Recently, several reports issued clevudine induced myopathy in the long term use.The aim of this study was to investigate antiviral effects and adverse events of clevudine monotherapy in patients with chronic hepatitis B (CHB).The subjects were 110 treatment-naïve CHB patients. They were treated with 30 mg clevudine/day for more than six months. Virological and biochemical tests, including that for serum creatine kinase (CK), were monitored at baseline and at 3-month intervals during treatment period.In HBeAg-positive patients, the cumulative rates of virological response were 74.0 %, 68.5 %, and 67.3 % after one, two, and three years of clevudine treatment, respectively. Cumulative rates of HBeAg loss or seroconversion were 17.8 %, 30 %, and 31.5 % after one, two and, three years of clevudine treatment, respectively. In HBeAg-negative patients, the cumulative rates of virological response were 97.3 %, 100 %, and 94.6 %, respectively. Virological breakthrough occurred in 27 patients. The rtM204I mutation in HBV polymerase was predominantly detected. Muscular adverse events were observed in 15 patients. All patients with myopathy recovered after the cessation of clevudine monotherapy. Fluctuations in CK level during the clevudine treatment period were frequently observed irrespective of development of myopathy. Multiple episodes of CK elevation were significantly related to the development of myopathy.Long-term clevudine monotherapy is effective for suppression of serum HBV DNA level and normalization of serum alanine amino transaminase levels, but associated with occurrence of rtM204I mutation. Clevudine-induced muscular adverse events are not uncommon, although they are totally reversible after cessation of the treatment. Muscular adverse events and serum CK level should be carefully monitored during long-term treatment with clevudine. Treatment of chronic hepatitis B virus (HBV) infection with nucleoside/nucleotide antiviral agents aims for the prolonged suppression of viral load and prevention of progress to decompensated liver cirrhosis and hepatocellular carcinoma. To achieve this goal, the antiviral agent should be a potent suppressor with sustained effects, and should be safe for long-term use . ClevudiThe present study was thus conducted to assess virological response and muscle-related adverse events associated with long-term monotherapy with clevudine.The study subjects were 110 chronic hepatitis B (CHB) patients who were enrolled consecutively between January 2007 and June 2009 at Konkuk University Medical Center. Hepatitis B surface antigen (HBsAg) had been detectable in the serum of all enrolled patients for more than six months, their serum alanine amino transaminase (ALT) level was greater than twice the upper limit of normal (ULN) for chronic liver disease, and they had not been treated previously with anti-HBV nucleosides or nucleotides. Those with HBeAg-positive CHB exhibited a serum HBV DNA level greater than 5 log10 copies/ml, while in those with HBeAg-negative CHB and/or with compensated cirrhosis, it was greater than 4 log10 copies/ml. Exclusion criteria included patients with chronic renal failure, decompensated liver disease, and co-infection with hepatitis C, hepatitis D, and HIV. Pregnant or breastfeeding women and women of child-bearing age who were unwilling to use barrier contraceptive devices were also excluded. Cirrhosis was diagnosed clinically based on imaging findings  and compatible clinical features . The study protocol was approved by the Institutional Review Board of Konkuk University Medical Center (KUH 1010277). All of enrolled patients gave their written, informed consent for storing their sera and for the biochemical tests and HBV genetic studies.All 110 treatment-naïve patients received 30 mg of clevudine once daily for more than six months from January 2007 onwards. They were monitored at baseline and every three months during clevudine therapy. They were assessed clinically by obtaining their medical history, and subjecting them to a physical examination, electrocardiography, and testing of blood HBeAg status  and serum HBV DNA levels. At the three-monthly follow-ups during the treatment period, the patients were assessed clinically to determine their tolerability, and subjected to a physical examination, blood chemistry, and HBV status testing. Serum HBsAg levels were tested after 12 months of therapy. Serial blood samples were collected and stored frozen at -80℃ from each patient at the time of initiation of each antiviral agent, every three months during treatment, and at the time of viral breakthrough. Serum HBV DNA levels were assessed using the COBAS Amplicor polymerase chain reaction (PCR) assay, which has a lower limit of detection of 300 copies/ml .Primary non-response was defined as a decrease in serum HBV DNA to less than 2 log10 copies/ml at six months therapy . PrimaryAll data were entered into a database and analyzed using SPSS 13.0 for Windows software . Continuous variables are presented as the mean ± SD values, and categorical variables as absolute and relative frequencies. Serum HBV DNA level and virological breakthrough were analyzed by t-test. The incidence of undetectable levels of serum HBV DNA levels and virological breakthrough were analyzed using Pearson’s 2 tests. P values less than 0.05 were considered statistically significant.A total of 110 patients were studied. The baseline characteristics of patients are summarized in Of HBeAg-positive patients, serum HBV DNA was undetectable in 64.4 %, 74.0 %, 68.5 %, and 67.1 % at six, 12, 24, and 36 months of clevudine treatment, respectively; the corresponding values for HBeAg-negative patients were 94.6 %, 97.3 %, 100 %, and 94.6 % . There wP = 0.345). Thus, initial serum HBV DNA level was not a risk factor for developing viral breakthrough. The rate of undetectable levels of HBV DNA at six months of treatment is significantly lower in the group that did not develop viral breakthrough (P = 0.001). Other factors, such as age, sex, HBeAg status, cirrhosis, underlying disease, fatty liver, and follow-up duration, were not correlated with developing viral breakthrough. Virological resistance was examined by RFMP for rt204 and rt180 at the time of virological breakthrough in 25 patients. The rtM204I mutation in the HBV polymerase gene was detected in 24 patients; the remaining patients had wild-type rt204 and rt180. Of 24 cases with the rtM204I mutation, three had co-dominant rtM204I and rtL180M mutations. One patient had a mixed pattern of rtM204I/V and rtL180M mutations.Viral breakthrough occurred in 27 patients during the follow up period. The cumulative incidence of viral breakthrough in HBeAg-positive patients was 11.8 % and 22.7 % at 12 and 24 months of therapy, while that in HBeAg-negative patients was 9 %, 23 %, and 24.5 % at 12, 24, 36 months, respectively. Baseline serum HBV DNA levels were 7.1 ± 1.4 log10 copies/ml and 6.4 ± 1.2 log10 copies/ml in the groups that did not and did develop viral breakthrough, respectively; the difference was not statistically significant (P = 0.001). 93.3 % in myopathy group and 40.5 % in non-myopathy group showed at least two episodes of abnormal elevation of CK (P = 0.001). 40.0 % in myopathy group and 15.5 % in non-myopathy group showed at least one episode of toxic grade above 1 of CK (P = 0.026). No patient of both groups showed toxic grade above 3 of CK. There were frequent fluctuations in CK level during clevudine treatment in both groups. Hematologic and biochemical factors were not associated with developing myopathy during clevudine therapy (P = 0.048) and abnormal episodes of CK elevation (P = 0.006) including toxic grade 1 or 2 CK elevations (P = 0.008)  .This was an observational study of the efficacy and adverse events during long-term monotherapy with clevudine in treatment-naïve CHB patients. Clevudine monotherapy showed a potent antiviral effect within 12 months of treatment in both HBeAg-positive and HBeAg-negative patients. Phase III clinical trials of clevudine therapy demonstrated that 59.0 % of HBeAg-positive patients and 92.1 % of HBeAg-negative patients had undetectable levels of HBV DNA after 24 weeks of treatment . Similar


Among those with HIV, anemia is a strong risk factor for disease progression and death independent of CD4 count and viral load. Understanding the role of anemia in HIV treatment is critical to developing strategies to reduce morbidity and mortality. We conducted a prospective analysis among 10,259 HIV-infected adults initiating first-line ART between April 2004 and August 2009 in Johannesburg, South Africa. The prevalence of anemia at ART initiation was 25.8%. Mean hemoglobin increased independent of baseline CD4. Females, lower BMI, WHO stage III/IV, lower CD4 count, and zidovudine use were associated with increased risk of developing anemia during follow-up. After initiation of ART, hemoglobin improved, regardless of regimen type and the degree of immunosuppression. Between 0 and 6 months on ART, the magnitude of hemoglobin increase was linearly related to CD4 count. However, between 6 and 24 months on ART, hemoglobin levels showed a sustained overall increase, the magnitude of which was similar regardless of baseline CD4 level. This increase in hemoglobin was seen even among patients on zidovudine containing regimens. Since low hemoglobin is an established adverse prognostic marker, prompt identification of anemia may result in improved morbidity and mortality of patients initiating ART. Anemia has been shown to be the most frequent hematological abnormality in HIV-infected patients globally , 2. EvenThe major cause of anemia is impaired erythropoiesis resulting from the release of inflammatory cytokines and decreased production of hematopoietic growth factors, coupled with malabsorption and impaired recycling of iron , 10. AddIn this prospective cohort study conducted in Johannesburg, South Africa, we set out to determine the prevalence, incidence, and predictors of anemia among patients initiating first line ART. We also sought to explore whether the degree of immunosuppression at ART initiation or the initiating ART regimen (zidovudine versus other) impacted the magnitude of hemoglobin increase while on treatment.The Themba Lethu Clinic is one of the largest HIV care and treatment clinics in South Africa with over 22,000 patients initiated onto ART since 2004 . We inclAt Themba Lethu, ART is provided according to the South African Department of Health Guidelines . All patFor all patients, person-time accrued from ART initiation until the date of the begining of (1) incident anemia; (2) last clinic visit; (3) loss to follow-up; (4) death; (5) transfer; or (6) completion of 24 months of follow-up. Loss to follow-up was defined as 4 months since last clinic visit. At Themba Lethu, mortality is ascertained via family or hospital report, active tracing, and linkage with the South African National Vital Registration Infrastructure Initiative. We used the National Institutes of Health Division of AIDS definitions for Grading the Severity of Adult and Paediatric Adverse Events to define anemia . Anemia We calculated the prevalence and incidence of anemia at ART initiation and during follow-up, respectively. Prevalence ratios (PR) and 95% confidence intervals (95% CI) for the association of baseline characteristics with prevalent anemia were estimated using modified Poisson regression. Cox proportional hazards models were used to evaluate the association between incident anemia and baseline (defined as 90 days before to 7 days after ART initiation) characteristics such as hemoglobin, CD4 count, body mass index (BMI), and WHO stage. We further examined overall changes in hemoglobin levels on treatment by stratifying results by CD4 count and ART regimen (AZT versus non-AZT containing regimen) at treatment initiation.Use of Themba Lethu Clinic data was approved by the Human Research Ethics Committee  of the University of Witwatersrand, and approval for analysis of anonymized data was approved by the Boston University Institutional Review Board.Between April 2004 and August 2009, 10,369 patients fulfilled the inclusion criteria. Among these, 102 (1%) did not have hemoglobin measurements, and 8 patients had abnormally high hemoglobin levels (>30 g/dL) at ART initiation leaving 10,259 patients for analysis. Excluded patients were largely similar in demographic and clinical characteristics from those who were included in analyses. The majority of patients were females (68%), with a median age of 36 years (IQR 31–43) and initiated on stavudine-lamivudine-efavirenz (89.4%) . The cohn = 322) of the patients were initiated on to AZT-based regimens. Of note, among these 322 patients initiated onto AZT-based regimens, 47 patients corresponding to 14.6% already had hemoglobin levels of less than 10 g/dL at baseline. Additionally, we found that those with a WHO III/IV versus I/II , patients with a low CD4 count , and those with tuberculosis at initiation  were more likely to have anemia at ART initiation. Overall prevalence of anemia at ART initiation was 25.8% (95% CI: 25.0%–26.7%). In multivariable analysis, females  and younger patients  were more likely to be anemic at ART initiation , while pA total of 7,612 patients had normal hemoglobin levels at baseline. The incidence rate for anemia among these patients was 3.1/1000 person-years (PY). Incidence of anemia was the highest in the first 3 months after ART initiation at a rate of 34.7/1000 PY and decreased to 16.7/1000 PY between 3 and 6 months on treatment. This incidence was then fairly stable over the remaining 18 months of follow-up, between 7.7 and 8.4/1000 PY. In adjusted analyses  female gWe found that AZT use was associated with over twice the rate of developing anemia during follow-up . In a sensitivity analysis to decrease measurement error, we further restricted the analysis to patients without an indication for AZT. Our hazard ratios were similar though the association between AZT and incident anemia was slightly attenuated, from 2.19 to 1.88 (95% CI: 1.17–3.03). Among the 10,259 patients initiating ART, hemoglobin increased over 24 months on ART . BetweenThis is a study of patients under routine public-sector HIV care in Johannesburg, South Africa, over a quarter of patients presented at treatment initiation with anemia Hb < 10 g/dL). This is slightly higher than previous estimate of 18% of prevalence of anemia among HIV-infected patients in Southern Africa but in line with estimates for other regions of Africa (between 24% and 37%) . We esti0 g/dL. TIn our data, incident anemia was associated with female gender, lower BMI, advanced WHO stage, and lower CD4 count. The association of anemia with lower BMI is likely to reflect malnutrition, as a number of nutritional deficiencies contribute to anemia. The increased risk in anemia among patients with immunosuppression is in agreement with previous published reports , 21–24. The present study also showed that over 24 months of follow-up, hemoglobin increased for patients initiating ART, similar to findings from resource-rich environments . This inOur study has several strengths including large number of clinical events owing to the relatively large sample size. Also, we use routine clinical data collected from a real life clinical setting. However, the findings of this study should be considered alongside its limitations. We did not collect comprehensive information of clinical conditions that may be possible underlying causes of anemia, resulting in unmeasured confounding of the risk factors for prevalent and incident anemia. Additionally, loss to follow-up may have led to selection bias, as outcome cannot be determined in patients lost or those who have died. In conclusion, we observed a high prevalence of anemia among HIV-infected patients initiating first-line ART in Johannesburg, South Africa. Over 24 months of follow-up, incidence of anemia was the highest in the first 6 months after ART initiation, but, overall, during follow-up, this remained low. Risk factors for prevalent and incident anemia were similar to those reported elsewhere. Our results further support findings that ART improves hemoglobin and this is seen regardless of regimen type (AZT containing versus non-AZT containing) and the degree of immunosuppression. Hemoglobin measurement is an inexpensive marker for HIV disease progression and should be measured more frequently in patients at risk for anemia in the first 6 months after initiation of ART. Our study suggests that ART improves hemoglobin levels regardless of magnitude of immunosuppression and ART initiating regimen.


We describe two patients who did not experience a viral rebound after cessation of HAART which was initiated for progressive disease. CD4 T-cell count remained stable in one patient and progressively declined in the other, despite apparent viral control. We failed to identify any immune activation or genetic markers that could offer an explanation for this unusual "secondary controller" status. But their viruses are clearly less fit compared to viruses from rebounders. Highly active antiretroviral treatment (HAART) has improved life expectancy and quality of life of HIV infected patients . HoweverIn 2008 we identified a patient who, after stopping HAART, had a VL below the detection limit of 1,6log for more than 2 years. After searching the data base of 1,700 HIV infected patients under follow-up at the clinic of the Institute of Tropical Medicine, another patient was identified, who spontaneously controlled VL after cessation of HAART for at least 6 months. We labeled these two patients "secondary controllers" (SC) as opposed to "elite controllers" (EC) who spontaneously control viral replication but are treatment naive.In order to identify correlates of SC status, we compared clinical characteristics, viral factors, genetic traits and immune activation markers in the two SC and patients who rebound after treatment cessation. We used the following criteria to define SC and rebounders: (1) being HIV-1 infected for at least 3 years; (2) HAART started in the chronic progressive phase; (3) HAART discontinued for the first time; and (4) VL below the detection limit of 1.6log at treatment interruption. Pregnant women taking HAART to prevent mother-to-child transmission and patients who started treatment in the acute phase of infection were excluded. SC were defined as patients who met the above criteria and who kept their VL below 3log for at least 6 months after treatment cessation. In order to make sure that the SC were not taking antiretroviral drugs, plasma samples were tested for the presence of protease inhibitors (PI) and non nucleoside reverse transcriptase inhibitors at the Bioanalytical Laboratory of the Catholic University of Louvain.The following parameters were assessed: HIV subtype, CD4 count, VL and number of days on HAART and were compared in SC and rebounders. In addition, we evaluated plasma markers of immune activation, including: neopterin; beta-2 microglobulin (B2M) ; soluble CD14 (sCD14) ; and lipopolysaccharide (LPS) . These measurements were done on stored plasma samples taken before starting HAART (T1), during HAART (T2) and 6 months after stopping HAART (T3). Soluble markers in plasma were measured instead of activation markers on cells (Roberts AIDS 2010), because at our clinic only plasma samples of patients are stored.In order to assess viral factors, the SC provided 100 ml of blood for purification of CD4+ T-cells. One part was used to extract DNA, as described elsewhere , for seq-3 with the viruses, IIIB and BAL as reference strains , and with virus isolated from SC patient 1 and from 4 different rebounders. P24 concentrations were measured by ELISA over time.In addition, we evaluated the relative replication capacity of P1. To assess viral fitness phytohemagglutinin (PHA) stimulated peripheral blood mononuclear cells (PBMC) from 3 healthy donors were infected at multiplicity of infection (MOI) 10This study was approved by the Institutional Review Board of ITM and the Ethics Committee of the University Teaching Hospital in Antwerp. Informed consent was obtained from all patients.When searching our database on 1700 patients in regular follow-up, we found that a total of 160 patients had stopped HAART for the first time, after a mean of 25 months of successful viral suppression. Treatment was stopped under medical guidance, mostly because of toxicity and/or because the patient wanted a drug holiday. At least two VL measurements after treatment cessation were available for 124 patients, which enabled us to classify them as "rebounders" or "SC". Median time to rebound (VL > 3log) was 7 weeks and median time to re-starting treatment was 4.5 months. Sixty-eight patients remained off treatment for at least 6 months. By that time all but 2 patients (the SC) had a VL that exceeded 3log. Both SC were negative on the drug assays.Patient 1 was diagnosed with HIV infection in 1985. In 1993 he started treatment with AZT, when his CD4 count was 228/μL. VL testing was unavailable at that time and no plasma samples from 1993 were available at the clinic in Antwerp. The first VL measurement dated from 1999 when the patient was already taking AZT monotherapy. It was 3.4 log and a PI based HAART regimen was started and continued till 2001, when the patient was switched to Nevirapine and 2 nucleoside reverse transcriptase inhibitors (NRTI's). He took this combination till treatment interruption in April 2005, when his CD4 count was 458/μl. In July 2005 the VL was 3.9log but from October 2005 until now it remained below the detection limit. The CD4 count, however, has been decreasing to 205/μL in February 2010 but the patient refuses to be restarted on treatment.Patient 2 was found to be HIV positive in 1997, but was probably infected since 1991. In 1998 her CD4 count was 377/μL and her VL 4.6log. She was started on HAART with a PI based regimen. In 1999 she switched to Nevirapine and 2 NRTIs. Treatment was stopped in March 2006. The VL remained undetectable after treatment interruption, though in April 2009 there was a blip in the plasma VL of 2,9log, which was detected with the Cobas ampliprep/cobas Taqman . The patient is still off treatment with a stable CD4 count above 1000/μL.Table Virus measured as p24 in culture supernatant of purified CD4+ T-cells from patients under HAART, typically emerges after 3 to 6 days. In contrast, in patient 1 it emerged on day 21, while in patient 2 virus culture failed despite three attempts. In 2009, however, the latter patient had a blip in plasma virus suggesting that the virus is not completely defective. Moreover the sequences of Gag, Pol and Env in the two SC did not show any obvious defects, and no CD8 T-cell associated escape mutations could be detected. Remarkably, phylogenetic analysis of env from patient 1 revealed the presence of two related but distinct viruses. Both viruses were lacking the lentivirus lytic peptides (LLP) 1 region, but could still be cultured.Autologous viruses from patient 1 could be cultured making it possible to investigate whether the virus of this patient was less fit in comparison to reference strains or virus of 4 different rebounders. Figure We identified two HIV-1 infected patients with an exceptional clinical course after stopping HAART. One patient kept stable and high CD4 T-cell counts for more than 4 years after treatment cessation, while the other patient had progressive disease with decreasing CD4 T-cell counts. Such discrepancy of evolution has also been described in elite controllers -9.To our knowledge there is only one previous study that had identified patients who maintained an undetectable viral load for several months after treatment cessation. Skiest et al mentioned 2 patients who had an undetectable viral load for 6 months after treatment was stopped as part of a study on the effects of treatment interruption. However, they just mention them without comparing them with patients whose viral load rebounded . We are One possible explanation for the SC status is that deleterious mutations have been induced in the virus by HAART. However, sequencing of the major structural genes of the 2 available viruses did not reveal any obvious defects. Virus could be cultivated in patient 1 only, but patient 2 had a spontaneous "in vivo" blip. We hypothesize that these replication competent viruses are less fit and/or are kept under control by the patient's immune system. Indeed, as shown in Figure Markers of immune activation are more disturbed in patients with rapid progression and closer to normal in EC ,13. NeopResearch on EC is very active because these patients may give clues for novel approaches to the development of a preventive HIV vaccine. Research on immune function in the secondary controllers in this study is ongoing; as we believe that they might provide new insights about correlates of protection and be useful for the development of therapeutic vaccines.Written informed consent was obtained from the patients for publication of this case report. A copy of the written consent is available for review by the Editor-in -chief of this journal.The authors declare that they have no competing interests.EVG performed all culture experiments and ELISAs for immune activation, contributed to the study design, data analysis and interpretation of the data; and wrote the paper. LH was responsible for the molecular biological aspects of this study and performed all sequence analyses. LB: performed the experiments to investigate the replicative fitness.SC performed all DNA isolations and amplifications. EF was responsible for the clinical aspects of the study. He identified the patients, abstracted the clinical information and recruited the patients in the study. AB contributed to the study concept, and the writing of the project proposal and the paper. PL performed all the statistical data analyses. GV supervisor and principal investigator of the project, contributed to the study design, data analysis and interpretation of the data. All authors have read and approved the final manuscript.EVG is a postdoc at the Institute of Tropical medicine. During her PhD she was trying to develop a therapeutic vaccination against HIV based on dendritic cells electroporated with mRNA encoding HIV Gag. In this studies patients on HAART were vaccinated with these DC. After vaccination it is the goal that they stop treatment and control the virus. Therefore these patients are so special because they can do it without vaccination. If we can find out why than we know to which parameter we have to look for protection in a therapeutic vaccination. AB, EF, GV are clinicians. EF is really working in the clinic and seeing patients. AB and GV are more interested in research. For EF is it also important if we can unravel why these patients control the virus after treatment interruption, maybe more patients can go on drug holiday.


Although highly active antiretroviral therapy (HAART) improves immunologic parameters, the relationship between DTH responsiveness and CD4 gains on HAART is not completely understood. We investigated CD4 reconstitution and the change in DTH responses from treatment baseline through 24 months of viral load (VL)-suppressive HAART in the U.S. Military HIV Natural History Study.Delayed-type hypersensitivity (DTH) testing is an n=302). DTH testing consisted of ≥3 recall antigens, and responses were classified by the number of positive skin tests: anergic (0–1) or non-anergic (≥2). Pre-HAART DTH results were compared for the outcome of CD4 reconstitution at 24 months of HAART. Improvement in DTH responses was also analyzed for those anergic before HAART initiation.Treatment-naïve subjects with VL <400 copies/mL after ≥24 months on HAART were included (p=0.104) and VL . Median CD4 gains 24 months after HAART initiation were similar between the non-anergic  and anergic groups . For individuals anergic before HAART initiation, DTH normalization occurred at 24 months post-HAART in the majority of participants . Normalization of DTH responses was not associated with CD4 count at HAART initiation  nor with AIDS diagnoses prior to HAART .Non-anergic responses were observed in 216 (72%) participants, while 86 (28%) individuals were anergic prior to HAART initiation. Demographically there were similar distributions of age at HIV diagnosis and HAART initiation, as well as gender and race or ethnicity. There were no significant differences between non-anergic and anergic participants in pre-HAART CD4 count  315–517 vs. 373 cells/μL, IQR 228–487; DTH responsiveness has been shown to predict HIV disease progression independent of CD4 count in untreated individuals. In the setting of HAART, pre-HAART anergy does not appear to impact CD4 gains or the ability to normalize DTH responses after 24 months of VL-suppressive HAART. The gDTH responsiveness is typically reduced in patients living with HIV compared to seronegative individuals , 6. For DTH responses have also been studied in patients receiving HAART. In addition to immune reconstitution measured clinically by CD4 cell gains, several studies demonstrated an improvement in DTH responses after initiation of HAART –14. HoweThe NHS is a prospective observational cohort of over 5500 military members, dependents, and beneficiaries who have been evaluated for HIV-1 positive status in the military healthcare system since 1986 . Study vThe NHS database was queried for participants on HAART who also received serial DTH testing for this retrospective analysis. Participants achieving a VL <400 copies/mL within six months of their initial HAART regimen and maintaining VL suppression for ≥24 months were included. The analysis was restricted to those achieving VL suppression because the outcome of immune reconstitution during HAART was the primary focus of this study and maximal CD4 gains occur when VL is fully suppressed. The association between pre-HAART DTH response and the outcome of VL suppression was not analyzed because data for some confounders, including HIV genotype and HAART adherence, were not available. Individuals were required to have DTH testing within six months prior to HI and repeat DTH testing performed at 24 months on HAART. DTH testing was performed according to standardized protocols, as previously described , 8, 13. p-values were calculated using t-tests or Wilcoxon tests for continuous variables depending on normality, and chi-squared tests or Fisher's exact tests for categorical variables depending on sparse cells. To estimate the impact of DTH status at HI on post-HAART immune function, we modelled CD4 gains at 24 months after HI with multiple linear regression while adjusting for other relevant covariates. Multiple logistic regression was used to understand factors associated with post-HAART DTH normalization among those who were anergic at HI.Baseline characteristic p=0.015). Median CD4 count at diagnosis was significantly higher in the non-anergic group  360–656) compared to the anergic group ; however, VL was no different .A total of 302 participants met inclusion criteria, of which 216 (72%) were non-anergic and 86 (28%) were anergic by DTH testing prior to HI . Demograp=0.104). VL at HI was also similar between groups. AIDS diagnoses prior to HI were more common in the anergic group  compared to the non-anergic group . The initial HAART regimen most commonly used was protease inhibitor (PI) based  compared to non-nucleoside reverse transcriptase inhibitor (NNRTI) based  and other regimens that typically contained both PIs and NNRTIs . For DTH test outcomes, there was a trend toward greater PI-based regimen use in the anergic group  compared to the non-anergic group .The year of HI was typically in the early HAART era (1998 for both groups). The median CD4 at HI was higher for non-anergic  versus anergic  participants, but this was not statistically significant (p=0.498). Linear regression was performed to investigate the factors associated with CD4 gains (p<0.001). Other variables, including diagnosis of AIDS prior to HI or type of HAART regimen, were not significantly associated with CD4 gains at 24 months.The median CD4 cell gains after HI were similar for non-anergic compared to anergic subjects, even when stratified by pre-HAART CD4 count ≤400 or >400 cells/μL . At 24 mD4 gains . Non-anen=86) was studied for factors associated with normalization of DTH responses at 24 months of HAART (p=0.129) nor AIDS prior to HI  was associated, with DTH normalization. Race or ethnicity was associated with DTH status at 24 months with African Americans  and Hispanic/others  more likely to become non-anergic than European Americans. Those whose first HAART regimen was NNRTI based compared to PI based were less likely to become non-anergic , but no other covariates were associated with DTH normalization.The subset of participants who were anergic at HI  jiroveci pneumonia prophylaxis was previously recommended in patients with CD4 >200 cells/μL combined with anergy [in vivo, DTH results can provide information about immune function in addition to the measurement of CD4 counts. DTH anergy has also been suggested as an indicator for initiating HAART in resource-limited settings [The utility of DTH testing has been demonstrated in many studies and was previously recommended in the evaluation and disease staging of patients living with HIV. For example, DTH responsiveness has been shown to be an independent predictor of opportunistic infections and death , 6, 8, 9h anergy . Since Dsettings , 11. AltDTH responsiveness to recall antigens in HIV-infected patients is associated with the magnitude of the CD4 count at the time of testing. For example, one study in our military population found that 60% of treatment-naïve patients with CD4 counts less than 200 cells/μL were completely anergic, whereas 86% with >400 cells/μL were non-anergic . Althouget al.[et al., 40% of participants did not achieve VL suppression on HAART, whereas our study included only VL-suppressed individuals [HAART has been previously shown to enhance DTH responsiveness in patients living with HIV who are anergic. In a study by Carr et al., DTH reset al., 22. A set al.. In the ividuals . In contividuals . We previviduals . Thus, iin vitro loss of T-cell function in patients living with HIV was progressive and sequential, beginning with impairments to antigen recall responses, then alloantigen responses, and ultimately mitogen-induced responses [One interesting observation in this study was that DTH normalization occurred in approximately 60% of anergic participants regardless of pre-HAART CD4 cell count and prior AIDS diagnoses. A previous study showed that esponses . Since rn=14) compared to PI-based HAART (n=64). Alternatively, this finding may also be related to race because four of the five participants on NNRTI-based regimens who failed to normalize DTH responses were European Americans. Further studies are necessary to investigate the impact of racial and genetic background on DTH responsiveness and immune recovery during HAART.Patients in the NHS cohort have free access to healthcare and medications, which mitigates social and economic issues that can confound analyses related to race and ethnicity in other cohorts. Multiple logistic regression analyses in the current study revealed that European Americans were more likely to be anergic before HI and less likely to normalize DTH responses during HAART. Since this finding was not attributable to low CD4 counts or the presence of AIDS diagnoses, other immunologic and/or genetic aspects may have contributed to the lack of DTH responses in this group. For example, CCL3L1-CCR5 GRG status, a genetic characteristic, has been associated with differences in DTH responsiveness . DTH norIn this study, the overall number of DTH responses was recorded, although detail about specific antigen responses was not available. Another potential limitation is that treatment occurred in the early HAART era. However, this was mitigated by the requirement of VL suppression for all participants included in this study. Other factors that may influence DTH responsiveness, such as vitamin deficiencies and nutritional status, were not measured, and some comparisons may have been limited by the relatively small sample size. The overall proportion of participants who were anergic at HI was lower (30%) compared to other studies, but this is likely due to relatively preserved pre-HAART CD4 counts (median 400 cells/μL) and the younger age (median 35 years) at the time of HI.DTH testing is a clinical tool that can be used to assess CMI. Although DTH responsiveness can predict HIV disease progression independent of CD4 count, pre-HAART DTH status does not appear to be predictive of CD4 gains after 24 months of VL-suppressive HAART. In participants with cutaneous anergy prior to HAART, a normalization of DTH responses occurred in approximately 60% of subjects after 24 months of suppressive HAART. In those with cutaneous anergy and advanced HIV disease, as evidenced by low CD4 cell counts and AIDS diagnoses, the capacity for both CD4 reconstitution and normalization of DTH responses during HAART appears to be similar to that of those without advanced disease.


Although dual-boosted protease inhibitors regimen is not recommended in children with HIV infection, such a strategy could be useful in subjects with a complex resistance profile. This study was aimed at assessing the long term efficacy and safety of a double-boosted protease inhibitor combination, fosamprenavir (fAVP) and atazanavir/ritonavir (ATV/r) in a cohort of HIV-infected children and adolescents who had failed with nucleoside reverse transcriptase inhibitors.Seven vertically infected children and adolescents who had previously failed highly active antiretroviral therapy and were resistant to nucleoside reverse transcriptase inhibitors, received a dual protease inhibitor (PI) regimen including fAVP plus ATV/r for 42 months. The patients were assessed at baseline, every month for the first 24 weeks of therapy and every 3 months until month 32. Physical examination, CD4+ cell count, HIV-RNA viral load, lipid profile and hepatic function were assessed throughout the follow up.th cent.le for the males and 94th cent.le for the females. HDL cholesterol showed protective values. Hepatic enzymes remained stable during the entire observation, whereas total bilirubin showed toxicity II/III grade in 6/7 subjects. No change in fat redistribution and insulin resistance was observed.During the study no serious adverse events were reported. CD4 absolute number increased over-time in all subjects. At baseline the median HIV-RNA was 6562 cp/mL (ranging 1048 -102772 cp/mL) and rapidly decreased below the limit of detection (50 cp/mL) after 2 months of the new treatment and remained undetectable in all cases through the entire study period. At the beginning of the study all cases showed a normal lipid profile. During the study period, 4/7 subjects showed total cholesterol, low density lipoprotein and triglyceride levels >97Dual-boosted protease inhibitor therapy was virologically and immunologically effective and it could be considered as a possible alternative to a rescue regimen in children and adolescents. However, hypercholesterolemia and hypertriglyceridemia need close follow-up and may limit the use of this therapeutic option. The introduction of potent combination antiretroviral (ARV) therapy has been found to improve disease progression and prolonged survival in HIV-infected adults and children -3. DespiThe rationale of using a dual-boosted PI regimen is to reach synergistic or additive activity against HIV and to further increase the genetic barrier to resistance, to obtain high plasma concentrations of both PIs with only one boost and to avoid toxicity with N(n)RTI-sparing regimens .The short-term efficacy of double-boosted PI in naïve adults has been disappointing ,9, whileIn pediatric patients, dual boosted PI regimen is not recommended because few data are available in this population . HoweverOur observational study evaluated the immuno-virologic efficacy and the safety profile of dual boosted PI-based therapy in a cohort of vertically HIV- infected pediatric patients who had failed nucleoside N(n)RTI based therapy. Inclusion criteria were: ongoing viral replication, multiple virological failures using a N(n)RTI-containing regimen, and presence of resistance-associated mutations to N(n)RTI and NNRTI in the current genotype., 1717 Deerfield Road, Deerfield, IL 60015-0778, USA) was performed in each patient before the introduction of the dual boosted PI-based regimen. Mutations were defined according to International AIDS Society-USA Drug Resistance Mutation Group ) to month 42 (175 mg/dL [135-194]) with a significant difference (P < 0.05). TG values >97th percentile were detected in 4/7 patients. (57 %) LDL cholesterol remained stable over-time, with baseline values of 75 (64-105) mg/dl and 106 (92-116) at month 42. LDL values > 97th cent.le for males and >95th cent.le for females were detected in patients 4/7 patients (57%). HDL exhibited protective values during the follow up.Fasting lipid profiles  are shown in Figure At baseline, one out of 7 patients had lipohypertrophy consisting in truncal fat accumulation, “buffalo hump” and breast enlargement. In this single patient, lipohypertrophy remained stable during the follow up. No other patients developed signs of lipodistrophy during the double-boosted PI therapy.At baseline all patients had a normal HOMA index (3.8 [1.2-10.1)], which was maintained stable during the follow up.This study showed that at month 42 a combination regimen of fAVP + ATV/r obtained an excellent virological (100% of subjects with HIV-RNA <50 copies/ml) and a good immunological (9% median CD4 increase) outcome in HIV-infected children and adolescents although there was an association of a worsening lipid profile. Our results showed a similar efficacy compared to the other two available studies of double-boosted PI in experienced HIV-infected children. These two studies assessed the efficacy of the combination of SQV and LPV/r in a cohort of 50 HIV-infected children; the first and the second assessment were performed at week 48 and 96 respectively ,14.At weThe dual-PI regimen in our study was chosen due to the genotypic resistance profile and the ongoing virological failure. The highly resistant genotypic profile was mainly due to an extensive exposure to the N(n)RTI class that has been considered the backbone of antiretroviral regimens. The presence of NAMs has been demonstrated to be highly correlated with reduced susceptibility to all N(n)RTIs , thus a The regimen was well tolerated and none of the patients discontinued the therapy due to side effects. Hepatic parameters remained stable overtime apart from TB, as expected. Lipids increased over 36-42 months of observation without causing therapeutic interruptions.Hyperlipidemia is a well-know complication of ARV therapy in pediatric patients. The European Pediatric Lipodistrophy Group showed that PI use, specifically ritonavir, was associated with dyslipidemia . HyperchOur study is somehow limited by the small number of subjects assessed. Since this observation was carried out in a small number of subjects within the out-patients clinic, we did not decide to conduct a case-control study.e.g. 2 PIs.Moreover, we did not have the possibility to obtain pK data, thus the relationship between drug exposure and toxicity could not be assessed. We understand that TDM studies are of pivotal importance when using drug regimens which include compounds with a potential interaction, In this study, we have proposed a therapeutic strategy that could be considered obsolete and most likely is not being used in most pediatric Infectious Disesase divisions. Nevertheless, in the presence of multiple failures, particularly with N (n)RTIs and NNRTIs, this strategy has overcome the complexity of the resistance pattern. This double-boosted protease regimen with ATV/r and fAVP was effective in suppressing HIV viremia and improving immunological outcome. During the study duration, hypercholesterolmia and hypertriglyceridemia were commonly observed and need close follow-up. The impact on bilirubin level and lipid profile suggests caution in using this regimen in the pediatric population. Thus, this regimen should only be considered as a possible option for a rescue treatment in HIV-infected children and adolescents who failed N(n)RTIs and more so in those with an extensive N(n)RTI resistance.The main contribution of this study lies in the clinical efficacy of a regimen composed by ATV/r and fAVP in effectively maintaining the viral suppression together with a good immunological outcome. An increase in lipid levels was demonstrated in the majority of children included in our study although none of them discontinued HAART. In the future, safer therapeutic strategies, with the same effectiveness, will ameliorate the performance status of our patients. In the meantime, these children will be safely followed to that point.Stefano Rusconi, Vania Giacomet, Chiara Mameli contributed equally to this work.Authors declared no conflict of interest.SR, VG and CM composed the manuscript. FA and OV analized the data. AV, GZ and MG revised the manuscript. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2334/12/179/prepub


Adherence to care and treatment are essential for HIV-infected individuals to benefit from antiretroviral therapy (ART). We sought to quantify the effects on treatment outcomes of missing visits soon after initiating ART.We analyzed data from HIV-infected patients initiating ART at Themba Lethu Clinic, Johannesburg, South Africa, from April 2004 to August 2008. We used log-binomial regression to evaluate the relative risk of missing visits during the first six months of ART on immunological response and virologic suppression. Cox models were used to evaluate the relationship between missed visits and mortality and loss to follow up over 12 months.Of 4476 patients, 65% missed no visits, while 26% missed one visit, 7% missed two and 1.6% missed three or more visits during the first six months on treatment. Patients who missed three or more medical or antiretroviral (ARV) visits had a two-fold increased risk of poor CD4 response by six months, while the risk of failing to achieve virologic suppression by six months increased two- to five-fold among patients who missed two and three or more medical or ARV visits. Adjusted Cox models showed that patients who missed two  and three or more  medical visits had an increased risk of death, while those who missed two ARV  or three or more medical  visits had an increased risk of loss to follow up.Thirty-five percent of patients missed one or more visits in the first six months on treatment, increasing their risk of poorer outcomes. These patients could be targeted for additional adherence counselling to help improve ART outcomes. Expansion of antiretroviral therapy (ART) treatment programmes in resource-limited settings has helped to substantially improve patient outcomes on ART ; howeverIn order to help improve overall treatment outcomes, treatment providers need to focus retention efforts on ART patients who are at increased risk of poor clinical outcomes and becoming lost to follow up. HIV clinics in resource-limited settings continue to struggle to keep patients in care and adhering to treatment in the early stages of ART -8, with Because late presentation for ART is so common , identifThis retrospective cohort study was conducted in the Themba Lethu Clinic in Johannesburg, South Africa. Themba Lethu is one of the largest ART clinics in South Africa, with more than 26,500 patients enrolled in care since April 2004, more than 17,700 of whom have initiated ART. The clinic staff provides care according to South African national Department of Health guidelines . PatientUse of Themba Lethu Clinic data was approved by the Human Research Ethics Committee of the University of the Witwatersrand. Approval for analysis of the data in a de-identified manner was granted by the Institutional Review Board of Boston University.3 and initiated onto standard government first-line ART regimens of stavudine (d4T) or zidovudine (AZT) with lamivudine (3TC) and either efavirenz (EFV) or nevirapine (NVP) between April 2004 and August 2008. We further excluded pregnant women, those who had less than three scheduled medical and three scheduled antiretroviral (ARV) pickup visits during the first six months of ART, and patients with less than 21 months of potential follow-up time (see section on person-time).Our analysis included HIV-positive patients who were eligible for ART based on the 2004 South African national treatment guidelines . Eligibl3 by six months on ART, while failure to achieve virologic suppression was defined as a viral load of ≥400 copies/ml by six months on ART. When CD4 values were missing, they were interpolated by taking the mean of the proceeding and following measures when both were available (6%).We evaluated the relationship between missed medical and ARV pickup visits within the first six months after initiating ART on four outcomes: (1) mortality; (2) loss to follow up; (3) immunologic response; and (4) virologic suppression. Loss to follow up was defined as not having been to the clinic for at least four months. Data on mortality, including for those patients who are lost to follow up, is verified with the South African National Vital Registration Infrastructure Initiative -26. PoorMissing medical and ARV pickup visits during the first six months of initiating first-line ART was the primary independent variable. After ART initiation, ARV visits are scheduled monthly, while medical visits are booked at months one, four and five. Patients prescribed nevirapine are scheduled for two additional medical visits at two weeks and two months to monitor for liver toxicity. Thus, in the first six months on treatment, patients can have up to five ARV visits (the first is excluded as it is the date of ART initiation) and between three and five scheduled medical visits. Unscheduled visits were not counted in the exposure variable since the exposure of interest is missing a scheduled visit, so any non-scheduled visit could not be missed.We determined each participant's missed visit status by evaluating appointment attendance records. Appointment scheduling is managed with the TherapyEdge-HIV™ database, which is used to schedule all visits and to record the actual date on which a scheduled visit was completed. As patients are given two extra days worth of pills each month in case they cannot attend their scheduled ARV visits on time, it is thus likely that in the first six months on treatment, patients who are more than seven days late for a pickup would be without medication. Accordingly, we defined a missed visit as being at least seven days late for that scheduled visit. For our primary analysis, missed visit status over the first six months of initiating ART was categorized as patients who missed 0, 1, 2 and ≥3 visits. We analyzed missed visits stratified by type of visit as either medical or ARV drug pickup.For death and loss to follow up analyses , person-time accrued from nine months after treatment initiation. We chose the period of nine months in order to exclude any deaths or losses to follow up that occurred during exposure (missed visits in the first six months on ART). We excluded an additional three months in order to reduce the impact of reverse causality . Person-time accrued until the earliest of: (1) date of death; (2) date of loss to follow up; (3) date of transfer; or (4) completion of 12 additional months of follow up.3) and failure to achieve virologic suppression (< 400 copies/mL) by six months on ART. Log-binomial models were adjusted for age, sex, WHO stage III/IV condition, body mass index (BMI), baseline haemoglobin and baseline CD4 count  and baseline ART regimen (AZT vs. d4T and NVP vs. EFV). We estimated crude and adjusted hazard ratios of mortality and loss to follow up by missed visit status using Cox proportional hazards models. Hazard models were also adjusted for age, sex, baseline regimen, total number of scheduled visits and CD4 count by the ninth month of ART.Descriptive statistics were performed to look for baseline differences in the distribution of missed visits. Log-binomial regression was used to evaluate predictors associated with missing a visit during the first six months of initiating ART. We also estimated the relative risk of missing a visit in the first six months on both poor CD4 response , were predominately female (62.8%), had a median age of 36.1 years (IQR 31.3-42.2), and were typically prescribed d4T-3TC-EFV (89.4%) as a baseline regimen  attended all scheduled medical and ARV visits, 1178 (26.3%) missed one visit, 312 (7.0%) missed two visits, and 73 (1.6%) missed three or more visits during the first six months on treatment. Patients had a median of four (IQR 3-5) scheduled medical visits and a median of five (IQR 4-5) scheduled ARV pickup visits. There were a total of 115 deaths (2.6%) and 277 patients were lost to follow up (6.2%) during the 12 months of observation. Median follow-up time (which began nine months after ART initiation) among patients who died and missed 0, 1, 2 and ≥3 visits  was 4.5 months (IQR 1.8-9.2), 5.7 months (IQR 2.8-9.8), 3.5 months (IQR 2.0-6.8) and 2.2 months (IQR 0.6-3.5), respectively. Median follow-up time among patients who were lost to follow up and who missed 0, 1, 2 and ≥3 visits  was 6.5 months (IQR 3.8-8.8), 7.1 months (IQR 3.9-9.4), 5.4 months (IQR 3.2-7.6) and 6.3 months (IQR 3.2-8.9), respectively.Using log-binomial regression, we identified no strong independent predictors of missing a medical or ARV visit in the first six months on ART. We found that males, younger patients and those with BMI of <17.5 had a somewhat increased risk of missing a visit in the first six months Table . PatientIn adjusted models, we found that patients who missed three or more medical visits had the highest risk of poor CD4 response by six months on ART   Table . In termPatients who missed 2 and ≥3 medical or ARV visits also had a nearly two- to four-fold increased risk of becoming lost during the follow up period. As with death, missing one medical or ARV visit was not strongly associated with an increased risk of becoming lost to follow up.Recent estimates state that 5.2 million HIV-positive people have initiated ART in resource-limited settings, an increase of 1.2 million since the end of 2008 . InitiatDocumenting the amount of missed visits early on in treatment is critical because of its potential implications for poorer treatment outcomes. Previous studies have shown that patients who miss visits soon after initiating ART are at increased risk of early mortality and loss ,17. In oOur results show that patients who miss visits early on in care are less likely to achieve virologic suppression after six months of ART compared with patients who attended all scheduled visits. As adherence is critical to the success of their treatment -13, missAlternatively, some of the relationship between missed visits and poor treatment outcomes may result from missed opportunities to detect and treat opportunistic infections. These conditions may still be diagnosed and treated at a later time, but opportunistic infections after initiation of ART are associated with poor immune recovery ,36. A loThe reasons that patients miss visits early on in care are likely to be multi-factorial and complex. While we found no strong predictors of missing medical visits, our findings show that male gender, younger age, low baseline BMI, AZT-based versus d4T- and NVP-based versus EFV-based regimens are predictive of missing visits early on in treatment. Data from industrialized countries has shown that younger patients ,28,29,38Our study is one of the first to report on the specific relationship between missed visits in the first six months of ART and clinical HIV treatment outcomes in a resource-limited setting and longer term outcomes. Still, our findings should be considered alongside the study's limitations. First, because our study reports data from a single government HIV clinic, our results may not be generalizable to the overall population. Second, data on specific reasons for missed visits is not collected and therefore we could not determine what the major barriers are to maintaining a consistent schedule of care and treatment.Finally, we did not have actual ARV drug dispensing data, and while missing scheduled drug collection visits is likely to mean that a patient is without medication, we cannot rule out the possibility that the individuals were dispensed additional months of drug supplies. However, in this study we considered only visits in the first six months after treatment initiation, a period when the pharmacy is least likely to dispense several months of medication. This is also a period when a patient needs to see a clinician in order to have his or her ARV prescription renewed, and so we anticipate that this occurred infrequently.Our results suggest that patients who miss multiple visits in the earliest period of treatment but remain in care are at increased risk of poorer ART outcomes. Currently, a substantial proportion of funding and manpower in resource-limited settings is focused on reducing mortality and loss to follow up among those started on ART. Our findings suggest that targeting those who miss visits early on in treatment could have benefits in terms of longer term mortality reductions. Future studies need to focus on identifying the barriers to adhering to a visit schedule so that interventions and support services can be directed at those at risk.MM and IS are affiliated to Right to Care, which provided some of the funding for the current research and also supports the provision of treatment for the patients in the study. All authors  have no other competing interests to declare.AB contributed to the study concept and design and to analysis and interpretation of the data, in addition to drafting and critical revision of the manuscript for important intellectual content. MM contributed to the analysis and interpretation of the data and critical revision of the manuscript. IS contributed to the acquisition of data, provided critical revision of the manuscript and obtained funding. MF contributed to the study concept and design and to analysis and interpretation of the data, in addition to drafting and critical revision of the manuscript for important intellectual content. All authors have read and approved the final manuscript.


Despite the availability of antiretroviral therapy (ART), suboptimal treatment outcomes have been observed among HIV-seropositive illicit drug users. As there is an urgent need to improve responses to antiretroviral therapy among this population, we undertook this study to evaluate the role of physician experience on rates of plasma HIV-1 RNA suppression following initiation of ART.Using data from a community-recruited cohort of HIV-positive illicit drug users, we used Cox proportional hazards regression to model the time to plasma viral HIV RNA < 500 copies/mL among antiretroviral-naïve subjects initiating ART. Physician experience was defined as a continuous variable measured per 100 HIV-infected patients previously enrolled in the province-wide HIV treatment registry by that physician at the time a patient was enrolled.Between May 1996 and December 2008, 267 individuals initiated ART among whom 227 (85%) achieved a plasma HIV RNA < 500 copies/mL during the study period. In a multivariate analysis, greater physician experience was independently associated with higher rates of plasma HIV RNA suppression  after adjustment for adherence to ART. Other factors associated with viral suppression included engagement in methadone maintenance therapy , ≥ 95% adherence to ART , baseline CD4 count  and baseline plasma HIV-1 RNA .In this setting of universal HIV/AIDS care, illicit drug users with more experienced physicians exhibited faster rates of plasma viral load suppression. These findings argue for specialized services to help optimize HIV treatment outcomes among this population. With over 30 million cases distributed worldwide, the HIV/AIDS pandemic is a global public health emergency . In areaDespite the availability of effective treatment, several individual, social and structural factors have posed barriers to effective ART outcomes among illicit drug users. While access and adherence to the prescribed drug regimen is recognized as the most important determinant of treatment success , other fData for this study were obtained from the AIDS Care Cohort to evaluate Exposure to Survival Services (ACCESS), an ongoing observational prospective cohort of HIV-positive illicit drug users ,9. FolloData on HIV clinical monitoring and drug-using behaviour were augmented with information on HIV care and treatment outcomes from the province-wide centralized ART dispensary and HIV clinical monitoring laboratory at the British Columbia Centre for Excellence in HIV/AIDS (BC-CfE) ,9. As a This study included all ACCESS participants who were ART-naïve at recruitment, initiated treatment during the study period, and had at least one clinical test measuring CD4 cell count and plasma HIV-1 RNA level during the first 12 months on ART. Our main outcome, time to plasma HIV-1 RNA suppression, was operationalized as the date of the first observation with a plasma viral HIV-1 RNA < 500 copies/mL.The primary explanatory variable of interest was physician experience, defined as the number of patients that the participant's prescribing physician had previously enrolled in the province-wide HIV treatment registry. This was considered as a continuous variable and, since physicians could become more experienced over time, was fixed for each participant at the time they initiated ART. In British Columbia, antiretroviral prescribing physicians could be located anywhere in the province and not at only one institution. Patients selected antiretroviral-prescribing primary care or specialist physicians in a non-random manner through self-selection or referral from other physicians. Physician experience was fixed as a baseline characteristic at the time the patient initiated ART as we believe that this was the best way to estimate physician experience. We recognized certain behaviours (e.g. drug use activity) could confound this association so these measures were treated as time-updated variables based on each participant's semi-annual follow up visit.10 increase) and the year of ART initiation (per more recent year). Illicit drug use measures included daily cocaine use (yes vs. no) and daily heroin use (yes vs. no) and referred to the six-month period prior to the interview. Patients were prescribed antiretroviral therapy consistent with therapeutic guidelines which, beginning in 1996 for all patients, recommended triple combination therapy. While the drugs used over the study period changed markedly over time, consistent with previous analyses, we adjusted for whether protease inhibitors were used in the initial regimen or not as a strategy to adjust for confounding that could occur as a result of regimen type. Given the large number of nucleosides used in the backbone of the ART regimen during the study period, we elected to not adjust for this in the analysis.Several variables that could influence the association between physician experience and plasma HIV-1 RNA suppression were also assessed, including age, gender , Aboriginal ancestry (yes vs. no) and Downtown Eastside residence (yes vs. no). Clinical variables included ART adherence (≥ 95% vs. < 95% in first year of treatment), current enrolment in methadone maintenance therapy (no vs. yes), protease inhibitor as part of the first ART regimen (yes vs. no), CD4 cell count at baseline (per 100 cells), plasma HIV-1 RNA level at baseline  women and 105 (39.3%) individuals who reported being of Aboriginal ancestry. The baseline characteristics of the study sample along with the median physician experience of different groups are presented in Table p-value < 0.001), as were older patient age , engagement in methadone maintenance therapy , ≥ 95% ART adherence , CD4 cell count , baseline plasma viral load , and the year of ART initiation During the study period, 227 participants (85.0%) achieved at least one observation of plasma HIV-1 RNA < 500 copies/mL, reflecting an incidence density of 65.2 per 100 person years (95% CI: 57.0-74.2). Table p-value = 0.031). Other factors that were associated with time to plasma HIV-1 RNA suppression in this multivariate analysis included methadone maintenance therapy , baseline CD4 count , baseline log10 HIV-1 RNA  and adherence to ART . When we fit a multivariate model including the three main associations shown in Table p-value = 0.003) after adjustment for Downtown Eastside residence, methadone maintenance therapy and daily cocaine injection.Table In a sub-analysis, we explored the impact of physician experience on time to viral load suppression when physician experience was defined as a categorical variable using tertiles. In the first tertile, physicians had treated less than 22 patients previously and the median time to plasma HIV-1 RNA < 500 copies/mL was 9.5 months (95% CI: 4.4-14.5). In the middle tertile, physicians had treated 22-81 patients previously and the median time to suppression was 6.2 months (95% CI: 3.0-13.4). In the most experienced tertile, physicians had treated greater than 81 patients previously and the median time to plasma HIV RNA suppression < 500 copies/mL was 3.0 months (95% CI: 2.3-4.7).In this study, we found that greater physician experience was positively associated with the time to viral load suppression among drug users initiating ART. This association persisted in a multivariate model after adjusting for CD4 cell count, baseline plasma HIV-1 RNA level, methadone use, and adherence following ART initiation.Our finding that physician experience was associated with higher rates of plasma HIV-1 RNA suppression is noteworthy. Prior to the advent of combination antiretroviral therapy, a US study found that higher physician experience was associated with longer patient survival . SimilarAlthough our measure of adherence has been previously validated to predict plasma HIV RNA responses , CD4 couOur study has some limitations to be noted. First, our cohort was not randomly selected and therefore may not be representative of all HIV-infected drug users in Vancouver or elsewhere. Second, although physician experience was defined based on an HIV treatment registry, for other variables we relied on measures of self-report, and therefore response bias could have undermined our ascertainment of sensitive behaviours, including illicit drug use and addiction treatment use. Finally, our analyses only includes follow-up information until 2008 and data collected when the lower limit of detection was 500 copies/mL. Future studies should continue to assess whether physician experience is associated with outcomes from HIV treatment, especially as modern HAART regimens are increasingly simple to take.In summary, we examined plasma HIV RNA responses after the initiation of ART among a long-running community-recruited prospective cohort of HIV-positive illicit drug users and observed that lower HIV-related experience of physicians was independently associated with lower rates of plasma HIV RNA suppression. These findings argue for more specialized treatment approaches to address the HIV treatment needs of this challenging population.Julio Montaner is supported by the Ministry of Health Services and the Ministry of Healthy Living and Sport, from the Province of British Columbia; through a Knowledge Translation Award from the Canadian Institutes of Health Research (CIHR); and through an Avant-Garde Award (No. 1DP1DA026182-01) from the National Institute on Drug Abuse at the US National Institutes of Health. He has also received funding from Merck, Gilead and ViiV to support research into Treatment as Prevention.SS, M-JM, TK, EW and JM conducted the study. EW, TK, AI and SS conceived the analysis. RZ performed all statistical calculations. M-JM, AI and SS drafted the manuscript and integrated revisions from all other co-authors. All authors reviewed and approved the final draft.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2334/12/22/prepub


While HAART allows for the reconstitution of immune functions in most treated HIV patients, failure to achieve a significant increase in circulating CD4+ T cells despite undetectable viremia occurs.A retrospective study was conducted to evaluate the treatment outcome in a subgroup of 232 patients who after 3.1 years of treatment had not achieved desirable immune reconstitution despite a good virological response to HAART.After a further 3.5±2.7 years of HAART, 41 (17.7%) patients achieved immune reconstitution (681.4±172.7 CD4 cells/µL), while 191 (82.3%) patients did not (306.6±109.16 cells/µL); the difference in the achieved CD4 counts between these subgroups was significant (P<0.01). One patient experienced treatment failure. Eleven patients died to the end of follow-up, of which ten with a continuously dissociated response. Factors associated with immune recovery included, usage of PIs and of drugs from all three classes , and a rise in CD4 count to over 200 cell/µL after the first 3.1 years of treatment . Achievement of a rise in CD4 count to over 200 cell/µL after the first 3.1 years of treatment, and usage of all three drug classes were independent predictor of immune reconstitution in the following period.If patients on HAART reach CD4 cell counts of above 200 cells/µL in the first three years, immune recovery is possible after at least six years of treatment, particularly if treated with drugs from all three classes.


Analysis of potentially different impact of Lopinavir/Ritonavir (LPV/r) on non-B subtypes is confounded by dissimilarities in the conditions existing in different countries. We retrospectively compared its impact on populations infected with subtypes B and C in Israel, where patients infected with different subtypes receive the same treatment.Clinical and demographic data were reported by physicians. Resistance was tested after treatment failure. Statistical analyses were conducted using SPSS.607 LPV/r treated patients  were included. 139 had HIV subtype B, 391 C, and 77 other subtypes. At study end 429 (71%) were receiving LPV/r. No significant differences in PI treatment history and in median viral-load (VL) at treatment initiation and termination existed between subtypes. MSM discontinued LPV/r more often than others even when the virologic outcome was good (p = 0.001). VL was below detection level in 81% of patients for whom LPV/r was first PI and in 67% when it was second (P = 0.001). Median VL decrease from baseline was 1.9±0.1 logs and was not significantly associated with subtype. Median CD4 increase was: 162 and 92cells/µl, respectively, for patients receiving LPV/r as first and second PI (P = 0.001), and 175 and 98, respectively, for subtypes B and C (P<0.001). Only 52 (22%) of 237 patients genotyped while under LPV/r were fully resistant to the drug; 12(5%) were partially resistant. In48%, population sequencing did not reveal resistance to any drug notwithstanding the virologic failure. No difference was found in the rates of resistance development between B and C (p = 0.16).Treatment with LPV/r appeared efficient and tolerable in both subtypes, B and C, but CD4 recovery was significantly better in virologically suppressed subtype-B patients. In both subtypes, LPV/r was more beneficial when given as first PI. Mostly, reasons other than resistance development caused discontinuation of treatment. WHO latest guidelines et al., compared LPV/r treatment outcome of B and C patients where both groups were treated under equal conditions in a developed country.Mutations conferring reduced susceptibility of the virus to LPV/r are well documented  versus B, following the introduction of LPV/r.In Israel, subtypes B, C and A/AE, are prevalent Because of the heterogeneity of the patient population and the retrospective nature of the study, a subtype-based comparison of treatment outcomes, particularly in terms of viral-load suppression and CD4 gains, might be confounded by several other factors that potentially influence the results. The median value of several parameters, including age, follow-up period, and time on LPV/r were similar in subtype B and C, and thus not confounding. The outcome of treatment significantly depended, in addition to subtype, on whether the patients had previous PI treatment experience or not, but here too, the proportion of patients for whom LPV/r was first PI was similar in the three subtype groups.Six of the seven AIDS-treating centers in Israel participated in this retrospective study. All patients (607) who received LPV/r for at least 3 months before August 2007 with documented treatment dates and known HIV RNA viral load (VL) levels and CD4+ T-cell counts (CD4 Counts) at starting and stopping/end dates were included. Patients were stratified according to subtype, gender, PI experience prior to LPV/r, and to whether they were on LPV/r treatment or had stopped it during the study. As subtype B patients were mainly MSM, this group of patients was also compared to subtype-C males. The study was approved by the local Ethics committees of the different institutions.http://hivdb.stanford.edu/pages/webservices, version 6.3.1  Sierra Webservice  Monitor test, version 1.5 ; the Cobas AmpliPrep/Cobas TaqMan HIV-1 test ; and real-time nucleic acid sequence-based amplification (NASBA) HIV-1 assay . The assays were performed according to the manufacturer's instructions. The detection level of the least sensitive test was 400 copies/ml and therefore results below this value were considered lower than detection level (LDL). The genotyping tests were all performed at the National HIV Reference Laboratory (NHRL). Viral RNA was isolated from patient blood plasma using the BioMerieux automatic extractor (Easy MAG) according to manufacturer's instructions. The Protease gene (codons 4–99) and RT gene (codons 38–247) were sequenced using Siemens' True-Gene™ kit as described before p<0.05.Chi-square test and Fisher's two-tailed exact test were used for analysis of discrete data  and Bonferroni's correction was applied to multiple comparisons of mutations frequency. T-test and one way Anova test were used in comparing continuous clinical data . A square root transformation was applied to the CD4 counts to approach a normal distribution for this variable and the t-test and Anova test were performed on the transformed values.Logistic regression analysis was implemented to predict viral load below detection level at the end of the study. All statistical analyses were performed using SPSS® (version 21.0). Data are presented as median [range] unless otherwise stated. The retrospective analysis of clinical and laboratory data, which were obtained from the medical charts of HIV-1 patients attending the different Medical Centers, was approved by the respective ethical committees. Specifically, permission was granted by the Kaplan Ethical Committee to analyze such data without the need of a signed informed consent by the patients. The samples obtained at the Sheba Medical Center that were used in this study belonged to patients who had signed an informed consent agreeing to participate in a range of studies.AY529598, KC184165, KC184166, KC184169, KC184185, KC184186, KC184219, KC184325, KC184345, KC184354, KC184392, KC213492, KC213521, KC213541, KC213556, KC213581, KC21358, KC213589, KC213607, KC213609, KC213612, KC213656, KC213662, KC213664, KC213665, KC213666, KC213683, KC213684 and KF134929 - KF135178.p<0.001). Other parameters, including median age (38.1±0.5 years), median time from diagnosis to starting LPV/r treatment (5.2±0.2 years), median time on LPV/r (23.7±0.8 months) and the fraction of patients for whom LPV/r was first PI were similar in the B and C groups (p = 0.1 to p = 1) but the median time from diagnosis to LPV/r treatment initiation for the non-BC group was shorter  and a larger fraction of these patients received LPV/r as first PI (p = 0.006).Six hundred and seven patients  from six AIDS clinics in northern, central and southern Israel were included. A summary of patient characteristics can be found in Median LPV/r treatment time was 23.7±0.8 months (range 3 to 95). For 305 patients (50.2%) LPV/r was the first administrated PI; 140 (46%) of those were naive to prior cART. The most common backbone regimens were ZDV+3TC (65.4%), 3TC plus either ddI or d4T or ABC (13.4%), and ddI plus d4T (9.8%). At the end of the study 429 patients (71%) were receiving LPV/r (“ongoing” group) and 177 had discontinued; of those, 13 died. There was no statistical difference between the subtype B and C or non-BC populations in number of former PIs (zero or more), and number of past regimens, but for a significantly larger fraction of non-BC patients (74%) LPV/r was first PI (p = 0.006). Among patients who were treated with other PIs about 30% received nelfinavir, 30% received indinavir, 30% received both and 10% were treated with other PIs (mainly saquinavir). The distribution of treatment regimens was similar in all subtypes groups.The various side effects reported by clinicians are shown in Viral load and CD4 results are described in p = 0.08 and 0.03, respectively). Additional ΔCD4 differences seen between MSM and C-infected males and between males and females reflect the predominance of MSM among B-patients.After stratification in terms of different groups as described we found, in particular, that the fraction of patients ending up below detection level did not differ significantly between subtypes B and C, despite the fact that treatment was interrupted significantly more frequently by B-group patients than by C (P = 0.001), and treatment interruption was strongly associated with a poorer virologic outcome p<0.001; . TreatmeOverall, the best outcome of LPV/r was observed when it was first PI . We also stratified patients by baseline VL and CD4 counts and compared clinical outcomes of the different groups. Patients were divided into three groups according to VL, and each VL group was subdivided according to the CD4 counts, creating altogether nine groups. The VL ranges were >100,000, 10,000–100,000 and <10,000 cp/ml, and for CD4 counts <200, 200–500 and >500 cells/µl . No signThree hundred samples from 237 patients on LPV/r with VL ≥1,000 copies/ml were genotyped. The latest sample from each patient while on LPV/r was used in the analysis. Mutations and common polymorphisms are depicted in et al.et al. (p = 0.5)p = 0.1–0.7) although significantly more subtype B samples were sensitive to all drugs  were fully resistant to LPV/r, 11 (5%) were partially resistant and 174 (73%) were sensitive, half of those to all available PIs. Of the 63 samples with resistance to LPV/r, 1 (2%), 23 (36%) and 10 (16%) samples, respectively, were fully sensitive to the newer PIs atazanavir/r, darunavir/r or tipranavir/r. The major factor influencing accumulation of mutations was the history of PI usage. When LPV/r was the first PI administrated resistance was found in 8.7% of samples, similar to Barber LPV/r was incorporated into cART regimens in Israel in 2001. Here we retrospectively evaluated our experience with this drug gained over the following several years, involving 607 patients treated with LPV/r for an average of two years (range 3 to 95 months). We compared the rates of achieving suppression of viral replication (VL<400 copies/ml) and median CD4 increases, as well as drug resistance pathways, in those infected with different HIV-1 subtypes, mainly C and B.et al. comparing CD4 recovery of subtype A- and subtype B-infected patients We did not find significant differences between the major subtypes, B and C, in the rate of viral suppression, but there were such differences in the median increase in CD4-cell count, in particular in those who achieved undetectable viral load  & 4. Sevp = 0.02), L63P and A71V (p = 0.04–0.001). T74S/A appeared in 28% of C samples and none of B (p<0.001). Almost all subtype-C patients had M36I and L89M which are in the consensus wild-type sequence of subtype C p = 0.2–1; The different subtypes appear to have selected somewhat different pathways to replicate in the presence of the drug . SamplesWe stratified samples according to viral load and presence of resistance mutations. As reported briefly earlier, we found that both groups, those who failed with mutations and those who failed without them, could each be divided into two subgroups according to their viral-load levels . Thus, tp = 0.4–0.8). It appears that the degree of success of the treatment in terms of viral load decrease and CD4 count increase as well as the actual presence or absence of side effects are not the only factors that influence the decision of patients to continue or discontinue treatment. For example, concerns regarding the risk of such side effects, especially in a more “educated” group, may be a factor that needs to be considered.Similar proportions of patients with different subtypes reported side effects  and between I54V and V82A (p = 0.09). Consistent with these trends, the triplet M46I, I54V and V82A was very significantly over-represented (p = 0.007). On the other hand, L24I and L33F tend to appear mostly in conjunction with multiple other resistance mutations, as they do not confer strong resistance to LPV/r on their own but strengthen the effect of other mutations or the interaction between them once the latter have been selected.With 157 C-subtype patients genotyped upon failure of LPV/r-including regimens, our study is now second only to van Zyl The retrospective nature of our study is a major limitation inherent to its design. We have attempted to provide subgroup analysis where possible. Another limitation is our inability to discern the effects of baseline structural differences between the subtypes on LPV/r impact from the effects of differences in adherence to, and persistence in, drug taking schedules prescribed by physicians. This difficulty exists because, although all patients are similarly treated by the same physicians, C patients largely belong to a group of immigrants from Ethiopia so that social and cultural factors may have affected adherence in ways that are hard to evaluate. Six out of seven AIDS treating centers in Israel participated in the study. Differences between the centers in keeping the patients on the drugs cannot be excluded, and since the ratio between B and C patients in each center varied, a bias could result. We believe, however, that such differences are secondary to the potential difference in adherence between subtype-B and subtype-C patients as groups. Assessing to what degree low-level residual viral replication contributed to differences in ΔCD4 was limited by the detection level of 400 copies/ml that we had to impose in the analysis as more sensitive tests were performed for only a fraction of the study population. Finally, as mentioned above, drug resistance might have escaped detection in some patients due to the limitations of the method of population sequencing. Ultra-deep sequencing would be required to assess this possibility.In summary, our data highlight the long-term efficacy and safety of lopinavir/ritonavir among patients with both subtypes B and C. Despite the great variety in these populations in ethnic, educational and other socio-economic parameters, the differences in clinical outcome between the subtype groups appeared to have limited clinical relevance. These findings strongly support the expanding use of LPV/r in resource limited settings with high prevalence of subtype C infection.


Despite the increasing use of antiretroviral treatment (ART) recent data on frequency and pattern of drug resistance mutations in Ethiopia is not available. Furthermore with increasing mobility of people HIV-1 subtypes other than the predominant subtype C may likely be introduced from the neighbouring countries. This study was aimed to determine the molecular characterization and pre-antiretroviral treatment resistance mutations among HIV-1 chronically infected ART naïve patients after the roll out of ART in Ethiopia.pol gene of the HIV-1 genome (N = 160) were amplified and sequenced using an in-house assay. Genotypic drug resistance was defined as the presence of one or more resistance-related mutations as specified by the consensus mutation of Stanford University HIVDB and the International Antiviral Society (IAS) mutation lists.Viral RNA was determined in 160 baseline plasma samples. The entire PR and the first 335 codons (76%) of the RT regions of the A predominance of HIV-1 subtype C (98.7%) was observed. The level of drug resistance is found to be 5.6% and 13.1% according to the Stanford University HIVDB drug resistance interpretation algorithms and the International Antiviral Society mutation lists, respectively. Mutations conferring simultaneous resistance to NRTIs and NNRTIs were not detected and no major PR mutation was found. However, a high rate of polymorphic changes both in PR and RT regions were observed. Moreover, twenty four (15%) monophyletic transmission clusters with bootstrap value of 99% were found.Strong evidence for consistent HIV-1C clade homogeneity and low influx of other variant into the country was found. The level of drug resistance observed in chronically infected treatment naïve patients which exceeds the WHO estimates suggests the need for incorporation of HIV-1 drug resistance testing prior to ART initiation. The occurrence of monophyletic transmission clusters affecting (24/160) individuals indicates their potential risk related practice. Thus, an intensified public health intervention program and monitoring of HIV drug resistance testing appears indispensible. In response to the global HIV-1 epidemic and to achieve universal access for the treatment of HIV-1 infected individuals in resource limited countries, antiretroviral treatment (ART) has been scaled up in the past decade  and theiIn Ethiopia, TDR before the introduction of ART (2002) and one year after its introduction (2005) was reported well below 3% ,8. Neverst with addition of protease inhibitors (PIs) in 2nd line, understanding of drug-resistance patterns in pol gene among non-B subtypes may help to optimize the selection of first-line regimens and limit the acquisition of cross-resistance. The objectives of the current study is to determine the HIV-1 genetic diversity and to identify the pattern of antiretroviral drug resistance mutations in pol gene of HIV-1 isolated from chronically infected treatment naïve Ethiopian patients.HIV pol gene encodes the reverse-transcriptase (RT), protease (PR) and integrase enzymes that are the major targets of antiretroviral therapy . As drugHIV-1 chronically infected treatment naïve patients (N = 160) with advanced diseases   above 18+ T cell count was measured using the FACSCount flow cytometer  following the manufacturer’s protocol. Plasma was separated by centrifugation and stored at −40°C.Five ml venous blood was collected in vacutainer tubes containing ethylene diamine tetraacetic acid (EDTA). Baseline CD4RNA extraction was done with the Abbott m2000sp automated sample preparation system using mSample preparation system RNA kit. Plasma viral load was determined with Abbott m2000rt Quantitative RealTime HIV-1 assay  with a lower detection limit of 40 copies/ml.The entire PR and the first 335 codons (76%) of the RT regions of the pol gene of the HIV-1 genome of 160 patients were amplified with an in-house protocol as described before . Brieflypol region. All primer positions are matched to HIV-1HXB2 (GenBank accession number K03455). Both forward and reverse overlapping sequences were manually edited with the Geneious software version 5.4 [Purified PCR products were subjected to direct sequencing of both sense and antisense strands using Big Dye Terminator Cycle Sequencing Ready Reaction kit . For each sample, six separate sequencing reactions were done using the two inner PCR primers (HIVpcrFor2 and HIVpcrRev2) and four additional internal primers: HIVseq1, HIVseq2, HIVseq3 and HIVseq4  and SIV) obtained from HIV Sequence Database at Los Alamos (http://www.hiv.lanl.gov) accessed on September 24, 2013. Phylogenetic inferences were performed by the neighbour-joining method with 1,000 bootstrap replicates under Kimura’s two-parameter correction using MEGA 5. The evolutionary distances were computed using the Maximum Composite Likelihood method and are in the units of the number of base substitutions per site [per site .http://hivdb.stanford.edu) accessed on September 24, 2013 and the latest definition of the International Antiviral Society (IAS) mutation lists [The existing drug resistance interpretation algorithms use lists of amino acid mutations that are associated with drug resistance, derived largely from research of HIV-1B infected individuals -20. Withon lists .+ T cell count and HIV RNA level were included in the statistical analysis using SPSS statistical software . Student t test was used and a p-value of < 0.05 was considered statistically significant.Age, sex, WHO clinical stage, CD4Ethical clearance and written informed consent from the University of Gondar Ethical Review Committee (RPO/55/291/00) and each study subjects was obtained, respectively.10 HIV RNA level of the patients was significantly lower (4.46log10 versus 5.51log10) in patients with higher (>200 cells/mm3) compared with lower (<200 cells/mm3) CD4+ T cell strata (P < 0.001).One hundred and sixty samples  were amplified and sequenced for HIV-1 subtyping and genotypic drug resistance analysis. The mean ± SD age of the subjects was 32.3 ± 8.5 years (range 18–66 years). The socio-demographic, clinical, immunological and virological characteristics of the patients are summarized in Table All sequences clustered with HIV-1C in HIV pol genome with a bootstrap value of 83%  and 5.6% (9/160), respectively were found to have one or more mutation conferring resistance to RTIs  according to the Stanford University drug resistance database and IAS mutation lists (Table http://www.hivfrenchresistance.org).As indicated in Table In both algorithms, four patients (2.6%) had NRTI resistance-associated mutations. K65R was observed in one patient. Mutations at position 210 from L to W and at 219 from K to E were observed in two and one patient, respectively. A revertant mutation T215A was also observed in 3 (1.9%) of the patients. Moreover atypical mutations at known subtype B drug resistance positions Table  conferri+ T cell counts  of patients with and without drug resistance mutations  of HIV-1 subtype C in Ethiopia similar with previous reports % of HIV- and provIn conclusion, there is strong evidence for consistent HIV-1C clade homogeneity and low influx of other HIV-1 subtypes to Ethiopia for the past 4 decades. The observed level of drug resistance which exceeds the WHO’s estimates suggests the need for routine HIV-1 genotypic drug resistance testing for treatment naïve patients and supports the hypothesis that scaling-up of ART in resource limited settings drive the development of drug resistance. The absence of major drug resistance mutations in PR gene suggest for scaling-up of PIs in Ethiopia. The occurrence of monophyletic transmission clusters in 15% of individuals indicate their potential of risk related practice and thus, an intensified public health intervention program targeting such individuals is warranted.Nucleotide sequences are deposited in National Centre for Biotechnology Information (NCBI), USA GenBank (Accession Number: KF026059-KF026220).We declare no competing interest.AM: Conception and design of the study, acquisition, analysis and interpretation of data, drafting the article and final approval of the version to be submitted; TL: Acquisition, analysis and interpretation of data, revising the draft article and final approval of the version to be submitted; UGL: Conception and design of the study, analysis and interpretation of data, revising the draft article and final approval of the version to be submitted; MM: Conception and design of the study, analysis and interpretation of data, revising the draft article and final approval of the version to be submitted.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2334/14/158/prepub