As part of efforts to improve the prevention of mother-to-child transmission in Northern Uganda, we explored reasons for poor viral suppression among 122 pregnant and lactating women who were in care, received viral load tests, but had not achieved viral suppression and had more than 1000 copies/mL. Understanding the patient factors associated with low viral suppression was of interest to the Ministry of Health to guide the development of tools and interventions to achieve viral suppression for pregnant and lactating women newly initiating on ART as well as those on ART with unsuppressed viral load.A facility-based cross-sectional and mixed methods study design was used, with retrospective medical record review. We assessed 122 HIV-positive mothers with known low viral suppression across 31 health facilities in Northern Uganda. Adjusted odds ratios were used to determine the covariates of adherence among HIV positive mothers using logistic regression. A study among health care providers shed further light on predictors of low viral suppression and a history of low early retention. This study was part of a larger national evaluation of the performance of integrated care services for mothers.Adherence defined as taking antiretroviral medications correctly everyday was low at 67.2%. The covariates of low adherence are: taking other medications in addition to ART, missed appointments in the past 6 months, experienced violence in the past 6 months, and faces obstacles to treatment. Mothers who were experiencing each of these covariates were less likely to adhere to treatment. These covariates were triangulated with perspectives of health providers as covariates of low adherence and included: long distances to health facility, missed appointments, running out of pills, sharing antiretroviral drugs, violence, and social lifestyles such as multiple sexual partners coupled with non-disclosure to partners. Inadequate counseling, stigma, and lack of client identity are the frontline factors accounting for the early loss of mothers from care.Adherence of 67% was low for reliable viral suppression and accounts for the low viral suppression among HIV-positive mothers studied, in absence of any other factors. This study provided insights into the covariates for low adherence to ART and low viral load suppression; these covariates included taking other medications in addition to ART, missed appointments in the past 6 months, feels like giving up, doesn’t have someone with whom to share private concerns, experienced violence in the past 6 months, and faces obstacles to treatment and confirmed by health providers. To improve adherence, we recommend use of a screening tool to identify mothers with any of these covariates so that more intensive adherence support can be provided to these mothers. Human immunodeficiency virus (HIV) causes a chronic infection that leads to a progressive disease. Without treatment, most persons with HIV develop acquired immunodeficiency syndrome (AIDS) within 10 years of infection, which results in HIV transmission to sex partners, substantial morbidity, and eventually death  don’t have real husbands. They are up and down. That’s why you will not get them easily because they are not in one place,” FGD participant, Nwoya.“Most of them [mothers] lack support of their partners, most of them are not educated and earn a living by selling food stuff that they grow,” FGD participant, Nwoya.“Many of the newly initiated mothers [on ART] have disclosure problems and sometimes don’t disclose their [HIV] status to their spouses,” FGD participant, Kitgum.“These are mothers who have not disclosed their [HIV positive] status to their husbands. They are mothers whose husbands are not aware that their wives are taking drugs…,” FGD participant, Lira“Mothers come from poor families. They lack transport and move on foot. They take drugs on empty stomach. There is gender based violence, [and when] husband chases the woman away, [and] they never come back for refill. Some would [could] stay 2 months without refill,” FGD participant, Gulu.These comments resonate with the social demographic characteristics described in Table The practice of assessing adherence had mixed responses. Some providers reported making use of appointment dates or pill count if the mother comes with a bottle, while others depended on self-recall by the mother on the number pills left. In some cases, providers reported to use both methods concurrently. However, it was also common for health providers to qualify their statements by adding that the heavy workload and long queues made it difficult for them to count pills as a way of assessing adherence.“We ask mothers to come with the remaining pills, but majority do not come with their pills and in this case, we use appointment dates,’’ FGD participant, Lira.“The good or fair assessment of adherence in registers is not real because we don’t delve into pill count because of the large numbers [of clients]…,’’ FGD participant, Gulu.“From the ART card there is that column for adherence where if a mother comes for refill we normally ask them looking at the appointment. Then we ask them of the number of the pills they have swallowed,’’ FGD participant, Apac.At a FGD in Awach, all providers agreed that the adherence rating on record in the registers was not accurate because it was not based on real adherence analysis but rather on the provider’s quick guess for purposes of recording. This was not so different with other providers elsewhere who mostly attributed this to the workload on any particular clinic day. There was mention that adherence cards which were used to assess adherence in the past are now stocked out. The following quotations support these findings.“I have one mother who told me that her husband does not allow her to take her medicine. So she hides and whenever the man discovers that she has the drugs he throws it in the pit latrine. And right now she has shifted to her mother’s place,’’ FGD participant, Apac.“Some mothers think they are healthy. So [they see] no need to take ARVs. When they get psychological reconstruction, they feel they are healthy,’’ FGD participant, Gulu.“Poor adherence [is a result of] some of the women combine [combining] drugs for morning and evening and take [taking] them once in the evening,’’ FGD participant, Kitgum.Health care providers commonly shared, on the basis of their own experience, that mothers miss their appointments and therefore run out of medicine, they share drugs between themselves and/or with their husbands when either party is unable to come to the facility, and that they stop taking drugs whenever they get new unsuspecting partners. Participants often cited various forms of distress including violence from husbands as prohibitive to mothers taking drugs. It was common for health care providers to state that mothers miss taking drugs especially when they have travelled away, while others think they have recovered from ill health and need not take drugs anymore. The following quotations represent the most common sentiments of providers with regard to mothers’ adherence.“…there’s one who stays in [the] village,  want to take their drugs [away from them claiming that]. …..That this transport you are wasting you can use for paying [school fees for] children in school after all you will still die even if you swallow the medicine,’’ FGD participant, Apac.“I have a client who kept piling her drugs because [she] believed that the drug will [would] kill her,’’ FGD participant, Lira.“Other mothers mistake Septrin for ARVs such that they are confidently swallowing cotrimoxazole in place of ARVs e.g. [we see this] when they run out of stock,’’ FGD participant, Lira.“…Pill burden, especially for those with other OIs [opportunistic infections] like TB,’’ FGD participant, Kitgum.A key area of exploration in the qualitative component of the study was to obtain providers’ perspectives about why mothers were not suppressing. Common reasons why the respondents believed mothers don’t suppress included: missing appointments and dropping out of care for some time without close monitoring, taking drugs erratically on the account of recovery and good health, torture and violence from their partners, or over time, failing to respond well to the medication. Long distances to the health facility and lack of transport also keep mothers away from care. The shortage of food affects drug use, as mothers will not take drugs on an empty stomach. Similarly, negative perceptions about ARVs, mothers experiencing side effects and stopping taking the drugs, or using them occasionally as and when they feel unwell, and stigma and discrimination keep clients in self-denial and affect adherence and appointments. Representative comments from providers in this regard:“They use common names in villages like maama John, and follow-up is not easy [with such names],’’ FGD participant, Gulu.“Disclosure to husband will cause divorce and cause mother [to] move back [be chased away] to her parents. So she keeps quiet and disappears [from routine care],’’ FGD participant, Apac.We inquired from health care providers why mothers drop out of care just 3 months after initiation on ART. They identified as contributing factors, that mothers lack proper identifying information for follow-up and non-disclosure to their partners for fear of reprimand, violence or separation. The following quotes provide an account of this phenomenon:“Sometimes midwives counsel them, but not to [their] expectation, especially that the [HIV] positive person needs a lot of time,’’ FGD participant, Gulu.“It’s the pregnancy that will lead them [to] know their status and once they know, it’s shocking and [they end up] leave [leaving] the facility. HIV status is detected only during ANC so it’s a shocking moment,’’ FGD participant, Gulu.“Some mothers will enter denial stage and go to another health facility. So one client can be registered twice, [and this is] common in town,’’ FGD participant, Kitgum.“[They have for the] Disbelief of the results and [they] will deny the results and they will go away….,’’ FGD participant, Gulu.Results of the study among HIV-positive mothers revealed that 17% of the 122 mothers never received adherence counseling before initiation on ART. This was confirmed by providers as follows:This study collected data on different variables from HIV-positive mothers with known low viral suppression only, since looking at these characteristics in all HIV-positive women would bias results; given that their characteristics may not be similar or have the same distribution in the women with good viral suppression.Unfortunately, the rate of adherence in our sample was significantly lower 67.2%) than has been observed in many other studies in Africa, even when it was self-reported adherence. For example, a study on adherence and low viral response in Kisumu, Kenya found adherence at 87.1% among women on Option B+ [7.2% thanUp to 23% of women in this study had multiple sexual partners. This was corroborated by health care providers that most women who have not achieved the required suppression have unstable relationships and were less likely to disclose their HIV status to their non-regular partners for fear of breakdown of their relationship. They were more prone to violence and vulnerable to discrimination by others even before the effect of other socio-economic conditions that generally affect women on ARVs. These could be termed as frontline factors, both of which could be used to explain this finding and were corroborated by the health care providers.The cultural context in Northern Uganda, a post-conflict region, is such that women are sexual subordinates and most often responsible for care of the household and children, and as a consequence, these practical barriers may adversely affect adherence among women. Perhaps, these among other factors, make the sample not comparable with women in other studies whose socio-economic variables could have different levels of influence on treatment adherence.Another key adherence barrier raised during the interviews included violence from their partners. Our findings reinforce the importance of partner counseling, including obtaining remedies for victims of violence whenever they exceed the confines of counseling.Adherence counseling prior to initiation of ART was associated with consistency in taking drugs. It was further discovered that mothers who had missed appointments in the last 6 months were more than two times less likely to adhere to their treatment than HIV-positive mothers who never missed their appointments at all. Adherence counseling before ART is one way to determine readiness to start treatment, diagnosing and addressing stigma, depression, and initiating a patient-provider relationship to even understand the patient better . Such a Even HIV-positive mothers who were treated discriminately since disclosure of their HIV status were only slightly less likely to take their drugs consistently as compared to those who were not discriminated/stigmatized. It is possible that patients with limited literacy might be reluctant to ask others for the kind of help they need to take their medicines correctly. Better-educated patients who are convinced of ART efficacy, perhaps as a result of educational programs, show a propensity towards better adherence . TherefoPatients’ beliefs, knowledge, and expectations regarding treatment strongly influence medical decision making . Our finAlthough ART is free of charge throughout Uganda’s public health system, there are ART-associated costs such as expenses in transport and food. These obstacles were cited by mothers and corroborated by health care providers. Mothers cited skipping medication because of lack of food or went without medication because they could not reach the health facility. This is consistent with other studies which have linked costs with poor adherence to treatment .A study in Botswana showed that adherence was higher when cost was not an obstacle and that poor patients were able to achieve excellent rates of adherence when they had access to free laboratory monitoring and subsidized ART . This isIn the FGDs, participants raised the issue that mothers were worried of disclosing their status to partners for fear of losing them. As such they either take drugs in hiding or cease taking them completely. Like many other studies, non-disclosure of HIV status was associated with poorer adherence. Our findings suggest unless mothers disclose, they cannot effectively take their drugs. This could perhaps explain the high rates of missed appointments (53.7%). This is a fundamental issue that disclosure health education and individual counselling sessions should embrace with utmost importance.Distance to health facilities was a common cause of missed appointments, as was drug stock-out as reported by mothers and validated by health care providers in the FGDs. Over half of the mothers come as far as 10 or more kilometers to the health facilities and find the transport cost prohibitive. This coupled with the seasonal deterioration of poorer roads during the rainy season makes it even harder to access facilities. Deliberate outreaches in hard-to-reach areas would offset this limitation as much as possible.ART regimens have toxicities and adverse side effects that vary from mild to severe and acute to chronic, which can prevent adherence . Our stuFood was an important barrier to medication in this study. Previous research has shown a relationship between food insecurity (uncertain access to safe and nutritious food) and lower rates of viral suppression among ART-treated patients in the US and Uganda. Pregnant and breastfeeding women may be especially vulnerable to food insecurity and insufficiency .Adherence of 67.2% among mothers with low viral suppression in this study is very low. The main covariates were distance to travel to health facility leading to missed appointments and running out of drugs, sharing antiretroviral drugs, drug side effects, lack of pre-ART counseling, gender-based violence, social lifestyles such as multiple sexual partners, non-disclosure to partners, and stigma and discrimination. Qualitative findings from providers corroborated these findings. To improve the quality of care provided to mothers, the information from this study can be used to screen pregnant women and lactating mothers on ART for any of the factors strongly associated with low adherence to ART  and then provide support through intensive adherence counselling and self-management or peer-to-peer support.


HIV-1 infection impairs tuberculosis (TB) specific immune responses affecting the diagnosis of latent TB. We aimed to (1) determine the proportion of HIV-1-infected adults with a negative QuantiFERON®-TB Gold in-tube (QFT-GIT) and Tuberculin skin testing (TST) that convert to QFT-GIT positive following TST, and (2) evaluate the relationship between conversion and baseline QFT-GIT results.HIV-1 infected adults being screened for a TB vaccine study in South Africa underwent QFT-GIT followed by TST. As per protocol, QFT-GIT was repeated if randomization was delayed allowing for evaluation of TST boosting in a proportion of participants.3 IQR 439–621) who had QFT-GIT repeated after median 62 days (IQR 49–70), 40.9 % (95 % CI 18.6–63.2 %) converted. Converters had a significantly greater increase in the background subtracted TB antigen response  following TST, 0.82 (IQR 0.39–1.28) vs 0.03 (IQR −0.05–0.06), p = 0.0001. Those who converted also had a significantly higher baseline TBAg-Nil 0.21(IQR 0.17–0.26) vs 0.02(IQR 0.01–0.07), p = 0.002. Converters did not differ with regard to CD4 count or ART status. ROC analysis showed a baseline cut off of 0.15 correctly classified 86.4 % of converters with 88.9 % sensitivity.Of the 22 HIV-1 infected, TST and QFT-GIT negative adults  contains supplementary material, which is available to authorized users. Mycobacterium tuberculosis (Mtb) in an otherwise asymptomatic person, either by Tuberculin skin testing (TST) or Interferon gamma release assays (IGRA)  and T-SPOT.TB) [The diagnosis of latent tuberculosis (TB) relies on the demonstration of an acquired cell-mediated immune response to SPOT.TB) .HIV-1 infection results in the depletion of Mtb specific CD4 cells  and impaTB > 5 spots) regardless of HIV-1 status. HIV-1 co-infection however has been shown to result in a reduced sensitivity of both IGRA platforms for tuberculosis [Unlike TST, IGRAs have adopted a single cut-off , or above 300/mm3 with suppressed HIV viral load, if receiving ART, were eligible for the vaccine trial. As part of the screening process all participants were assessed for active TB by symptom screen, chest radiograph (CXR) and sputum culture and for latent TB by QFT-GIT followed by TST. Those with no evidence of active or latent TB were eligible for randomization into the vaccine trial. However, if > 45 days had elapsed since screening initiation, the protocol determined that participants should be rescreened prior to randomization. This included repeating the QFT-GIT. In March 2012 there was a brief sponsor initiated suspension in recruitment to the vaccine trial. As a result when recruitment was reinitiated in April 2012 a number of screened participants breeched the 45-day rule and required rescreening. This pause in recruitment allowed assessment of boosting of the response following the initial TST presented here. The study was observational and did not have a control arm.The study was conducted, in Khayelitsha, Cape Town, within the screening period of a TB vaccine trial, which enrolled participants between August 2011 and April 2013 . The stuTST was administered as an intradermal injection of 2TU (0.1ML) of PPD-RT23  into the volar aspect of the forearm and performed immediately after blood had been drawn for baseline QFT-GIT. Maximal transverse induration was measured at 48–72 h by clinical research nurses trained to do so. Induration < 5 mm was considered negative.The QFT-GIT assay  was performed in accordance with manufacturer’s instructions. The following measures were implemented in order to minimise variation in assay results: (1) All samples were taken by research nurses trained and assessed to ensure consistent sampling methodology. (2) Following blood draw samples were kept in temperature-logged insulated transport boxes and samples were incubated within 4 h of blood draw. (3) Incubation time was standardized to 24 h. (4) QFT-GIT assay was performed by Good Laboratory Practice (GLP) trained research technicians in a Foundation of Innovative New Diagnostics (FIND) accredited laboratory with regular sponsor mandated Quality Assurance (QA) undertaken to ensure reproducibility of results.U test and parametric data compared using t-test. Non-parametric paired data was compared by Wilcoxon signed-rank Proportions were compared by χ2 test or Fisher’s exact test (if the contingency included a number ≤ 5). Receiver Operating Characteristic (ROC) analysis was performed as a non-parametric test.Statistical analysis was conducted in Stata ver. 12.1 (StataCorp). Gaussian distribution of data was determined by the Shapiro-Wilk test. Non-parametric independent data was compared using Mann-Whitney 3 (IQR 439–621). All participants had negative sputum culture and a CXR without evidence of active TB.Twenty-two HIV-1-infected adults screened for the vaccine trial with a negative TST and QFT-GIT required rescreening (due to brief suspension in recruitment) and therefore had a repeat QFT-GIT performed at a median of 62 days (IQR 49–70) after the TST. The TST was 0 mm in all participants. 90.9 % of participants were female and all were resident in Khayelitsha, a peri-urban township of Cape Town. Median age was 33 years (IQR 29–40). 16 participants (73 %) were on ART for a median of 2.0 years (IQR 1.5–3.5) and had a suppressed VL (either lower that detectable limit (LDL) or <40 copies/mL). Seven participants (31.8 %) had been treated for active TB a median of 3.7 years previously (IQR 2.0–4.2). Median VL for those not on ART was 17,309 copies/ml . Median CD4 count for the 22 participants was 477/mmp = 0.002) as was the change in TBAg-Nil post TST, 0.82 IU/ml (IQR 0.39–1.28) compared to 0.03 IU./ml , respectively. The repeat TBAg-Nil, post TST, was significantly higher in converters (0.84 IU/ml (IQR 0.65–1.50), p = 0.008) but not in non-converters 0.06 IU/ml (IQR 0.00–0.10), p = 0.86  demonstrated conversion from negative (<0.35 IU/ml) to positive QFT-GIT during the interval between tests. There was no significant difference in CD4 count, age, sex, proportion on ART, previous TB treatment or time between initial and repeat IGRA between those that converted and those that did not  of baseline TBAg-Nil to identify converters was 0.87 (95 % CI 0.69–1.00). TBAg-Nil cut-off values of 0.15 IU/ml and 0.17 IU/ml both correctly classified 86.4 % of converters, with 0.15 IU/ml cut-off having a higher sensitivity (88.9 % vs 77.8 %) and 0.17 IU/ml having a higher specificity (92.3 % vs 84.6 %)  and TST (<5 mm) convert to QFT-GIT positive (TBAg-Nil ≥0.35 IU/ml) within 7–10 weeks of a TST. We have also demonstrated that converters have a higher baseline QFT-GIT TBAg-Nil and that a baseline cut-off value of 0.15 IU/ml correctly classifies 86.4 % of converters with 88.9 % sensitivity. This rate of conversion is considerably higher than in previous studies conducted in a variety of settings with low HIV prevalence .Although our study was conducted in a setting of high TB incidence it is unlikely that the 40 % conversion rate represents acquired infection over a 7–10 week period. The estimated force of infection in this setting is less than 8 % per year, therefore the expected rate of new infection over a 7–10 week period would be less than 1.5 % . FurtherConverters in our study had a significantly higher baseline TBAg-Nil than non-converters (0.21 vs 0.02). In addition to this, following TST, converters also had a significantly greater increase in TBAg-Nil with non-converters generally showing minimal change post TST 0.82 vs 0.03) demonstrating that TST did not just simply result in a uniform boosting in IFNγ production in all participants. The low baseline response and minimal boosting in non-converters in our study are similar to the responses in HIV-uninfected participants that are TST and QFT-GIT negative (<0.35 IU/mL) at baseline. Sauzullo et al. found 69 TST negative Italian healthcare workers to have QFT-GIT TBAg-Nil <0.03 IU/ml and <0.08 IU/ml before and 6 weeks after TST, respectively 2 vs 0.03.ex vivo when the repeat QFT-GIT is performed resulting in greater IFNγ release, and a positive QFT-GIT. Our data also suggest that a lower cut-off may improve sensitivity of QFT-GIT for the detection of latent TB in HIV-1-infected persons. Intuitively, having a lower cut-off in HIV-1-infected persons for an immune-diagnostic test is logical as the cells which contribute to a positive result are especially susceptible to HIV-1 induced depletion [Taken together, our data supports the possibility that within this population of HIV-1-infected persons with negative TST and QFT-GIT there may be 2 distinct populations. A true negative group that have no evidence of immune sensitization by Mtb even following stimulation with mycobacterial peptides in the form of PPD and a false negative group that show initial evidence of weak antigen specific immune response to Mtb, as determined by IFNγ release in whole blood stimulated by ESAT-6, CFP-10, and TB7.7, that can be boosted by intra-dermal PPD injection. One explanation for this phenomenon is that pre-existing memory responses may be stimulated by PPD (which include antigens ESAT-6 and CFP-10) when TST is performed . This maTB assay. It was beyond the scope of the study to investigate whether participants that converted had any immunological differences in phenotype or function of antigen specific T cells in comparison to non-converters. In particular it would be of interest to investigate if converters had evidence of antigen specific central memory responses not present in non-converters as this would have supported the assertion that they represented distinct subgroups.A limitation of the study was that we were unable to validate a lower cut-off for QFT-GIT in HIV-1-infected persons. It would also be important to ensure that any novel cut-off for HIV-1-infected persons was validated for a range of CD4 counts. Furthermore our study was only conducted using QFT-GIT, it would be of interest to determine if a similar phenomenon occurred using the TSPOT.We have demonstrated that in an HIV-1 infected population with negative QFT-GIT and TST  there are 2 distinct groups. One group shows evidence of a weak Mtb specific immune response that boosts significantly following TST resulting in conversion of the test result and may represent a false negative group that may benefit from IPT. We propose that a lower cut off, between 0.15 and 0.17, may improve sensitivity of QFT-GIT and warrants further validation.


Timely linkage to care by newly-diagnosed HIV+ individuals remains a significant challenge to achieving UNAIDS 90-90-90 goals. Current World Health Organization (WHO) guidelines recommend initiating anti-retroviral treatment (ART) regardless of CD4+ count, with priority given to those with CD4+ <350 cells/μl. We evaluated the impact of not having a day-of-diagnosis CD4+ count blood draw, as recommended by South African guidelines, on time to linkage, using data from a prospective cohort study.Individuals (N = 2773) were interviewed prior to HIV counseling and testing at three public sector primary care clinics in the greater Durban area; 785 were newly-diagnosed and eligible for the cohort study; 459 (58.5%) joined and were followed for eight months with three structured assessments. Linkage to care, defined as returning to clinic for CD4+ count results, and day-of-diagnosis blood draw were self-reported.linkage) associated with not having day-of-diagnosis blood draw was 0.66 . By 4 months, 54.8% of those without day-of-diagnosis blood draw vs. 75.2% with one were linked to care (chi-squared p = 0.004). Of those who deferred blood draw, 48.3% cited clinic-related and 51.7% cited personal reasons. AHRlinkage was 0.60  for clinic-related and 0.53  for personal reasons relative to having day-of-diagnosis blood draw.Overall, 72.5% did not have a day-of-diagnosis CD4+ count blood draw, and 19.2% of these never returned. Compared with a day-of-diagnosis blood draw, the adjusted hazard ratio of linkage (AHRNewly-diagnosed HIV+ individuals who did not undergo CD4+ count blood draw on the day they were diagnosed—regardless of the reason for deferring—had delayed linkage to care relative to those with same-day blood draw. To enhance prompt linkage to care even when test and treat protocols are implemented, all diagnostic testing required before ART initiation should be performed on the same day as HIV testing/diagnosis. This may require modifying clinic procedures to enable overnight blood storage if same-day draws cannot be performed, and providing additional counseling to encourage newly-diagnosed individuals to complete day-of-diagnosis testing. Tracking HIV+ individuals via clinic registries should commence immediately from diagnosis to reduce these early losses to care. In most low and middle income countries (LMIC) current practice is to stage individuals’ HIV disease, either clinically or through CD4+ count, as a first step in care. In September 2015, World Health Organization (WHO) guidelines were revised to recommend that all HIV+ individuals be placed immediately on antiretroviral therapy (ART) regardless of CD4+ cell count, and that those with CD4+ count less than 350 cells/μl be prioritized for ART initiation . EarlierDuring the first decade of ART roll-out in South Africa, treatment thresholds tended to lag behind those recommended by WHO, but in recent years, guidelines generally have been in line with those recommended by WHO ,15. DuriIn a prospective cohort study, ‘Pathways to Engagement in HIV Care for Newly-Diagnosed South Africans’ , we examined what proportion of individuals had blood drawn for CD4+ count determination on the day of diagnosis, reasons for not doing so on that day, and the relationship between not having a day-of-diagnosis blood draw and time to linkage to care. The results of this analysis provide critical insights into how procedures required for access to care and treatment might promote or delay linkage to those services.The Pathways to Care cohort consisted of newly-diagnosed HIV+ women and men recruited from three public sector primary healthcare clinics (PHC) located in greater Durban, including two sites in urban areas and one in a rural area. KwaZulu-Natal, the province in which Durban is situated, has the highest HIV prevalence in South Africa, with 17.4% (15.8–19.2) of the population aged two years and older infected in 2012  The p-value associated with the interaction term was 0.40.In analyses stratified by gender (not shown in table), the AHRIn this prospective study that identified individuals prior to HCT and followed them for eight months from the day of diagnosis with high retention rates, we found that newly-diagnosed HIV+ individuals who did not have their blood drawn for CD4+ count on the same day as they were diagnosed had a significantly lower rate of linkage to care than those who did, and that both clinic-related and personal reasons for not having this test on the day of diagnosis were associated with a significantly lower rate of linkage. The effects were similar for women and men. Moreover, almost one-fifth of those who did not have this test done on their day of diagnosis never returned.There are three important implications of these findings. First, this study highlights the importance of identifying very early losses to care. Regardless of the reason, leaving the clinic without completing all the recommended services for that visit seemed to be a marker for being at risk for delayed or no return. Tracking HIV+ individuals enrolled in care—both pre-ART and after initiating ART—for potential loss to follow-up is now an accepted best practice , and ourSecond, clinic-related barriers were important in explaining about half of the cases with no day-of-diagnosis blood draw. Clinic procedures can make a difference in individuals’ ability to complete necessary services, especially in settings where poverty-related barriers may make it difficult to attend the clinic. Once identified, clinic-related barriers often can be addressed through changes in procedures.In this study 35% of individuals who did not have blood for CD4+ count drawn at diagnosis indicated that the clinic or lab was closing, or that they were told by staff to return another day. When we presented study findings to clinic staff, they explained that CD4+ blood draws were sometimes deferred because the van collecting specimens for transport to the laboratory had already left on that day, and blood was not stored in the clinic overnight. However, new evidence suggests this barrier could be eliminated without great expense; Skhosana et al. demonstrated high reproducibility of CD4+ count results of blood refrigerated overnight . If thosThird, personal reasons for leaving the clinic before having blood drawn for CD4+ count also were important, and these require further investigation, both in research and in practice. Although some explanations lacked detail, they likely reflect psychological processes related to receiving a diagnosis of a life-threatening and highly stigmatized disease, along with structural barriers related to work and family responsibilities. Such barriers may continue to exert an influence even if disease-staging prior to initiation ART is eliminated. Therefore, our findings also highlight how vital it is to ensure that newly-diagnosed HIV+ individual leave the clinic with a basic package of linkage services–immediate psychological support, an assessment of barriers to returning to the clinic, and a plan for follow-up contact and support. These services often are not routine in busy public sector clinics such as those where we conducted this study, but are essential to successful linkage to care.The findings of this study also are notable for their consistency across women and men. Whereas other research has shown that men are at greater risk than women for late diagnosis and loss to care ,20, the Strengths of this study include that it was a prospective cohort study representative of people undergoing HCT at these clinics, with close to a 60% of those eligible joining the study. Minimal differences between eligible participants who joined and did not join—by clinic and relationship status—were unlikely to have altered these findings, as all analyses were stratified by clinic, and relationship status was not associated with the exposure or outcome. Additional strengths included that we identified potential study participants and assessed potentially confounding variables prior to diagnosis, had less than 4% loss to follow-up, and that we obtained detailed information about dates, locations, and reasons for all clinic visits during the follow-up period. A limitation was that we relied on participant self-report of clinic visits and tests, including both the exposure and outcome. In these public primary care clinics individual medical records were not created until an HIV+ person initiated ART. Although dates of blood draw could have been obtained from the National HIV laboratory system, we did not use this source because it lacked data on our primary outcome–return to the clinic to obtain CD4+ count test results. Due to the careful way that we elicited visit data, we believe they were comparable in completeness and accuracy to what we would have obtained through clinical records had they existed. However, having multiple sources from which to obtain and validate this information would have increased accuracy.Other limitations are that we did not probe individuals’ explanations for why they did not have a day-of-diagnosis blood drawn for CD4+ count and we did not ask this question of people who had had this test done by the time of their baseline interview. Additionally, although we did not provide clinic visit reminders, payments to individuals when they returned for study interviews (which were conducted close to clinics of recruitment) may have encouraged them to keep clinic visits. Our estimates of time to linkage, therefore, may be higher than what would have occurred without the study.In summary, in a detailed analysis made possible by interviewing individuals beginning when they first presented for HCT, we have shown that adherence to the South African health service guideline for immediate blood draw for CD4+ count on the day of diagnosis was less than optimal in these clinics, and was associated with substantial loss to follow-up and delayed linkage to care. Engaging clinic staff in identifying and removing clinic-related barriers to these first steps in care can be an important tool for achieving the 90-90-90 targets set forth by UNAIDS . FurtherS1 Data(XLSX)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.


Patients with substance use disorders grow older thanks to effective treatments. Together with a high prevalence of comorbidities, psychological problems, and low social support, these patients are at high risk for medication non-adherence. Established treatment facilities face challenges to accommodate these complex patients within their setting. Electronic medication management aids (e-MMAs) might be appropriate to simultaneously monitor and improve adherence for these patients.We report the first long-term experiences with a novel remote electronic medication supply model for two opioid-dependent patients with HIV. John  and Mary  suffered from disease progression due to non-adherence. We electronically monitored adherence and clinical outcomes during 659 (John) and 953 (Mary) days between July 2013 and April 2016. Both patients retrieved over 90% of the pouches within 75 min of the scheduled time. Technical problems occurred in 4% (John) and 7.2% (Mary) of retrievals, but on-site support was seldom required. Viral loads fell below detection limits during the entire observation period.Continuous medication supply and persistence with treatment of over 1.7 years, timing adherence of more than 90%, and suppressed HIV viral load are first results supporting the feasibility of the novel supply model for patients on opioid-assisted treatment and polypharmacy. Along with the aging of the general population, the number of older users of illicit substances (or older drug users) is growing worldwide , 2. Age-. In Switzerland, OAT is offered by a wide range of providers, such as general practitioners, specialized clinics, and addiction centers [Opioid-assisted treatment (OAT) is recommended for opioid dependence . It is e centers . In the  centers . The out centers .Because existing nursing homes or home-care services are often not suited or willing to accommodate patients with SUDs, outpatient treatment and surveillance are provided as long as possible. Thanks to regular appointments, caregivers may ensure initiation and persistence with treatment, but may not be able to assure correct implementation of the dosing regimen. Many patients take their medications irregularly due to a lack of structure in their daily routine. As a result, treatment success may be compromised, resulting in health risks not only for patients but also for society. The cost of providing care to the aging older drug users is expected to increase, and innovative solutions to optimize medication management compatible with OAT are therefore required. In this context, we developed a novel medication supply model with interdisciplinary collaboration between the OAS, an HIV clinic, and the Emergency Pharmacy of Basel, in order to guarantee adherence by using an electronic medication management aid (e-MMA) for pre-packed polypharmacy located at patients’ homes. Various e-MMAs for polypharmacy exist, but only a few studies on implementing the devices in routine care have been published. Most studies are conducted outside of drug-using populations, do not describe the supply model in detail, and focus on the use and the benefits of e-MMAs for patients, such as improved safety of buprenorphine storage  or effecWe present two cases of outpatients living in social housing in Basel, Switzerland, who obtained medication including opioids from the OAS Basel. After some years, conventional care and adherence to medication were questioned, especially after missed appointments and flares of HIV viral load. Both patients accepted the novel supply model with electronic monitoring of the entire medication. Both patients consented to the publication of their cases.John was 50 years old when he entered the study on July 2, 2013. He had been diagnosed with HIV at the age of 25 as a result of intravenous drug use. A liver biopsy in 2014 showed cirrhosis and severe activity due to chronic HCV infection and alcohol abuse . He lost his girlfriend to suicide and lived with a friend who also suffered from SUD. He was unemployed and spent most of his days at home. After diagnosis of a long-QT syndrome in early 2013, he was switched from 150 mg methadone to 1200 mg (6 tablets) of long-acting morphine daily. Remaining treatment consisted of 5 tablets once daily: ritonavir 100 mg, darunavir 2 × 400 mg, tenofovir/emtricitabine 245 mg/200 mg, and pantoprazole 40 mg. Viral load reached 1000 copies per milliliter (copies/ml) during 2012 and early 2013. His viral load fell below detection limits after he was obliged to visit the OAS daily (instead of once weekly) to assure regular intake of his medication. However, he continued to miss appointments, and the situation remained unsatisfactory for him and his caregivers. Evaluation in the local memory clinic revealed a diagnosis of moderate Alzheimer’s disease accompanied a moderate depressive episode.9 cells) necessitated the introduction of a prophylaxis against Pneumocystis carinii in summer 2013. Her caregivers convinced her to resume therapy with the same treatment as before and her viral load started to decrease. She understood the need for treatment but lacked the motivation to adhere despite intensive psychological support. At the time, additionally to methadone and methylphenidate, she was taking: lopinavir/ritonavir 200 mg/50 mg 4 tablets once daily, darunavir/emtricitabine 245 mg/200 mg 1 tablet once daily, and sulfamethoxazole/trimethoprim 800 mg/160 mg 1 tablet every Monday, Wednesday, and Friday. Her hypertension was not an issue at that time, and an approach of watchful waiting was considered appropriate with the intention not to jeopardize adherence to HIV medication.Mary was 46 years old when she entered the study on 18 August 2013. She had been diagnosed with HIV at the age of 19. She suffered from hypertensive cardiopathy, chronic lymphedema in both legs, and suspected chronic obstructive pulmonary disease (COPD). She had a history of hepatitis C and had been a heavy smoker for years. Her OAT consisted of liquid oral methadone (170 mg), and she received sustained-release methylphenidate (90 mg) for attention-deficit/hyperactivity disorder (ADHD). She lived with a friend. Both were not working and rarely left home. In 2012, she started to take her HIV medication only sporadically and stopped altogether in early 2013. As a result, her viral load increased sharply to over 250,000 copies/ml. A low CD4 count  and in the living room (Mary). Dispense times were scheduled in line with consistent habits of daily life, i.e., 12 pm for John  and 8 pm for Mary (watching TV). John was followed for 659 days. At the time of drafting of this article, Mary is still using the e-MMA. We present data from 953 days. Adherence was electronically monitored during 655 days  and 911 days  more than 75 min after the scheduled time and forgot to retrieve 4 doses , appointments were kept and the laboratory results were satisfactory.In December 2015, Mary mentioned that when she was busy in the flat and the alarm would ring, she would sometimes press the OK button to retrieve the pouches but would walk away and go back to her business. Sometimes, she would then forget to take the medications later and save the pouches in a drawer. Between May and December 2015, she reportedly skipped medication intake on 21 days (10% over 8 months). Because viral load was constantly suppressed and CD4 counts recovered, prophylaxis of P. carinii. Adapting the content of the pouches is possible without modifying the intake habits. This insures that changes in treatment do not coincide with variable intake times and thus might prevent non-adherence.A novel medication supply model with electronic adherence monitoring of polypharmacy showed sustained treatment implementation and suppressed viral loads in two opioid-dependent HIV patients over 1.7 and 2.5 years, respectively. The drop of viral loads started when patients resumed therapy after intervention of their caregivers. It has been demonstrated that adherence interventions for long-term treatments need regular follow-ups to remain effective . HoweverElectronic monitoring makes changes in dosing patterns instantly apparent and allows for a timely intervention. Feedback from electronic monitoring has been shown to effectively improve adherence . The onlOur study has several strengths. First, we included patients from a population with a high probability of non-adherence and a high prevalence of time-sensitive medication regimens, such as highly active anti-retroviral therapy (HAART) for HIV. Thus, the success of our intervention in these complex patients demonstrates the potential of our supply model. Second, we measured adherence to polypharmacy. Typically, devices for electronic monitoring are designed for single preparations. The monitoring of polypharmacy thus requires multiple devices and may complicate the management of medications. With our e-MMA, all medications were dispensed in unit-of-use pouches, which enhanced the likelihood of concurrent intake.We acknowledge some limitations. First, measuring adherence with the e-MMA might overestimate adherence because medication retrieval does not equal ingestion. Literature suggests that electronic monitoring might underestimate adherence , althougSecond, although John and Mary experienced a benefit from the dispenser, case reports cannot generate results to claim effectiveness of an intervention. Additionally, the generalizability of our results is limited. We evaluated the e-MMA in two patients that matched the envisaged target groups suggested in a qualitative study of the dispenser . Living Finally, we did not evaluate the financial implications of the novel supply model. Costs for repackaging of medications are reimbursed in Switzerland for patients with polypharmacy . Costs of the e-MMA and additional costs for service and support are currently not reimbursed by health insurances. However, savings from improved adherence might offset the additional costs, as shown in previous studies where better adherence resulted in significant cost savings , 33.Continuous medication supply and persistence with treatment over more than 1.7 years, timing adherence of more than 90%, and suppressed HIV viral load are the first results supporting the feasibility of the novel supply model. Further trials should aim at evaluating the effectiveness of the supply model in terms of clinical, humanistic, and economic outcomes and in patients that do not necessarily match the envisaged target groups.


HIV care programs in resource-limited settings have hitherto concentrated on antiretroviral therapy (ART) access, but HIV drug resistance is emerging. In a cross-sectional study of HIV-positive adults on ART for ≥6 months enrolled into a prospective cohort in Uganda, plasma HIV RNA was measured and genotyped if ≥1000 copies/ml. Identified Drug resistance mutations (DRMs) were interpreted using the Stanford database, 2009 WHO list of DRMs and the IAS 2014 update on DRMs, and examined and tabulated by ART drug classes.Between July 2013 and August 2014, 953 individuals were enrolled, 119 (12.5%) had HIV-RNA ≥1000 copies/ml and 110 were successfully genotyped; 74 (67.3%) were on first-line and 36 (32.7%) on second-line ART regimens. The predominant HIV-1 subtypes were D (34.5%), A (33.6%) and Recombinant forms (21.8%). The commonest clinically significant major resistance mutations associated with the highest levels of reduced susceptibility or virological response to the relevant Nucleoside Reverse Transcriptase Inhibitor (NRTI) were; the Non-thymidine analogue mutations (Non-TAMS) M184V—20.7% and K65R—8.0%; and the TAMs M41L and K70R (both 8.0%). The major Non-NRTI (NNRTI) mutations were K103N—19.0%, G190A—7.0% and Y181C—6.0%. A relatively nonpolymorphic accessory mutation A98G—12.0% was also common. Seven of the 36 patients on second line ART had major Protease Inhibitor (PI) associated DRMS including; V82A—7.0%, I54V, M46I and L33I . Also common were the accessory PI mutations L10I—27%, L10V—12.0% and L10F—5.0% that either reduce PI susceptibility or increase the replication of viruses containing PI-resistance mutations. Of the 7 patients with major PI DRMs, five had high level resistance to ritonavir boosted Lopinavir and Atazanavir, with Darunavir as the only susceptible PI tested.In resource-limited settings, HIV care programs that have previously concentrated on ART access, should now consider availing access to routine HIV viral load monitoring, targeted HIV drug resistance testing and availability of third-line ART regimens. Antiretroviral therapy (ART) improves survival and quality of life of HIV-infected individuals and controls HIV transmission, however HIV-1 drug resistance limits the benefits of ART . Samples with HIV RNA viral load of 1000 copies/mL or higher were genotyped. For ARV drug resistance testing, HIV RNA was extracted using the QIAmp Viral RNA mini kit (Qiagen), the entire protease and amino terminus of the reverse transcriptase was amplified, cleaned and sequenced using the ABI 3500 machine (Applied Biosystems). Sequences were base-called using Sequencher v5.2.4 and sequence alignments performed using BioEdit v7.2.5  and SeaVParticipants with HIV-1 RNA viral load of 1000 copies/mL or higher, whose samples were successfully amplified and genotyped were included in this analysis. The socio-demographic characteristics including age and sex were examined and tabulated. ART data consisted of the type of ART regimen and duration on ART. The three ART regimen types were; Triple nucleoside (3 NRTIs), 2 NRTIs with a NNRTI, and a PI-based regimen. Duration on ART was grouped into below 5 years, 5–9 years, and above 9 years. Drug resistance mutations to one antiretroviral class only , two classes only  or three antiretroviral classes (NRTI and 1 NNRTI and PI) were tabulated. For participants with a major PI DRM, we showed the resistance levels to each antiretroviral drug.Out of the 953 individuals on ART for 6 or more months, 119 12.5%) had a viral load ≥1000 copies/ml. The five samples that did not amplify and the 4 with insufficient quantities were excluded from this analysis. Of the 110 individuals whose samples were sequenced for drug resistance testing, 73 (66.4%) were females, 95 (86.4%) were aged 35 years and above, 89 (80.9%) had been on ART for 9 or more years, with 74 (67.3%) on first and 36 (32.7%) on second-line ART regimens at enrolment. Sixty-five (59.1%) had CD4 cell counts of 350 cells/ml or less and 67 (60.9%) had HIV RNA of 10,000 copies/ml or more. The predominant HIV-1 subtypes were D (34.5%), A (33.6%) and Recombinant forms (21.8%) : M184V—20.7% and K65R—8.0%, while M41L and K70R (both 8.0%) were the commonest TAMs. The commonest major NNRTI resistance mutations known to reduce susceptibility or virological response to NNRTIs were: K103N—19.0%, G190A—7.0% and Y181C—6.0%. A relatively nonpolymorphic accessory mutation A98G—12.0% was also common. Of the 33 identified PI resistance mutations, 7 were major mutations and 26 minor mutations. The most common major PI-resistance mutations associated with the highest levels of phenotypic resistance were: V82A—7.0% and I54V, M46I, L33I . The identified accessory PI resistance mutations that either reduce PI susceptibility or increase the replication of viruses containing PI-resistance mutations included: L10I—27.0%, L10V—12.0% and L10F—5.0% , Fosamprenavir (FPV), Lopinavir (LPV) and Nelfinavir (NFV)—to each was found among 4 patients. Intermediate level drug resistance to Saquinavir (SQV) was found among 4 patients, Tipranavir (TPV) and IDV among 3 patients. The HIV among six of the 7 patients with major PI mutations were susceptible to Darunavir with one expressing low level resistance to this drug, and two expressing susceptibility to Tipranavir . NRTIs have fewer side effects and toxicity than NNRTIs, but drug resistance could diminish their efficacy. As expected in Africa, the predominant NRTI mutation observed was M184V . Type 1 The observed PI resistance mutations  are clinically significant because they are associated with highest levels of phenotypic resistance and/or strongest evidence for interfering with successful PI therapy. Among the seven patients, these mutations conferred high level resistance to ritonavir boosted Lopinavir and Atazanavir, which are the readily available PIs for second-line ART regimens in our setting. Darunavir, the only susceptible PI tested is still expensive and unavailable in public ART centers in Uganda where PIs are reserved for second-line ART regimens. The emergence of HIV drug resistance is inevitable, owing to the high replication and mutation rates of HIV and ART being a life-long treatment. Therefore, as more people are switched to second-line ART, cases of second-line drug failure will increase and necessitate access to third-line or salvage regimens. In Uganda, routine viral load monitoring is not yet standard care, making early identification of treatment failures to prevent transmission of DRMs impossible. Uganda is yet to recommend HIV genotyping prior to ART initiation, therefore diagnosing transmitted HIV drug resistance is impossible. The strength of our study is the long duration on ART among our participants. A source of selection bias might be that some patients with virological failure might have died before enrolment. We also had limited information on confounders like adherence so our findings may not be generalizable. In conclusion, HIV drug resistance is a major challenge for HIV care programs in resource limited settings that have hitherto concentrated on increasing ART access. Therefore, ART programs in these settings should avail routine HIV viral load monitoring for prompt detection of virological failures, targeted HIV drug resistance testing to detect HIV drug resistance among ART failure as well as access to third-line ARV drugs like Darunavir.


Human immunodefiecency virus infected patients did not adhere correctly to their Antiretroviral Therapy because of the drugs adverse effects. Thus, continuous evaluation of the adverse effect of Antiretroviral Therapy will help to make more effective treatment. The aim of this study was to assess the prevalence of Adverse Drug Reaction and associated factors on Antiretroviral Therapy among Human immunodefiecency virus infected Adults at Hiwot Fana Specialized University Hospital, Eastern Ethiopia.A Hospital based retrospective study was conducted among 358 of adult patients clinical records on antiretroviral Therapy from April1 to June30, 2014.The overall prevalence of Adverse Drug Reaction among Human immunodefiecency virus infected patients on antiretroviral Therapy was 17.0%. Of reported Adverse Drug Reaction, 80.3%, 18% and 1.7% occurred in patients on Stavudine, Zidovudine and Tenofovir based regimens respectively. The common Adverse Drug Reaction were lipodystrophy (fat change) (49.2%), numbness/tingling (27.9%), peripheral neuropathy (18%) and (8.2%) anaemia (8.2%). Patients on Stavudine containing regimens were more likely to develop Adverse Drug Reaction compared to Zidovudine  and Tenofovir .The overall prevalence of Adverse Drug Reaction among Human immunodefiecency virus infected patients in this study was 17% and more common on those patients taking Stavudine based regimen. Lipodystrophy and peripheral neuropathy were significantly associated with stavudine-based regimens, while anaemia was significantly associated with zidovudine based regimens. Thus regular clinical and laboratory monitoring of patients on Antiretroviral Therapy should be strengthened. Antiretroviral Therapy (ART) is the cornerstone of management of patients infected with Human immune deficiency virus (HIV). The initiation ART has resulted in significant decreases in HIV-related morbidity and mortality in both the developed and developing countries –3. It isStudy area and population: Harari People National Regional State is located in the Eastern part of Ethiopia which is 515 kms away from the capital city- Addis Ababa. It had a projected total population of 203,438 in 2010. It has 36 kebeles , which are the lowest administrative clusters in the region. The health service coverage of the region, which is calculated based on the number of health institutions giving health services per population in the region, is estimated to be about 100%. There were 6 Hospitals and 8 health centers in the region [e region . ART proStudy design and period: A Hospital based retrospective review of ART clinical records were conducted among patients who initiated ART between March26, 2006 and March 26, 2014.Sample size estimation and sampling techniques: The sample size was determined by using single proportion formula by setting 95% confidence interval, P= 0.303 of patients taking ART develop ADRs [lop ADRs , margin Data collection: Data were collected using structured questionnaire which was adopted from routine standard ART monitoring card. Information on patient's details like socio demographic characteristics, WHO clinical staging of the disease at the starting of ART, duration of treatment, drug details, types, severity and outcome of ADRs of reported by patients during clinical examination at time coming to ART unit for their appointment visit were collected by clinical nurses who are working at the hospital ART Clinic. Haematological and metabolic ADRs, which could be done by laboratory examination, were not collected from ART clinical records. The data collection format was checked on daily bases by investigators and supervisors for its completeness and consistency with the patient's clinical records of the hospitals.Data entry and analysis: The collected data were double entered, cleaned and analyzed by using SPSS soft-ware Version16. The finding of this study was presented by using mean, standard deviation and simple frequencies tables. The overall prevalence of ADRs was determined as the proportion of individuals who developed the different ADRs by clinical examination. Odd ratio with 95% confidence interval was used to describe association between the selected study variables (i.e. Outcome and independent variables). Univariate and multivariable logistic regression analysis was performed to explore independent variables that were predictors of adverse drug reactions (ADRs). The criterion for significance was set at P < 0.005. The severity of ADRs were categorized from grade I to IV based on criteria used the WHO severity grading [Ethical consideration: The study protocol was approved by Institutional Health Research and Ethical Review Committee of College of Health and Medical Sciences, Haramaya University. And the patients’ clinical records were reviewed anonymously and all information obtained from clinical records was kept confidential.Characteristics of the study subjects: A total of 358 HIV/AIDS patient ART clinical records were reviewed in this study. The mean age of patients was 34 (SD + 9.8) years, ranging from 18-71 years. Most of the patients were on the age group of 30-39 years (41.4%), female (68.4%), Urban (83%) and elementary school (44.7%). One hundred fifty (41.9%) of the study participants were at WHO clinical stage III. With regards to functional status, most (65.9%) of them were working. Majority (74.6%) of the study participants were on the first line ART regimen. D4T (30)/3TC/ NVP (33.2%) were the most commonly prescribed ART regimen at their initial time initiation of ART. The most frequently used drug other than ART among the study participants were Cotrimoxazole (75.7%) and Isonizide (INH) (14.5%) ( (14.5%) .Antiretroviral treatment (ART) Drug regimen and follow up status of HIV/AIDS patients: At the time of study, 33.2% of the study participants were still on their initial regimen whereas 38% of them had changed one or two drugs during their follow up. Currently (during the study period), most (69.7%) of the HIV/AIDS patients on ART were in the first line regimens. The main reasons for changing initial regimen of ART drugs were new drug availability (45.6%) and adverse drug reaction (41.2%) followed by Immunological failure 10(7.4%), due to new TB 9(6.6%), risk of pregnancy, clinical failure, virological failure, immunological failure and clinical failure, and immunological failure and viral failure each accounts 1(1%). Two hundred fifty eight (72.1%) were still on ART follow up in the hospital ART clinic whereas 48(13.4%), 29(8.15) and 24(6.7%) of the study participants were transferred out to other ART clinic site, died and interrupted their follow up respectively. ART treatment adherence rate among study participants was also reviewed from participant's card. About (5%) had poor adherence. The main reasons for poor adherence were lost / ran out of pills 10(55.6%) and delivery/ travel problems 4(22.2%), followed by nutritional problem 1(5.6%), taking alcohol1 (5.6%), too ill 1(5.6%) and combination of stigma 1(5.6%).Magnitude, clinical symptoms and severity of adverse drug reactions (ADRs): The overall prevalence of ADRs among these study participants was 17.0%. The most frequently identified ADRs clinical symptoms were lipidtropy (Fat change) (44.3%) and Numbness/tingling (4.9%). Most (80.3%) of the patients had grade III ADR. The main management/measure taken to resolve ADRs were one drug change (62.3%), reassurance (18%) and regimen change (14.8%). The majority 27/61 (44.3%) of the study participants with ADRs were died who were on grade III. In addition, most of the study participants, who developed ADRs, were WHO stage III (47.5%) ( (47.5%) . Nausea, (47.5%) .Factors associated with adverse drug reactions: The prevalence of ADRs was higher in those study participants on D4T based ART drug regimen especially D4T/3TC/EFV. This was significantly different when compared to study participants in other treatment regimens (P<0.005). The prevalence of ADRs was higher in those study participants who were more than 50 years of age, in females, ambulatories, in WHO stage III, with initial CD4 count 201-300 cells/µl and grade III severity level. But the difference was not statistically significant when comparisons were made within each category member (P>0.05). Further analysis with univariate and multivariate was done. In both univariate and multivariate analysis all ART drug users were less likely to develop ADRs as compared to D4T/3TC/ NVP users (p<0.05). It was compared based on the base ART drugs; AZT and TDF based ART drug regimen base users were less likely to develop ADRs when compared to D4T base users  and Cameroon 19.5%)  but it w.5%  but The other common ADR was peripheral neuropathy 18%). This was similar to the study carried out in Cameroon 21.2%) and Kenya (20.7%) %. This w. However.2% and KThe prevalence of ADRs was 17% and more common on those patients taking Stavudine based regimen. Lipodystrophy, peripheral neuropathy and anaemia were the common ADRs. Lipodystrophy and peripheral neuropathy were significantly associated with stavudine-based regimens, while anaemia was significantly associated with zidovudine based regimens. Therefore this study recommends, regular clinical and laboratory monitoring of patients on ART should be strengthened before and after initiation of ART according to guideline of institution particularly for stavudine and zidovudine based regimens. It also recommends further large scale study to assess the magnitude and associated factors ADRs by using clinical and laboratory examination.The prevalence of ADRs was more common on those patients taking Zidovudine -based regimens;Lipodystrophy and peripheral neuropathy were significantly associated with stavudine-based a regimen;Anemia was significantly associated with zidovudine based regimens.The prevalence of ADRs was more common on those patients taking Stavudine based regimen;Lipodystrophy was the common ADRs.


Talaromyces marneffei infection is a major cause of death in HIV-infected individuals in South and Southeast Asia. Talaromycosis immune reconstitution inflammatory syndrome has not been well described. Here we report the clinical features, management, and outcomes of three HIV-infected patients with talaromycosis-associated paradoxical immune reconstitution inflammatory syndrome in Ho Chi Minh City, Vietnam. The development of skin nodules on her face and body prompted her to seek care at our hospital, where she was diagnosed with HIV infection by three different enzyme-linked immunosorbent assay (ELISA), and with talaromycosis by skin and blood culture. She had no history of injection drug use (IDU). The CD4+ T-cell count was 10 cells/µL. Her laboratory tests revealed pancytopenia and elevated aspartate aminotransferase (AST) (T. marneffei and other organisms. Multiple acid fast bacilli (AFB) smears of sputum and lymph node were negative, and chest x-ray showed no abnormality. The diagnosis of talaromycosis IRIS was made based on the temporal association between the timing of ART, the appearance of the inflamed skin lesions and joints, and the significant rise in CD4+ count. The dose of itraconazole was increased from 200 mg/day to 400 mg/day for 12 further weeks, and ART was continued. She was discharged after 2 weeks and continued to improve on ART. She received a total 23 months of itraconazole therapy. At her last follow up visit in August 2015, she was well, and her CD4+ T-cell count was 192 cells/µL.se (AST) . She wasse (AST) , enlargePatient 2: A 35 year-old male with a history of IDU from Ho Chi Minh City presented in May 2013 with a two week history of fatigue and scaly skin lesions on his face and body without fever. He had been diagnosed with HIV and T. marneffei infection by skin microscopy and culture at another hospital and had started itraconazole therapy (currently on a 200 mg/day maintenance dose). He was started on first-line ART 3 months prior to presentation, at which time his CD4+ T-cell count was 9 cells/µL, and reported good adherence to ART and itraconazole maintenance therapy. The skin lesions on this presentation were significantly larger and different from before and were scaly psoriasis-like papules on his face and trunk . Hemoculture was negative. She was treated with intravenous amphotericin B deoxycholate and intravenous oxacillin for MSSA super-infection of the skin lesions, both for 2 weeks plus one week of ibuprofen for joint pain. Her fever and skin lesions resolved after 10 days; however she developed productive cough and cervical lymph node enlargement during hospitalization. FNA of the lymph node showed no organism; however her sputum AFB smear was positive and she was started on antituberculosis therapy with isoniazid, rifampicin, ethambutol, and pyrazinamide. She was discharged after 19 days of hospitalization on antituberculosis therapy, and the dose of itraconazole was increased to the maximum dose of 600 mg/day for further 10 weeks to account for the rifampicin-induced reduction in itraconazole levels, as therapeutic monitoring for itraconazole was not available in Vietnam. The patient had a gradual improvement of symptoms over 3 months. At the last follow up visit in June 2016, she was completely well, and the CD4 T-cell count had increased to 170 cells/µL.and pain . She hadand pain . Her hem310 copies/ml or an increase in CD4+ T-cell count from baseline. In resource poor areas receipt of and adherence to an effective ART regimen can be sufficient. The clinical features should be consistent with an inflammatory process, and the clinical process should not be explained by a new infectious process, by drug toxicity or drug resistance The plethora or presentations of IRIS are in part driven by the underlying pathogen. Although there is no gold standard or a single test that can confirm the diagnosis, a number of pathogen specific definitions of IRIS have been proposed All three of our patients required hospitalization for exclusion of other active infections, re-treatment with high dose antifungals, and supportive care. All patients recovered with continuation of ART. Although talaromycosis IRIS can be clinically challenging and can have significant morbidity, the outcomes of our patients and of those reported in the literature with unmasking talaromycosis IRIS appear to be better than seen in IRIS associated with cryptococcal meningitis, where mortality is as high as 34% Co-infection with talaromycosis and tuberculosis is common, with microbiology-confirmed tuberculosis coinfection seen in 22% of patients This case study highlights the clinical manifestations and management in patients with paradoxical talaromycosis IRIS and call for research to define the incidence, risk factors, and clinical impact of this understudied complication in patients with advanced HIV infection in Asia.We declare that there are no competing interests.


In contract, patients with a combination of end-of-treatment HBsAg ≥ 50 IU/mL and HBV DNA ≥ 2 × 105 IU/mL at entry experienced high virological and clinical relapse . No patients experienced hepatic decompensation when clinical relapse occurred after timely retreatment. A combination of HBV DNA levels at entry and end-of-treatment HBsAg levels was useful for predicting the post-treatment HBV relapse in HBeAg-negative patients with HBsAg ≤ 200 IU/mL.The predictors of hepatitis B virus (HBV) relapse and HBsAg loss after cessation of nucleos(t)ide analogues (NA) in HBeAg-negative patients with end-of-treatment HBsAg ≤ 200 IU/mL remains unclear. The study recruited 119 chronic hepatitis B (CHB) patients who achieved end-of-treatment HBsAg ≤ 200 IU/mL, were treated with lamivudine (n = 34) and entecavir (n = 85). The 5-year rates of post-treatment virological relapse, clinical relapse, and HBsAg loss at 60 months were 39.4%, 27.6%, and 45.9%, respectively. Cox regression analysis revealed that HBV DNA at entry and end-of-treatment HBsAg levels were independent predictors of virolgical and clinical relapse. HBV genotype C and end-of-treatment HBsAg were independent factors of HBsAg loss. Patients with a combination of end-of-treatment HBsAg < 50 IU/mL and HBV DNA < 2 × 10 Clinical manifestations of CHB can range from asymptomatic to severe chronic liver disease, such as cirrhosis and hepatocellular carcinoma (HCC)6. Nucleos(t)ide analogues (NAs) are very effective in treating CHB and viral suppression. However, virological relapse is common after the cessation of NAs, and long-term treatment is required10. It remains controversial whether or when NAs can be discontinued.Chronic hepatitis B (CHB) is a major global health problem12. However, some studies found that serum HBsAg levels were weakly or moderately correlated with serum HBV DNA, and were weakly or no correlated with intrahepatic cccDNA14. Low HBsAg can also predict subsequent HBsAg loss and stratify the risk of HCC16. Previous studies demonstrated that serum HBsAg < 100 IU at the end of treatment was a predictor of sustained response after discontinuing NAs18. Our previous studies also showed that end-of-treatment HBsAg levels <200 or <150 IU/mL were useful predictors for HBV relapse in HBeAg-negative patients after stopping lamivudine and entecavir treatment, respectively20. However, patients with such low HBsAg levels still have a chance of HBV relapse.Previous studies showed that hepatitis B surface antigen (HBsAg) quantification has been shown to correlate with HBV DNA and intrahepatic covalently closed circular DNA (cccDNA) levels20. Thus, further studies are needed to find predictors associated with a high rate of HBV relapse after discontinuing NA therapy in HBeAg-negative patients with low HBsAg levels. The aim of this study is therefore to investigate the rates and predictors of off-therapy HBV relapse and HBsAg loss in HBeAg-negative patients with end-of-treatment HBsAg level ≤200 U/L.Our recent study showed a virological relapse rate of about 25% at 36 months after cessation of entecavir therapy in HBeAg-negative patients who achieved end-of-treatment HBsAg < 150 IU/mLThe 119 patient included 95 men and 25 women with a mean age of 52.0 ± 11.7 years at entry. Table Among the 119 patients, 45 experienced virological relapse during the follow-up period. The cumulative rates of virological relapse at 12, 36, and 60 months were 25.2%, 38%, and 39.4%, respectively. Table P < 0.001), respectively.The end-of-treatment HBsAg level was an independent factor for virological relapse. An HBsAg level of 42 IU/mL was the best cut-off value of for predicting virological relapse within 5 years (AUROC: 0.707). Thus, we used HBsAg level of 50 IU/mL as the optimal value. In patients who had HBV DNA < 50 and ≥50 IU/mL, the virological relapse rates at 60 months were 12.1% and 55.8% . Thus, we used this level as the optimal value. In patients who had HBV DNA< 2 × 105 and ≥2 × 105 IU/mL, the virological relapse rates at 60 months were 22.2% and 61.3% (P < 0.001), respectively.HBV DNA levels at entry were also independent predictors of virological relapse. An HBV DNA level of 2 × 105 IU/mL at entry and end-of-treatment HBsAg levels of 50 IU/mL to assess the combined risk for developing virological relapse. We divided the patients into four subgroups: Group I: HBV DNA < 2 × 105 IU/mL and HBsAg < 50 IU/mL; Group II: HBV DNA ≥ 2 × 105 IU/mL and HBsAg < 50 IU/mL; Group III: HBV DNA < 2 × 105 IU/mL and HBsAg ≥ 50 IU/mL; and Group IV: HBV DNA ≥ 2 × 105 IU/mL and HBsAg ≥ 50 IU/mL. The virological relapse rates at 60 months in Group I to IV were 5%, 35.1%, 23.1%, and 82.5% (P < 0.001), respectively   and entecavir groups (P < 0.001).We further combined HBV DNA of 2 × 10ely Fig. . After m01) Fig. . These tP < 0.001) and end-of-treatment HBsAg levels  are independent predictors for developing clinical relapse. Of the 45 patients with virological relapse, 14 experienced virological and clinical relapse at the same time, 16 did not experience clinical relapse after virological relapse at the last follow-up, and 15 experienced clinical relapse after virological relapse .Among the 119 patients, 29 experienced clinical relapse during the follow-up period. The cumulative rates of clinical relapse at 12, 36, and 60 months were 12.7%, 23.2%, and 27.6%, respectively. The risk factors shown in Table P < 0.001), respectively.The end-of-treatment HBsAg and HBV DNA levels at entry were independent predictors of clinical relapse. An HBsAg level of 42 IU/mL was the best cut-off value of for predicting clinical relapse within 5 years (AUROC: 0.705). Thus, we used HBsAg level of 50 IU/mL as the optimal value. In patients who had HBV DNA < 50 and ≥50 log IU/mL, the virological relapse rates at 60 months were 5.4% and 41% (5 IU/mL (AUROC: 0.864), which we therefore used as the optimal value. The clinical relapse rates at 60 months in patients who had HBV DNA < 2 × 105 and ≥2 × 105 IU/mL were 11.4% and 51.2% (P < 0.001), respectively.The best cut-off value of HBV DNA for predicting clinical relapse within 5 years was 2 × 105 IU/mL at entry and end-of-treatment HBsAg levels of 50 IU/mL into four subgroups as mentioned (Group I to IV). The clinical relapse rates at 60 months in Group I to IV were 0%, 18.9%, 13.4%, and 72.3% (P < 0.001), respectively   and entecavir groups (P < 0.001).We again combined HBV DNA of 2 × 10ely Fig. . After m01) Fig. . These tP < 0.001), clinical relapse , and HBsAg loss . Compared to increased HBsAg levels, any decline from end-of-treatment HBsAg levels at 6 months post-treatment correlated with significantly lower rates of virological relapse , clinical relapse , and HBsAg loss . In group A patients, the decline levels of HBsAg from end of treatment to 6 months post-treatment was not associated with HBV relapse due to very low rates of virological and clinical relapse. In Group B and C patients, patients who had any decline of HBsAg levels from end of treatment to 6 months post-treatment had a significantly lower rates of virological relapse  and clinical relapse  than those with increased HBsAg levels.The HBsAg levels at 6 months after cessation of entecavir and lamivudine treatment were a significant factor for predicting virological relapse . The cumulative rates of HBsAg loss in patients with HBsAg ≤ 50 IU/mL, 51–100 IU/mL, 101–150 IU/mL, and 151–200 IU/mL at 60 months were 54.7%, 66.5%, 33.2%, and 20.3%, respectively . Table 21. Thus, the rate of HBV relapse was not as high in our patients with lower end-of-treatment HBsAg levels.We found that the rates of virological and clinical relapse at 60 months were 39.4% and 27.6%, respectively, after cessation of lamivudine or entecavir treatment in HBeAg-negative patients who achieved end-of-treatment HBsAg ≤ 200 IU/mL. The Cox regression analysis revealed that higher baseline HBV DNA and end-of-treatment HBsAg levels were independent predictors for virological and clinical relapse. A recent systemic review study showed that the pooled rates of virological relapse at 36 months were 69.9% after NA discontinuation in initially HBeAg-negative patients20. A recent study also recognized HBsAg levels <100 IU/ml as a new therapeutic end point22. However, we found that the virological relapse rates in patients with end-of-treatment HBsAg ≥ 50 IU/mL were still high (55.8% at 60 months). In contrast, the virological relapse rate in patients who achieved end-of-treatment HBsAg < 50 IU/mL was relatively low (12.1% at 60 months).Our recent studies showed that in HBeAg-negative patients, HBsAg values of 200 and 150 IU/mL could predict sustained responses after lamivudine and entecavir treatment, respectivelyFor the analysis of clinical relapse, we also found that the virological relapse rates in patients with end-of-treatment HBsAg ≥ 50 IU/mL were still high (40.1% at 60 months). In contrast, the clinical relapse rate in patients with end-of-treatment HBsAg < 50 IU/mL was very low (5.4% at 60 months). Thus, the end-of-treatment HBsAg < 50 IU/mL was more useful for predicting HBV relapse after cessation of entecavir and lamivudine treatment in HBeAg-negative patients.5 IU/mL at entry was a significant independent factor for sustained response after cessation of entecavir therapy in HBeAg-negative patients9, which our results support. Furthermore, we combined HBV DNA of 2 × 105 IU/mL at entry and end-of-treatment HBsAg to assess the combined risk of virological and clinical relapse and found that the virological and clinical relapse rates were very low  in patients with a combination of HBsAg < 50 IU/mL and HBV DNA < 2 × 105 IU/mL. In contrast, the virological and clinical relapse rates were high  in patients with a combination of HBsAg ≥ 50 IU/mL and HBV DNA ≥ 2 × 105 IU/mL. Thus, close monitoring is needed after the cessation of entecavir and lamivudine therapy in HBeAg-negative patients with a combination of end-of-treatment HBsAg ≥ 50 IU/mL and HBV DNA ≥ 2 × 105 IU/mL at entry.A previous study showed that HBV DNA ≤ 2 × 1022. Our study also found that the HBsAg level at 6 months post-therapy was a significant factor for predicting virological relapse, clinical relapse, and HBsAg loss. Furthermore, patients who had any decline of HBsAg levels from end of treatment to 6 months post-treatment had a lower virological and clinical relapse rates in high-risk groups than those with increased HBsAg levels. Thus, it is worthwhile to check HBsAg levels at 3 to 6 months after cessation of therapy for early diagnosis of HBV flare, especially in high-risk groups for HBV relapse.Investigations of the role of post-treatment HBsAg levels in HBV relapse after cessation of NA therapy were rare. A recent study showed that the HBsAg levels were higher in cases of relapse at 6 months off therapy than among non-relapse cases23. A recent study found that 9% of the HBeAg-positive patients and 14% of HBeAg-negative patients became HBsAg-negative at 3 years after NA discontinuation24. Our study is consistent with these studies24. We also found that patients with HBsAg ≤ 50 IU/mL, 51–100 IU/mL, and 101–150 IU/mL had similar rates of HBsAg loss . Patients with HBsAg of 151–200 IU/mL had a relatively low rate of HBsAg (20.3% at 72 months). Furthermore, the 6-year cumulative rate of HBsAg loss was 61.2% in 74 patients without virological relapse. NAs have potent antiviral activity but do not have a direct immunomodulatory effect. Thus, HBsAg seroclearance is rare during NA treatment even with the most potent NA27. The host immune mechanisms that are modulated during the long period of HBV suppression by NA treatment react effectively to the resumed HBV replication. However, for such an immune clearance to occur, the immune system needs not only to become modulated but also to be re-exposed to the antigens of the replicating virus. Thus, stopping NA might more often lead to HBsAg loss than maintaining NA therapy.Despite the high HBV relapse rate, we found a relatively high rate of HBsAg loss after cessation of NA therapy in HBeAg-negative patients who achieved end-of-treatment HBsAg ≤ 200 IU/mL. The 6-year cumulative rate of HBsAg loss was 54.9% in these patients. A previous study of HBeAg-negative patients reported a high rate of HBsAg loss of 39% within 6 years after adefovir therapy discontinuation, and 13 (72%) out of 18 patients who achieved a sustained response after stopping adefovir therapy were clear of HBsAg28. However, the mechanism of the impact of genotype C infection on HBsAg loss in HBeAg-negative patients remains unclear.In addition to HBsAg, we found that HBV genotype C was another independent factor for predicting off-therapy HBsAg loss in these patients. A recent study in Taiwan showed that compared to genotype B, genotype C infection is associated with a higher chance of spontaneous HBsAg loss in HBeAg-negative patients29. But a recent study showed that 2 of 27 patients with cirrhosis developed hepatic decompensation when off NA therapy22. In our study, none of patient experienced hepatic decompensation during follow-up or timely retreatment, even among cirrhotic patients. Thus, NA therapy might be safely stopped if proper off-therapy monitoring is provided to restart therapy in a timely manner in HBeAg-negative patients who achieve end-of-treatment HBsAg ≤ 200 IU/mL.Most of the current practice guidelines recommend continuing NAs until the loss of HBsAg because of the risk of hepatitis flares and failure. However, recent studies have shown that NA therapy can be safely stopped, even in cirrhotic patients, if proper off-therapy monitoring is provided to restart therapy in a timely mannerlarge-scale studies are needed to confirm the results. Second, we only included Asian populations, in which HBV genotypes B and C are predominant30, as well as patients who discontinued entecavir and lamivudine treatment. Thus, it is unclear whether the same HBsAg and HBV DNA cut-offs are applicable to other HBV genotypes or for those who have discontinued tenofovir therapy.Our study has a few limitations. First, the case number is limited, so further 5 IU/mL at entry experienced the lowest rate of HBV relapse, and this was a better marker for predicting HBV relapse. Patients with a combination of HBV DNA ≥ 2 × 105 IU/mL at entry and end-of-treatment HBsAg ≥ 50 IU/mL had a high HBV relapse rate, and close monitoring is suggested. Patients who achieved end-of-treatment HBsAg ≤ 150 IU/mL had a high rate of HBsAg loss during long-term follow-up. NA therapy might be safely stopped if proper off-therapy monitoring is provided to restart therapy in a timely manner in HBeAg-negative patients who achieve end-of-treatment HBsAg ≤ 200 IU/mL.In conclusion, after cessation of lamivudine or entecavir treatment in HBeAg-negative patients who achieved end-of-treatment HBsAg ≤ 200 IU/mL, the 5-year rates of virological and clinical relapse and HBsAg loss at 60 months were 39.4%, 27.6%, and 45.9%, respectively. Although the HBV relapse rate was not very high in these patients, patients with a combination of end-of-treatment HBsAg < 50 IU/mL and HBV DNA < 2 × 1019. From 2007 to 2011, 240 HBeAg-negative CHB patients who received entecavir treatment and had stopped for at least 12 months were eligible for the analysis. All patients fulfilled the antiviral agent stopping criteria according to the Asian Pacific Association for the Study of the Liver (APASL) 2012 guidelines31. Taiwan’s National Health Plan reimbursed CHB patients for oral antiviral agents for 18 months before November 2009 and for 36 months since then. Thus, these patients discontinued NA therapy if they did not want to bear the expenses themselves. Patients were excluded if there was any evidence of autoimmune hepatitis, alcoholic liver disease, markers of hepatitis C virus (HCV), hepatitis D virus, or human immunodeficiency virus (HIV). Patients were also excluded if they lost HBsAg during treatment or had decompensated liver cirrhosis, HCC, or immunosuppressive therapy. Of the 356 HBeAg-negative patients, 119 patients (28 cirrhotic patients) who achieved end-of-treatment HBsAg ≤ 200 U/L were enrolled in this study (34 lamivudine (treatment duration: 95.8 ± 45.7 weeks), 85 entecavir (treatment duration: 174.2 ± 51 weeks)).From 2002 to 2007, 116 HBeAg-negative NA-naïve CHB patients who received lamivudine treatment and had stopped lamivudine for at least 12 months were considered for analysis. All patients fulfilled the antiviral agent stopping criteria according to our previous study9/L) × [ALT (U/L)]1/2}, where AST represent aspartate aminotransferase32.Liver biopsy was performed in 14 patients before treatment. None of 14 patients showed cirrhosis. Cirrhosis was diagnosed according to repeated ultrasounds with consistent findings suggestive of cirrhosis in addition to clinical features such as splenomegaly, thrombocytopenia, ascites, or gastroesophageal varices. FIB-4, a simple noninvasive test for liver fibrosis, was defined by the following formula: age (years) × AST(U/L)/{platelets  elevation >2× the upper limit of normal (ULN) (40 U/L) plus HBV DNA > 2000 IU/mL after stopping entecavir treatment31. Consolidation duration was calculated from the first demonstration of undetectable HBV DNA to the end of treatment. The criteria of retreatment were HBV DNA ≥ 2000 IU/mL with ALT > 2× ULN in non-cirrhotic patients and whenever HBV DNA ≥ 2000 IU/mL was detected regardless of ALT level in cirrhotic patients.Post-treatment virological relapse was defined as a serum HBV DNA level > 2000 IU/mL in two consecutive measurements obtained at least 3 months apart after the cessation of NA treatmentThe study was conducted in accordance with the Helsinki Declaration of 1975. The study was approved by the Ethical Committee of the Chang Gung Memorial Hospital (103-7677B). All patients provided written informed consent.After cessation of lamviudine and entecavir therapy, the patients were followed up every 1 to 3 months. Additional biweekly or weekly visits were arranged if ALT levels increased by more than 2× ULN. The follow-up studies included liver biochemical tests and serological hepatitis B markers. HBV DNA was monitored every three months in the first 12 months and then every 6 months thereafter. HBV DNA was further assessed when clinical relapse was found. If HBV DNA increased beyond 2000 IU/mL during follow-up, HBV DNA was retested 3 months later for confirmation. Ultrasonography was performed for HCC surveillance every 3 to 6 months. Serum HBsAg quantification was performed at baseline, 12 months, the end of treatment during NA therapy, and 6 months after cessation of NA therapy. All serum HBsAg levels were quantified retrospectively using stored serum samples, which had been frozen at −20 °C until use.The presence of HBsAg, HBeAg, anti-HCV antibodies, and anti-HDV antibodies were determined using commercial assay kits . HBsAg titers were quantified with ElecsysHBsAg II Quant reagent kits  according to the manufacturer’s instructions with a lower limit of detection of 0.05 IU/ml. HBV DNA was quantified using the COBAS TaqMan HBV test  with a lower limit of detection of 20 IU/ml.30.The HBV genotypes were determined using restriction fragment length polymorphism on the surface gene (between nucleotide positions 256 and 796), as described previouslyt test was used for continuous variables. The cumulative incidences of post-treatment virological and clinical relapse as well as HBsAg loss were analyzed using the Kaplan-Meier method with a log-rank test. Univariate and multivariate analyses were performed using Cox proportional hazards regression models. A time-dependent receiver-operating characteristic (ROC) curve analysis was used to assess the best HBsAg cut-off level for predicting virological and clinical relapse within 5 years33. All statistical tests were two-sided with the significance level set to 0.05.Data are presented as the mean ± standard deviation (SD), proportion, or median (range). To compare values between the two groups, the chi-square or Fisher’s exact tests were applied to analyze categorical variables, while the Student’s