistance or re-adherence counselling if the clinician suspects that adherence has not improved. Both resistance testing and third-line drugs  were available at the site after 2012 through a PEPFAR grant.Intervention cohort: In addition to intensified adherence counselling (which was the standard of care at the time), the intervention included the use of an EAMD until the first follow-up viral load test (3-6 months after enrollment). The intervention was designed to complement standard of care at the clinic, with little disruption to clinic duties or flow and apart from one additional study visit to coincide with routine medical visits.We piloted the intervention among eligible English speaking patients enrolled in the second-line clinic between 13 August 2013 and 22 November 2013 with a single recent (within the last 2 months) elevated viral load (≥400 copies/ml). Patients also had to have a cell phone and be willing to receive an SMS message of their choice if they missed a scheduled dose. Intervention cohort patients were identified and recruited from the second-line clinic. An EAMD, otherwise referred to as a Wisepill™ device, was offered to any patient that met the study eligibility criteria and provided informed consent.The Wisepill™ device is a portable rectangular box (110 mm long × 45 mm wide × 12 mm deep) which holds approximately 30 large or 60 small pills and contains a cell phone SIM card, a 1100 mAH lithium polymer rechargeable battery and a microchip to record time-date stamp within the unit . This wiAt enrollment (which coincided with intensified adherence counselling) study staff recorded participant and device information so that each participant could be linked to their adherence data and clinical information. Participants returned to the clinic 1 month after enrollment where study staff ensured proper use of the device . The clinician also reviewed the adherence data collected and had a focused discussion with the participant about their adherence pattern. This additional visit was not part of the usual standard of care that patients receive as part of the second-line clinic. When participants returned for the follow-up viral load test (3–6 months after enrollment) the device was returned and the clinician reviewed the adherence data with the patient. Finally, study staff administered a qualitative interview to determine the feasibility and acceptability of using the device. Questions addressed the participants opinion of using the device, including ease or difficulty of use; potential sigma and loss of confidentiality; and how useful they thought the device was to monitor adherence in real-time.For the intervention cohort, the proportion achieving adherence was calculated as the number of “recorded or actual” doses divided by the number of “scheduled (as prescribed)” doses. For this definition, “recorded” doses occurred when the device was opened (morning or evening) within a 3 h window either side of the scheduled time. If it occurred outside of this window period, the dose was defined as “missed”. Where multiple “recorded” doses were recorded in one period (morning or evening), the dose closest to the scheduled dose was considered and all other duplicates were disregarded. Required scheduled dose was calculated as the total number of days multiplied by the number of doses prescribed per day. The total number of days from enrolment was determined as follows: until study staff deactivated the device (definition 1), until the last event recorded (definition 2), or until the repeat viral load date (definition 3). We considered two cut-offs to define good adherence; either taking at least 80 % (missing ≥20 %) or taking at least 95 % (missing ≥5 %) of the prescribed medication .All patient-level data, including information on demographics, medications, laboratory test results and other clinical information were extracted from TherapyEdge-HIV™ prior to the analysis. Viral load testing is not done by the clinic, but by a central lab and therefore those performing the viral load tests were blinded to the study cohorts. Blood samples are sent to the National Health Laboratory Service (NHLS) and viral load and CD4 count results are uploaded directly into TherapyEdge-HIV™ from the NHLS on a daily basis . BecauseIn addition, we also estimated cumulative viral suppression by 12 months. For patients who were alive in care, not suppressed by 6 months (VL1), we followed patients for an additional 6 months (VL2) to determine if they could suppress by 12 months after eligibility.Continuous variables are described using medians with corresponding interquartile ranges (IQR) and categorical variables are expressed as simple proportions. For the intervention cohort log-binomial regression was used to estimate the association between poor adherence , was estimated using log-binomial regression. As the outcome was relatively common (incidence of ≥10 %), we calculated relative risks and corresponding 95 % confidence intervals (CI). In addition, we conducted a secondary analysis restricted to only patients with a repeat viral load. Models were adjusted for gender, first elevated viral load, time to elevated viral load and CD4 count at first elevated viral load. Loss to follow-up (LTFU) was defined as being ≥3 months late for the last scheduled visit with no subsequent visit. All-cause mortality was ascertained through patient tracing and linkage with the South African National Vital Registration System for patients with a valid South African national identification number (61 %) , 25.n = 49) to assess the feasibility and acceptability of using the EAMD. Participant responses were captured on a paper case report form and the raw data was transcribed and collated. Responses were categorized and coded into key themes, and those with similar content were summarized, with important quotes noted as has been described by Kleiman . Using different definitions of adherence  were recorded. No participant’s device had a signal at any point indicative that it could not pick up a signal or communicate with the server. A small number (13.1 %) of the drug intakes had a signal strength below 10  and 10.8 % of all drug intakes had a battery strength less than 3700 mV suggesting that the battery was low and needed recharging. Drug intakes were however still recorded even when the battery voltage dropped to the lowest at 3400 mV.We report good uptake of the device with only two patients who were offered it refusing to participate. Overall, 96 % of participants reported that Wisepill™ helped them to remember to take their medication. Most patients did not have a problem telling people what the device was when asked, one participant said he just told people it was a “machine that helps me remember to take my medication”. Participants reported that the device was “easy to carry around”, “it’s portable”, “I can keep all my tablets in one place”, “it reminds me to take my medication on time”, “looks like a modern day gadget”, “better than carrying packets of pills” and “it is a good box”. Participants felt that the device not only reminded them to take their pills when they forgot but it helped them to be more careful and responsible about taking their pills because they knew that someone was “watching” or monitoring them. On the negative side participants reported that “it would send reminders even when I took my medication”, “I didn’t like that it had to be charged”, “it is too big”, “people asked about the box” and “timing of SMS was inconvenient”.Participants suggested that the device should be combined with a reminder , preferably in one device. Participants also reported that there should not be long delays between forgetting to take their medication and receiving a SMS reminder. Some patients also complained about getting multiple reminders despite actually taking their medication. Participants also mentioned that the device could be smaller and that it should be designed to fit into trouser pockets for convenience, especially in men.Overall, participants were optimistic about the device and their responses indicated that they found the device “acceptable and useful”. One participant reported that “It was great to be reminded to take my medication” while another participant mentioned that “This box is very precious, it is a nice box. I don’t have to be reminded by my wife. I wish I can keep it”.We conducted a pilot study in Johannesburg, South Africa to determine if short-term use of electronic patient adherence monitoring devices can improve adherence in patients failing second-line antiretroviral therapy. By 6 months after eligibility, 44.9 % (95 % CI 31.4–58.9) of participants in the intervention cohort re-suppressed their virus without a regimen switch. This is similar to what has been reported for re-suppression on first-line ART in South Africa (40 %) . A signiDespite the fact that participants found the device “acceptable and useful”, Wisepill™ resulted in modest improvement in the primary outcome over intensified adherence counselling. We demonstrate that compared to those who received intensified adherence counselling, patients receiving combination intervention of electronic adherence monitoring and intensified adherence counselling were slightly more likely to be alive, in care and virally suppressed at 6 months after an elevated viral load on second-line ART (44.9 vs. 38.5 %), for an additional cost of 151 US dollars per patient managed. In order for the intervention to be considered, either the effectiveness would have to be improved  or the cost of the intervention would have to be dramatically reduced (<18 US dollars). We could not demonstrate that EAMD could significantly improve long-term adherence, as measured by viral suppression, but the results are encouraging since patients that re-suppress early are more likely to remain virally suppressed . Our finOverall re-suppression rates were low but expected given the study population. A recent study among patients experiencing treatment failure on protease-inhibitor based second-line ART showed that 46 % (51/111) of patients reach re-suppression at first follow-up viral load after an adherence support intervention similar to the intensified adherence cohort described here . Others Virological failure may be caused by a number of factors, including drug resistance, length of time on treatment and poor adherence , 30. We We also report that adherence dropped at 2 and 3 months post-enrolment. Recent evidence from this cohort has revealed social and behavioural factors including use of herbal/traditional medicine, alcohol and depression were significantly associated with failure to re-suppress viral load on second-line ART following intensified adherence counselling (results not shown). Therefore, interventions to address these barriers should be considered as part of the adherence intervention package to improve adherence over time , 34.Our findings should be considered in light of the study limitations. Firstly, compared to those in the Wisepill™ intervention cohort, the intensified adherence cohort and standard adherence cohort had a considerable amount of missing viral load data (25.8 and 25.2 % by 6 months and 12.7 and 5.0 % by 12 months). Patients with missing viral loads were considered failures , potentiSecond, we could not confirm adequate cellular signal at the participants’ home or work prior to enrolment in the study . We did Third, this study was not a randomized controlled trial aimed at measuring the effect of the intervention but rather a pilot study to generate preliminary estimates of effectiveness, generate preliminary cost data and evaluate the feasibility of EAMD. The pre-post design may result in secular trends or periodic variation. Study enrolment took place near the holiday period which may have resulted in a longer duration between counselling and the repeat viral load for some. To overcome this we used the first repeat viral load within 6 months of eligibility. Furthermore, some participants travelled out of South Africa and cell phone coverage did not include international roaming. These patients would have recorded missed drug scheduled doses in their adherence pattern and also would not have received SMS reminders when low battery was detected.Participation in a study may in itself motivate adherence. Patient motivation can be enhanced by free accessible care, approachable and supportive healthcare workers, broad social acceptance of ART, and past first-hand experiences with AIDS-related co-morbidity and mortality . AlthougLast, we restricted the intervention period to 3 months. It may be important to consider a longer intervention period (±6–9 months) for patients on second-line ART as adopted in other studies , 38. MurAdherence strategies increase the durability of second-line ART, decrease the need for costly third-line regimens and prevent unnecessary genotyping tests . Our fin


HIV treatment has reduced morbidity and mortality. By 2012, it was estimated that 60.4% of eligible South Africans accessed antiretroviral treatment; however, treatment adherence and retention remain the greatest challenges. There is a growing belief that social capital, seen as "the features of social organization that facilitate cooperation for mutual benefit", is important in promoting HIV treatment retention. The aim of this study was to establish whether social capital is associated with HIV treatment outcomes.This was a cross-sectional analysis of data from a cohort study that investigated how patient outcomes were linked to clinical characteristics, and included exploratory factor and logistic regression analysis. Data from 943 patients were analyzed. Outcomes for the analysis were visit non-adherence, unsuppressed viral load, and treatment failure. Sixteen percent of patients (n = 118) had unsuppressed viral loads; 19% (n = 179) were non-adherent; and 32% (n = 302) experienced treatment failure. Social capital had two dimensions that were described by two factors. There was no association between either factor and visit non-adherence. Social capital factor 1 was marginally associated with lower risks of unsuppressed viral load and treatment failure at 12 months ; but not with visit non-adherence . After controlling for confounders, the odds of both unsuppressed viral load and treatment failure decreased with an increase in social capital factor 1.This study suggests that social capital, in terms of the number of groups to which an HIV-infected person belongs, the diversity of the groups, availability of child support, and time available for community projects, is protective against poor HIV treatment outcomes. Implementers and policy makers in the areas of HIV treatment and prevention need to consider the inclusion of social capital in the design of HIV/AIDS treatment program. Social capital is important in promoting health globally. Putnam et al.  defined HIV treatment program have measurably reduced morbidity and mortality . South ASocial relationships are influenced by external events. In the case of HIV, the greatest challenge may be that of stigma . Social There is a growing belief that social capital is vital in promoting HIV treatment retention. Pronyk et al.  and otheThe need to explore the association between social capital and HIV treatment outcomes has been suggested in several reports –14. WoutWe aimed to establish whether social capital is associated with HIV treatment outcomes, particularly virological failure, in HIV patients on ART at 12 months in South Africa : A therapy used in treating HIV infection, usually given as a combination of three drugs—first line usually made up of two nucleoside reverse transcriptase inhibitors and one none-nucleoside reverse transcriptase inhibitor.Social Capital: The features of social organisation, such as trust, norms and networks that can improve the efficiency of society by facilitating coordinated action.Visit non-adherence: Fewer than 5 visits over the first 12 months after start of ART.Unsuppressed viral load: >400 copies/mL at 12 months after start of ART.Treatment failure: ≤ 400 copies/mL or missing viral load at 12 months after start of ART.This was a cross-sectional analysis of data from a large cohort study  examininClinics located in five provinces in South Africa  were chosen if they had more than 80 patients that had initiated ART during the 12-month study period. Of the 34 clinics, 17 were located in towns, 14 in cities and three in rural areas. Private solo practitioners ran 25 clinics, six were private group practices, and three were NGO clinics.The study participants consisted of HIV positive adults who were on ART for more than one month at the selected clinics from January 2006 to December 2010. Patients who attended a clinic visit (and were on ART for approximately 2 weeks to 2 months) during the study period for each of the selected clinics were included in the study (N = 1324).Patients were interviewed as they exited from their first clinic visit, after giving written informed consent to participate in the primary study.It is important to emphasize that no validated tool was available for measuring social capital at the time of the study. The questionnaire was drawn up from previous studies on ART adherence and perceptions around HIV and health , and on s included questions concerning each of the three categories of social capital and scores were assigned to responses. For example, “groups and networks” included questions on the household member's participation in various types of social organizations, and relationships with close friends. For “trust and solidarity”, questions considered if, in general, people can be trusted, and the levels of trust toward neighbors, strangers and other service providers. For “collective action and cooperation”, questions were asked about how members of a community or household worked on joint projects and/or in response to a crisis.Information was collected to determine social capital as defined by groups and networks; trust and solidarity; and collective action and cooperation. The questionnaireSocial capital was measured using eight categories see , viz.number of groups: the number of groups or organisations to which the participant belonged . One point was allocated per group, with a maximum score of five.group diversity: the diversity of the members of the different groups or organisations to which the participant belonged. One point was allocated per group, with a maximum score of six.friends: the number of close friends that the participant had. One point was allocated per friend, with a maximum score of five.trust: if the participant had trust in people in general. A score of zero was allocated for a negative response; five points was allocated for a positive response.level of trust: the level of trust someone had towards a category of people or groups like neighbors, strangers, national leaders or police. Points were allocated from one to five, based on a Lickert scale (from ‘no trust’ to ‘complete trust’)time: how much time someone had available to contribute to a community project. A score of zero was allocated to a negative response; five points was allocated to a positive response.availability of child support: if someone was at hand to care for the participant’s child/children. Two points were allocated for a relative being available; five points were allocated for a person who was not a relativeability to borrow money from someone, including friends and relatives. Points were allocated from one to five, based on a Lickert scale (from ‘definitely not’ to ‘definitely’).Because the Cronbach’s alpha for the eight categories was very low, viz. 0.23, indicating low internal consistency, exploratory factor analysis was carried out in order to find fewer variables to represent social capital.Routine data that were collected as part of the program, from patients at each clinic visit after initiation of ART, included demographic information and laboratory electronics records . Baseline CD4 count was any CD4 test done within one month prior to start of ART. Changes in treatment were also included in the analysis.The outcomes for the analysis were visit non-adherence, unsuppressed viral load, and treatment failure. Visit non-adherence was defined as fewer than five visits in the first year after starting ART. Visits following ART initiation were scheduled at 2 weeks, 6 weeks, 3 months, 6 months, 9 months and 12 months after the start of ART . Unsuppressed viral load was defined as HIV RNA viral load >400 copies/mL at 12 months after the start of ART. HIV RNA was assayed using polymerase chain reaction . Treatment failure was defined as either no viral load result or viral load >400 copies/mL after 12 months. We also ran a sensitivity analysis with different cut off values for treatment failure, i.e. <1000 copies/mL and <200 copies/mL. The three outcomes were dichotomized into visit adherence or non-adherence; suppressed or unsuppressed viral load; and treatment failure yes or no, respectively.Social capital was the main predictor variable. Potential confounders were age, gender, education level, employment status, socio-economic status (including composition of residence), wealth/income, and health .Data were analyzed using Stata version 12. Odds ratios in a logistic regression model were calculated for categorical variables and, where a trend was evident, a chi-square test for trend was conducted. The main variables of interest were social capital, defined by the two factors, and the three treatment outcomes. Variables that caused the effect sizes of the social capital factors to change by 10% or more were included as confounders of the three treatment outcomes in the final model. Associations between social capital and the treatment outcomes were also analyzed.This study was approved by the University of the Witwatersrand Research Ethics Committee (certificate number: M130249).Of the 943 patients with social capital information, all had treatment outcome data available. Most had viral load results recorded at 12 months  and data on visit adherence at 12 months . Of the 759 with viral load results, 641 (84.5%) had suppressed viral loads; 759 of the 938 (80.9%) were adherent (≥5 visits in the first year), and 302 (32%) of the 943 had treatment failure.Factor analysis suggested that social capital is explained by two factors . Factor Mean social capital scores at baseline are reported in Tables In the univariable analysis, the use of tenofovir in the regimen was associated with visit non-adherence . There was no association between non-adherence and either of the social capital factors. In the multivariable model, none of the variables was associated with visit non-adherence.Being older than 40 years appeared to be protective against treatment failure . The statistical model showed that the odds of treatment failure decreased with increasing social capital factor 1, i.e. the greater the group diversity, the more groups to which one belonged, and the greater the availability of child support, the lower the risk of treatment failure. There was no evidence of an association between social capital factor 2  and treatment failure.The sensitivity analysis using cut off values for treatment failure of <1000 copies/mL and <200 copies/mL did not change the statistical significance of the association between treatment failure and either social capital factor.Being older than 40 years was protective against unsuppressed viral load . However, the following factors were associated with an increased risk of having an unsuppressed viral load: being single , and use of a nevirapine in the regimen . When the model was adjusted for age and whether or not the patient was using a second line of treatment, there was an association between factor 1social capital and unsuppressed viral load .This is the first study to measure the association between social capital and ART outcomes. Social capital factor 1 was associated with a lower risk of unsuppressed viral load and a lower risk of treatment failure at 12 months, but not with visit non-adherence. After controlling for confounders, the odds of both unsuppressed viral load and treatment failure decreased with increased social capital. This study is the first to show that social capital is linked to a lower risk of having an unsuppressed viral load for individuals on ART. In addition, we found that demographic factors associated with high social capital were older age and being employed.These findings are in keeping to those from studies by Cattell et al.  and KawaIndividuals with middle and higher social capital had a lower risk of treatment failure. This situation is similar to that reported in a study on cancer patients in the US . By talkSocial capital is a difficult concept to understand as it is measured in different ways and has a variety of definitions. Several studies have identified useful proxy measurements for social capital using different types and combinations of qualitative, quantitative and comparative research methodologies. This is one of the few studies that have looked at social capital and HIV treatment outcomes in South Africa, and the only one that has used a method of scoring social capital to show its effect on HIV/AIDS patients with regard to treatment outcomes.Our study had some limitations. The first was the timing of the interviews. Social capital was measured at one-time point, but outcomes were measured at 12 months after the start of ART. There was an assumption that the social capital measured at the starting point remained constant while the person was on ART. Nevertheless, the study design provided prospective data on a sample of patients that would not have been available in a cross-sectional study. Another limitation was that we included only patients who were at the study clinics and available for interview. We are therefore likely to have overestimated treatment success due to selection bias. However, our study was not aimed at understanding treatment success but rather at understanding social capital and its effect on treatment success. In addition, as we did not have validated tools for social capital, we used modified social capital scores. This made it difficult to compare the findings with those from other studies.We used visit adherence as a proxy for adherence to treatment as early studies show significant impact of social responsibility upon treatment adherence. We also included viral suppression, as this is the gold standard used to measure treatment adherence. We used a composite outcome of unsuppressed viral load and missing viral load or no viral load result to account for those who did not return for the viral load results as treatment failure. We assumed that the reason for missing viral load results was that patients did not return for their viral load visits, but there might have been other reasons, such as problems with the laboratory and the data management system.At the same time, our review of different studies where most of researcher points on several gaps concerning validity of social capital scores. We did not have validated tools for social capital. In this study, we assumed that the reason why higher social capital was associated with older age could be that our study included 18 years and older. Further research should be done in younger groups and using validated tools.The results of this study show that, as the level of social capital increases, treatment outcomes improve. Most probably, this is linked to having the practical means to assist with the ability to continue taking treatment, assistance with child support, being part of a group or being part of a community project. In addition, it may be that HIV-infected people with high social capital tend to develop confidence, and that support results in improved treatment adherence and retention in the programme.Implementers and policy makers involved in HIV treatment and prevention need to consider social capital in the design of HIV/AIDS treatment programmes and encourage support groups and other mechanisms for improving social capital. The association between social capital and employment also provides further evidence for the benefits of increased employment opportunities.S1 File(DOCX)Click here for additional data file.S2 File(DTA)Click here for additional data file.S3 File(DTA)Click here for additional data file.S4 File(DTA)Click here for additional data file.


Approval of Ledipasvir/Sofosbuvir for the treatment of chronic hepatitis C (HCV) includes the truncation of therapy from 12 to 8 weeks in treatment naïve, non-cirrhotic patients with baseline HCV RNA levels <6 million IU/mL (6.8 log10 IU/mL). The aim of this study was to evaluate this clinical cutoff with a different widely used commercially available HCV RNA test. Results from samples tested prospectively with Roche High Pure TaqMan HCV 2.0 test (HPS) were compared to those tested retrospectively with the Abbott RealTime HCV RNA test (ART). Using 6 million IU/mL as the cut-off, pre-treatment results were concordant in 70.4% of cases. When results with the same test measured at screening and baseline, clinical decisions could be impacted in 14.4% and 6.2% of cases for HPS and ART respectively. Using only HCV RNA cutoff of 6 million IU/mL, 29.55% of subjects would receive a different and potentially incorrect treatment duration based solely on HCV RNA test method used. A further 6–14% of subjects would have treatment decision change based on the day the sample was taken. Worldwide it is estimated that 64–103 million people are chronically infected with the hepatitis C virus (HCV)10 IU/mL) based on the findings of the ION-3 trial121415The recent approval of all-oral direct acting antivirals (DAAs) targeting viral proteins involved in the virus life cycle deliver very high rates of sustained virologic response (SVR) with much improved side effect profiles and have revolutionized the treatment of chronic HCV456789In this study we evaluate this new clinical cutoff (6 million IU/mL) used to shorten LDV/SOF therapy. A secondary aim of this study is to investigate the performance of highly sensitive, automated, commercially available HCV RNA viral load assays and the impact of detecting low levels of HCV RNA at EOT and post treatment week 12.A total of 2903 samples from 631 subjects who enrolled in AbbVie’s SAPPHIRE I Phase III trial were included in this study . SAPPHIR19To evaluate the potential impact of quantitative viral load bias reported by HPS and ART around the 6 million IU/mL clinical cutoff, the results from pre-treatment samples with values for both methods (n = 741) were compared for concordance. These samples represented 627/631 from screening and 113/631 from baseline. The interval from Screening to Baseline in this study averaged 23 days with a minimum of 7 days, a maximum of 92 days and an overall standard deviation of 12.88 days. The potential impact of natural viral fluctuations prior to the initiation of treatment was explored where possible, by comparing results from subjects sampled at screening and baseline with the same viral load method for concordance at the 6 million IU/mL cutoff. Screening and baseline viral load results tested with the same method were available for 631 subjects for HPS and 113 for ART. The number of data points available for analysis with ART was lower due to insufficient residual sample volume. To understand how viral loads measured by these two methods and any bias between them vary over the dynamic range of both assays, correlation was measured at key intervals and the data adjusted for the bias was plotted and a linear regression performed. To investigate differences in detection at the low end of the assay dynamic range, samples from all subjects and timepoints with sufficient volume were tested with ART (n = 2903) and concordance measured against the test of record, the HPS method, with 25 IU/mL and 100 IU/mL as the cutoff for detection, respectively.t test was performed to test whether the mean bias was statistically different from zero. P values <0.05 were considered to be statistical significant. In order to visualize the differences between the two assays a Bland-Altman bias plot of the bias versus the average value of the two assays was generated.All data were analyzed with PC SAS 9.3 . A least squares linear regression was performed to examine the association between the HPS assay result (log IU/mL) and the Abbott RealTime HCV result (log IU/mL). The data were plotted and the Pearson correlation coefficient, the coefficient of determination, the point estimates for the slope and intercept of the regression line were calculated. Additionally an analysis of the bias between the two assays was performed. For each specimen, the bias was defined as the difference between the ART HCV and HPS results. The summary statistics of the difference were calculated  along with their confidence limits. A All patients provided written informed consent relevant to the use of the current study.10 HCV RNA IU/mL. A Bland Altman plot analysis is shown in A total of 2903 paired HCV RNA level results from the 631 patients enrolled in the study were obtained. There were 1011 specimens with detectable HCV RNA results that were within the dynamic range for both tests; significant correlation between the 2 measurements was observed  were compared, clinical decisions (treatment duration) could be impacted in 14.4% cases for HPS (8.1% extended and 6.3% truncated) and 6.2% for ART (3.5% extended and 2.6% truncated). The mean difference in viral load between these two time points with the same test was between 0.07 log IU/mL and −0.02 log IU/mL for HPS and ART respectively.The correlation between ART and HPS measured at various intervals accross the dynamic range of HCV viral loads seen in this study revealed that the ART-HPS bias went from +0.30 log IU/mL for viral loads <2 log IU/mL (n = 28) to −0.56 log IU/mL at viral loads greater than 6.6 log IU/mL (n = 428). The linear regression of the data adjusted for mean bias at key viral load intervals can be seen in The approved label for sofosbuvir/ledipasvir indicates that treatment naive non-cirrhotic patients with a baseline viral load <6 million IU/mL are eligible to short therapy of 8 weeks while all others should receive a full 12 weeks of therapy1220212223252628In this study, using only HCV RNA cutoff of 6 million IU/mL, 29.55% of subjects who would be eligible for treatment truncation based on the HPS results would not have been eligible based on the ARTresults. A further 6–14% of subjects would have treatment decision change based on the day the sample was taken due to natural fluctuations in pretreatment viral loads. When analyzing these natural fluctuations it should be noted that there were greater number of cases tested at both time points by HPS than by ART which may have some impact on the differences seen between methods.The results of this study also support published findings of other investigators that the ART HCV RNA assay is a more sensitive test and detects virus in a higher number of patients on treatment, that the positive bias in quantitation between ART and HPS will more likely result in a quantifiable result at the low end of the dynamic range with ART and that the number of concordant undetectable results significantly increases with time on treatment1516171819202122232425In this study, using only HCV RNA cutoff of 6 million IU/mL, 29.55% of subjects who would be eligible for treatment truncation based on the HPS results would not have been eligible based on the ART results. A further 6–14% of subjects would have the treatment decision changed based on the day the sample was taken due to natural fluctuations in pretreatment viral loads. The patient profile of subjects enrolled in SAPPHIRE I and the number of subjects with baseline viral loads <6 million IU/mL using HPS (64.5%) compares very closely with the ION-3 registration trial (59%) used to establish this treatment truncation rule. It will be important to follow-up on real-life data that are based on ART and other automated platforms such as the COBAS® AmpliPrep version COBAS® TaqMan assay  measurements to determine whether the potentially higher number of patients eligible for shortened treatment durations will result in a higher risk for relapse. The mean bias around the 6 million IU/mL clinical cutoff of 0.61 log IU/mL observed in this study would imply that a viral load of between 1.5 and 2.0 million IU/mL with ART (6.18–6.30 log IU/mL) would be equivalent to 6 million with HPS (6.78 log IU/mL). This threshold is supported by ROC analysis using all 741 available pre-treatment data points and HPS as truth/gold standard where the intersection of the sensitivity (>6 M) and specificity (≤6 M) is 6.19 log IU/ mL. In another study comparing CAP/CTM with ART the bias at 6.78 log IU/mL was smaller (0.41 log IU/mL vs 0.61 log IU/mL) which clinically significant diferences also exist between the manual version of the test used in the clinical trials (HPS) and the automated versions widely used in the field (CAP/CTM)Current guideline recommendations based on an absolute quantitative baseline HCV RNA value derived from a single test not widely used in clinical practice may be inappropriate and may result in misclassification with regard to this clinical cutoff. At the minimum, ranges of HCV RNA levels should be used and ideally stated for all commercially available assays.The findings of this study would supports those of other investigators which demonstrated limited or no utility for very sensitive, quantitative viral load testing at baseline to inform treatment duration and at end of treatment as an indicator of subsequent SVR17Limitations of this study include the fact that ART results were generated retrospectively and that there were greater number of cases tested at both time points by HPS than by ART.How to cite this article: Cloherty, G. et al. Hepatitis C RNA assay differences in results: Potential implications for shortened therapy and determination of Sustained Virologic Response. Sci. Rep.6, 35410; doi: 10.1038/srep35410 (2016).