The global burden of Hepatitis B virus (HBV) and HIV co-infection is enormous. The risk of developing cirrhosis and hepatocellular cancer is associated with HBV DNA levels. The main objective of the study was to determine proportion of Hepatitis B viremia in ART-naïve and ART-experienced co-infected Ghanaian patients and factors associated with HBV viremia after at least 36 weeks of lamivudine with or without tenofovir containing ART.Hepatitis B and HIV co-infected patients who were ART-naïve or had received at least 9 months of lamivudine-containing ART were enrolled in a cross-sectional study at Korle-Bu Teaching Hospital. Demographic and clinical data were collected and samples obtained for Hepatitis B serology, liver function tests and HBV DNA. Factors associated with viremia were determined using univariate and multivariate logistic regression analysis.Of 3108 HIV-infected patients screened, 257 (8.3 %) were HBsAg-positive, of which 235 enrolled. Overall, 152 (64.7 %) were ART-experienced and 83 (35.3 %) were ART-naïve. Eighty-nine-percent of ART-naïve and 42.1 % of ART-experienced patients had HBV DNA > 20 IU/mL. In multivariate analysis of all patients, being ART-naïve  and elevated ALT  were associated with Hepatitis B viremia. In treatment experienced patients, elevated ALT (OR 4.8 CI 2.0 – 12.1) and male sex  were associated with Hepatitis B viremia.Majority of ART-naïve (89 %) and 42 % of ART-experienced patients had detectable hepatitis B viremia > 20 IU/mL. An abnormal serum ALT was significantly associated with hepatitis B viremia in HBV and HIV co-infected patients irrespective of treatment status. Baseline and on-treatment ALT may be a useful non-invasive predictor of Hepatitis B viremia in resource-constrained countries in sub-Saharan Africa where infection is endemic and viral load tests are not widely available. Globally, it is estimated that there are 240 million persons chronically infected with HBV infection, with a majority of them living in low- and middle-income countries . The preThe risk of developing cirrhosis, hepatic decompensation and HCC is associated with HBV DNA levels , 7. HIV In Ghana, the reported prevalence of HBV ranges from 12 to 15 % among blood donors . At the Korle-Bu Teaching Hospital has 2500 beds and is the main tertiary referral centre in Accra serving the majority of the southern half of Ghana. The Fevers Unit is the principal referral centre for infectious diseases in Southern Ghana with approximately 15,000 HIV patients under regular follow-up in the out-patient clinic and over 7000 patients on regular anti-retroviral therapy. It has an in-patient department staffed with doctors, nurses, counselors, para-clinical and laboratory staff which offers service to a wide range of patients with infectious diseases including HIV.This study used a cross-sectional design to recruit HBV-HIV co-infected patients attending the Fevers (Infectious Diseases) Unit, Korle-bu, Accra, Ghana between 2012 and 2014. Adult HIV patients attending the Fevers (Infectious Diseases) Unit during the study period (2012–2014) were screened for HBsAg using the Core HBsAg rapid test  following informed consent. HIV and HBsAg-positive study participants were enrolled into two groups. The treatment-naive group (t0) were HBV-HIV co-infected patients who had not initiated therapy. The treatment-experienced group (t1) were HBV-HIV co-infected patients who had been on at least 9 months of 3TC containing anti-retroviral therapy. Patients were excluded if they declined to sign a consent form for participation. Demographic and clinical data were collected from enrolled patients using standardized data collection form. Medical records were also reviewed where additional clinical and laboratory data including CD4 cell count extracted as patients did not know their most recent CD4 count. Adherence status was determined using the validated Simplified Medication Adherence Questionnaire (SMAQ) . All extThe Institutional Review Board (IRB) of the University of Ghana Medical School, Accra, Ghana and Lifespan Hospitals, Providence, Rhode Island reviewed and approved the study.Enrolled patients had blood drawn into an EDTA and serum separating tube (SST). Blood in EDTA was transported to the Virology Laboratory, Korle-Bu, on ice, centrifuged within 4 h of collection using EBA 20 table-top centrifuge , and plasma frozen at −80 in ~ 600 ul aliquots. HBsAg positivity was confirmed serologically using a third generation ELISA Surase B 96 (TMB) . Plasma was also tested for Hepatitis B e-antigen (HBeAg), Hepatitis B e-antibody (anti-HBe) using EASE BN-96 (TMB) and Hepatitis B-core antibody (anti-HBc) using ANTICORASE MB-96 (TMB) from General Biologicals, Hsinchu, Taiwan. Blood in SST was tested for liver function. Plasma HBV DNA was determined using fully automated COBAS® TaqMan® Analyzer . The lower limit of detection of the Roche TaqMan assay was 20 IU mL-1 and the linear range 20 – 170,000,000 IU/mL.t-test for continuous variables or Chi-square test for categorical variables. Logistic regression with variable selection by stepwise was used to find the joint effect of the participants  on HBV viremia using all the data and also using treatment-specific data, and the findings were confirmed by the Smoothly Clipped Absolute Deviation (SCAD) method [P value < 0.05 was considered significant.Statistical analyses were performed using Software SAS 9.3 . Bivariate analyses of association between participant factors and treatment status or HBV viremia were assessed by Student’s ) method , 17. For10IU/mL was 8.8 (IQR: 4.8–17.6) and 2.9 (IQR: 2.1–6.8) respectively. The mean (SD) duration of ART in treatment-experienced patients was 4.2 (2.7) years. Treatment-experienced and naïve patients were similar except that treatment-experienced patients were older, had higher mean weight, higher mean BMI and higher mean CD4 count Table During the study period, 3108 HIV-infected patients were screened for HBV with HBsAg, of which 257 patients (8.3 %) were HBsAg positive. Two hundred and thirty-five (235) patients were recruited into the study, with 22 patients either declining consent or had incomplete data. The baseline characteristics of all patients and by ART status are shown in Table Of the 152 treatment-experienced patients, 100 (65.8 %) were on ART with 3TC (3TC), while 52 had received 3TC + TDF containing ART, for at least 9 months prior to sampling (3TC + TDF). Of the (3TC + TDF) group, 37 (71.2 %) were on combination therapy from the onset of treatment while 15 (28.8 %) had been on another 3TC containing regimen before changing to 3TC + TDF combination therapy.In the 3TC-only group, the median duration of treatment was 4.85 years (IQR: 3.0–7.1). In the 3TC + TDF group, median duration of treatment of patients on 3TC and TDF from the onset was 1.23 years (IQR: 1.1–1.6) while 4.87 years (IQR: 3.8–5.9) in patients on another 3TC regimen before changing to 3TC + TDF combination therapy.Overall, 74 (89.2 %) of the 83 ART-naïve and 64 42.1 %) of the 152 ART-experienced patients had HBV DNA > 20 IU/mL (Table  % of theP = 0.019). An elevated serum AST and ALT above ULN were also more likely to be associated with detectable HBV during ART (P = 0.0004) and (P = 0.0001), respectively. In treated patients, being HBeAg-positive and HBeAb-negative was significantly associated with detectable HBV DNA (P < 0.0001). Treatment regimens (3TC) and (3TC + TDF) had similar likelihood of hepatitis B DNA suppression, with undetectable Hepatitis B virus in 42.0 and 42.3 % respectively (P = 0.97). Of the 83 treatment-naïve patients, only 9 (10.8 %) had HBV DNA < 20 IU/mL.Bivariate analysis of factors associated with viremia (HBV DNA > 20 IU/mL) in the treatment-naïve and experienced patients are shown in Tables In multivariate analysis of all patients, naïve patients were more likely than the treatment-experienced group to have detectable HBV DNA . Additionally, elevated serum ALT level above ULN compared with normal ALT was associated with 3.7-fold increase in likelihood of detectable HBV DNA . Among the treatment-experienced patients, elevated serum ALT level above ULN compared with normal ALT  and Male sex compared Female  were more likely to have detectable HBV DNA . In all patients, ART status and serum ALT level above ULN were associated with HBV viremia. Among the treatment-experienced patients, male sex and ALT level above ULN were associated with Hepatitis B viremia in both univariate and multivariate analysis. Earlier guidelines recommend that HBV DNA be fully suppressed by 12 months of TDF-based therapy in Hepatitis B-HIV infected patients . Howevern = 67) had HBV DNA < 20 IU/mL after 5 years of treatment [n = 15) achieved undetectable HBV DNA in 4 years [n = 111) of co-infected patients demonstrated a virologic response with HBV DNA < 60 IU/mL [A longer duration of TDF therapy is required for achievement of virologic response (HBV DNA < 20 IU/mL) in co-infected patients. Among HBeAg-positive patients, 92 % (reatment . In HBeA 4 years . Similar60 IU/mL . Tenofov60 IU/mL .n = 106) of co-infected patients in Kumasi, Ghana [In our study, HBV DNA was undetectable in 42.0 and 42.3 % irrespective of whether TDF was included in the regimen. The median duration on therapy and IQR for 3TC-only and 3TC + TDF were 4.85 years IQR (3.02–7.16) and 4.87 years IQR (3.81–5.85) respectively. This suggests the influence of other factors in sub-optimal therapeutic response. Poor adherence is often considered as an important factor in patients with persistently detectable HBV DNA on nucleoside analogue therapy . It is ai, Ghana . Among pi, Ghana . Overalli, Ghana .In this study, HIV RNA was not determined. Its cross-sectional design also did not allow for a temporal assessment of the influence of study predictors such as pre-treatment HBV DNA, adherence on virologic response in treated patients. Furthermore, the small sample size of treatment naive patients limited the evaluation of predictors and detectable HBV DNA.The serum aminotransferases are sensitive indicators of liver cell inflammation . The mosWe found that males had higher baseline HBV DNA in comparison to females in naïve and ART-experienced patients. The reason is unclear, and will be in keeping with an increased liver-related mortality rate in men with HIV and HBsAg compared with those with only HBsAg (14.2/1000 compared with 0.8/1000) . ReviewsRoutine Hepatitis B screening is essential in the evaluation and management of HIV patients. Elevated HBV DNA level is associated with overall mortality in HBV/HIV co-infected patients, especially during the period before development of AIDS , and it


Spirulina platensis, may be effective and efficient in delaying HIV disease progression by frequently reported improvement in immune response.Micronutrient deficiencies occur early in Human Immunodeficiency Virus (HIV) infections they have reverse effects on the nutritional status. The diet supplementation with a natural nutraceutical rich in proteins and micronutrient like S. platensis supplementation and standard care or standard care and local balanced diet without S. platenis. Selected hematological and biochemical as well as CD4 count cells, viral load copies were assessed at three separate times.A prospective single-blind, randomized, multicenter study conducted on 320 HIV-1 ARV-naïve participants for 12 months. Participants received either 3–30.87 × 103 copies/mL) of the patients who received a supplementation of S. platensis was found after 6 months of treatment. Haemoglobin level was also significantly higher in the same group while the fasting blood glucose concentration decreased after 12 months compared to control.Among the 169 ART-naïve participants enrolled in the study, the female was mostly represented (67.1 %). The significant increase of CD4 count cells (596.32–614.92 cells count) and significant decrease of viral load levels (74.7 × 10S. platensis to diet combined with a reasonable balanced diet has significantly increased the CD4 cells and reduced the viral load after 6 months. Further studies are recommended among a large specific group of people infected by the HIV in order to investigate the mechanisms involved on the effect of S. platensis on immune system.A daily supplementation with  One of the several Public Health problems faced in Sub-Saharan Africa is HIV/AIDS. In Cameroon, about 4.3 % of the population are infected and this prevalence is particularly higher among women and individuals aged between 15 and 49 years . In AfriMalnutrition potentialises the effects of HIV infection on the individual by promoting chronic fatigue and disease progression, the latter lead to the morbidity and earlier death . MalnutrSpirulina platensis a cyanobacterium available in sub-Saharan Africa has been used as food source. S. platensis represents an important staple diet in humans and has been used as a source of protein and vitamin supplement in humans without any significant side-effects [S. platensis in vitro and in vivo as well as its therapeutic functions [S. platensis with a local balance diet on the level of CD4 cells count and viral load during six months by treatment naïve HIV–infected persons.To fight against nutrients deficiency mainly to HIV patients, the World Health Organization (WHO) recommends nutrients intake of each required micronutrients, which may be taken through nutrient supplementation , 11. Due-effects . It cont-effects , 12. Sevunctions , 14. In unctions –17. ThisWe carried out a longitudinal study in a randomized cohort from January 2011 to February 2012. Patients aged between 18–65 years, HIV–infected naïve to antiretroviral treatment and with CD4 count ≥400 cells/μL were eligible. Patients who had their CD4 below 400 cells/μL during the follow-up were excluded. Informed consent was obtained from every subject prior to the participation for the study. The study was approved by the Cameroon National Ethic Committee under the following reference number: 123/CNE/SE/2011, Cameroon. The subjects were recruited through the medical file obtained from the Day Care Clinic of Central Hospital of Yaounde (CHY) and of the Biyem-Assi District Hospital. Investigation and intervention were carried out at the Etoug-Ebe Health Center.S. platensis to their usual diet during the first six months. The S. platensis was given in package of powder doses of 10 grams per day. The next six months were for the follow-up without S. platensis powder. Subjects were treated “as usual care” treatment in Cameroon. To monitor the biological parameters a standardized questionnaire for demographic characteristics, clinical, quality of live was filled by the subjects at the baseline, during and at the end of the trial.Selected participants (169 subjects) were included in the study. They were divided into two groups: 87 subjects to the control group (first group) and 82 subjects to the intervention group (second group). HIV infected patients were selected after checking patients files for CD4 counts over 400 cells/μL who were ARV naïve to treatment . CD4 T cells values were expressed as cells/μL. The second tube was used for the determination of the viral load with an Abbott Real time PCR . HIV-1 Amplification Strategy with a detection limit of 40 copies/mL. The hemoglobin and the fasting blood sugar levels were determined in the third tube using a standard method and the « ULTRA ONE TOUCH” glucometer  respectively. The level of glycemia higher than ≥ 110 mg/dL was identified like cardiovascular risk factors.t-test version 3.3.2 was used to compare the CD4 lymphocytes counts and viral load of both groups. The statistical SPSS program (version 16.0) was used for all the analysis. Results were considered significant at p-value less than 0.05.Data were presented as mean ± standard deviations or when indicated as percentage. Chi-square was performed to determine the significance of differences in the prevalence of opportunistic infections in both groups. Student’s A total of 169 patients were enrolled on the study among them 50 men and 119 female  than men 23.9 %) in the study with a sex ration of 2.38  and significantly increased until 12 months in the spirulina group while in the control group a linear reduction (R2 = 0.96) of CD4 cells was noted  the S. platensis extract acts by avoiding the virus to attach and penetrate to the cell membrane to infect the cell [ii) the calcium-spirulant (Ca-SP) and sodium spirulant (Na-SP) found in the S. platensis extract interfered with replication of viruses including HIV-1, they increase the activation of macrophages, T-and NK cell activities [iii) Polysaccharides, phycocyanin, glycolipids and sulfolipids idenfied the S. platensis extract increase the immunity by enhancing bone marrow reproduction, growth of thymus and spleen and biosynthesis of serum protein; (iv) the presence of phenolic acids, tocopherols and ß-carotene and other antioxidant molecules identified in S. platensis exhibited the antioxidant protection with many benefits both in vitro and in vivo. Since the baseline characteristics between the two groups were similar, the beneficial effects noted in the Spirulina group demonstrated that nutritional supplementation for 6 months led to an improved recovery of the immune system and an improvement in the body’s ability to fight infections.This present randomized control study included 169 patients divided into two groups either 87 (control group) or 82 (spirulina group). Our sample size and the duration of the experiment were respectively higher and longer than previous study conducted in Cameroon which had 73 and 53 patients for 12 weeks , 18. Thesistance . Our resdiseases . Severalpatients , 22. Ourtivities . Also sethe cell . Therefotivities , 26; (iiS. platensis and six months of follow up of HIV-infected, ART-naïve patients in the early stages of disease significantly delayed time to HIV disease progression and reduced the opportunistic diseases. Daily consumption of S. platensis coupled with nutritional counseling significantly was found to be strongly associated to the significantly reduction of the viral load and increase the CD4 cells counts but the mechanism by which S. platensis may induce these beneficial effects is not well understood. Other studies need to be conducted on the HIV-naive patients to elucidate this mechanism.This clinical trial demonstrated that six months supplementation of


Since the novel H7N9 avian influenza outbreak occurred in China in 2013, neuraminidase inhibitors (NAIs) such as oseltamivir and peramivir have been used as first-line drugs to treat the influenza virus infection. This study aimed to compare the efficacy of oseltamivir-peramivir combination therapy versus oseltamivir monotherapy.A retrospective study of 82 H7N9 confirmed patients was conducted by reviewing medical charts at the First Affiliated Hospital of ZheJiang University in China from April 1, 2013 to Feb 28, 2014. The patients’ clinical information was collected systematically, and we compared the virology and clinical data between oseltamivir monotherapy group (43 patients) and oseltamivir-peramivir combination group (39 patients).p >0.05), respectively. The median decline of Day 2 to Day 0 (initiation of NAIs therapy) viral load was 0.00 and 0.69 log10 copies/μl (p >0.05) respectively in the monotherapy vs. combination therapy groups. The incidence of new Acute Respiratory Distress Syndrome during NAI administration was 63.89 and 77.78 % (p >0.05); while the mortality rates were 25.58 and 43.59 % (p >0.05) in the oseltamivir group vs. oseltamivir-peramivir group.The median duration from NAIs administration to H7N9 virus-negative in oseltamivir monotherapy group and oseltamivir-peramivir combination group was 6.50 and 7.00 days (Our results suggest that in adults with H7N9 virus infection, the use of oseltamivir-peramivir combination therapy was not superior to oseltamivir monotherapy. Since the novel H7N9 avian influenza outbreak occurred in China in 2013, neuraminidase inhibitors (NAIs) such as oseltamivir and peramivir have been important and efficacious in curbing the viral infection . FurtherInfluenza A (H1N1) virus in vitro and in mice [Influenza A H3N2 virus infection during 2008–2009 showed the oseltamivir-zanamivir combination therapy was not more effective than either oseltamivir or zanamivir monotherapy [One such strategy was the combination of two NAIs to treat influenza infections. The basis of the combination therapy was that two or triple antiviral drugs may have additive synergistic effects and reduce drug resistance at the same time –6. Combi in mice . Another in mice . In addi in mice . Furtherotherapy . HoweverThis study was approved by the First Affiliated Hospital of ZheJiang University ethics board.During the outbreak, patients with influenza symptoms onset , pneumonia of unknown origin, or patients who had recently been in close contact with birds or a H7N9-confirmed patient were screened in this study. Respiratory specimens  and blood samples were collected for H7N9 virus laboratory tests and conducted in these suspected cases. There were three methods for H7N9 laboratory diagnosis: real-time reverse-transcriptase-polymerase-chain-reaction assay (RT-PCR) assay, viral isolation, and H7N9 serological testing by modified hemagglutinin inhibition assay , 13. TheInfluenza A (H7N9) virus infection, and acceptance of oral oseltamivir monotherapy or oral oseltamivir and intravenous peramivir combination therapy. All the 82 cases enrolled in the study were admitted during the study period from April 1, 2013 to Feb 28, 2014.This retrospective study was performed at the First Affiliated Hospital of ZheJiang University. Enrollment criteria included age ≥18 years with confirmed We reviewed medical charts and used standardized forms to gather information retrospectively. Clinical and laboratory information was collected systematically from admission to discharge for every patient, including demographic information, baseline and follow-up clinical information.2/ FiO2 ≤200 mmHg with PEEP ≥5 cm H2O), associated with bilateral opacities on chest X-ray, which could not be fully explained by cardiac failure or fluid overload.After admission, respiratory specimens  were collected daily to determine H7N9 viral RNA by PCR analysis. The second negative result of two respiratory samples collected in two-consecutive days was considered the time to stop NAIs therapy and represented an undetectable viral RNA level. We defined the duration between NAI administration and undetectable viral RNA level as the time from NAI administration to virus-negative. As we could not determine the exact virus infection time, following another report , we defiThe primary outcome was the time from NAI administration to virus-negative. The second outcome was the decline of virus load (measured by log10 virus load) between Day 0 (the day NAI therapy was initiated) and Day 2 in patients with confirmed H7N9 virus infection. Based on the viral shedding kinetics in seasonal influenza patients treated by NAIs, the Day 2 viral outcome seemed the most suitable to evaluate virology effects , 16. TheP value <0.05 was considered significant. All analyses were conducted using SPSS for Windows (version 16.0).Categorical variables were calculated by frequency analysis. The numerical variables of normal distributions was represented by means (±standard deviations), abnormal distributions was represented by medians . Two sample Student’s t test was performed to assess the significance of the time from NAI administration to H7N9 virus-negative, the decline of log10 virus load between Day 0 and Day 2. Chi-square test was performed to compare the rate of new ARDS after NAI administration and the total in-hospital mortality. All analyses were two-tailed and the From April 1, 2013 to Feb 28, 2014, about 1950 patients were tested for H7N9 viral infection, and 87 patients were positive. Five of them received other antiviral drugs other than oseltamivir or peramivir. Therefore, a total of 82 adult patients were enrolled in our study. The mean age was 58.21 years (±14.31) and 68.29 % (56 patients) were male. Forty four (53.66 %) patients had one or more coexisting conditions. Hypertension, diabetes, coronary heart disease were the most common coexisting conditions . Eight patients (9.76 %) received NAIs therapy within 48 h after the symptom onset, and 35 patients (42.68 %) within 5 days, 47 patients (57.31 %) more than 5 days. Forty two patients (51.22 %) received glucocorticoid therapy, including intravenous methylprednisolone and intravenous dexamethasone. There were 41 (50 %) patients developed bacterial or fungal infection or colonisation by positive sputum culture during hospitalization, including Among the 82 patients, 39 patients (47.56 %) were enrolled in the first H7N9 wave from April 1, 2013 to May 31, 2013, and 43 patients (52.43 %) were enrolled in the second wave from Nov 30, 2013 to Feb 28, 2014. In the first H7N9 wave, the median age was 63 years and 46.15 % (18 patients) were ≥65 years, while in the second wave, the median age was 57 years and 51.16 % were 50–64 years. Compared with the first wave, more patients in the second wave had no coexisting condition  and oseltamivir-peramivir combination therapy group .Based on the different NAIs drugs they accepted, the 82 patients were divided into oseltamivir monotherapy therapy group  were treated with oseltamivir dosage of 75 mg orally twice daily, 19 patients (44.19 %) were treated with 150 mg twice daily, and one patient (2.33 %) received 75 mg once. In the oseltamivir-peramivir combination therapy group, 24 patients (61.54 %) were treated with 75 mg oseltamivir twice and 600 mg peramivir once daily; five patients (12.82 %) were treated with 75 mg oseltamivir twice and 300 m g peramivir once daily; six patients (15.38 %) were treated with 150 mg oseltamivir twice and 600 mg peramivir once daily; two patients (5.13 %) were treated with 150 mg oseltamivir twice and 300 mg peramivir once daily; one patient (2.56 %) changed oseltamivir dosage to 150 mg twice daily after 10 days of 75 mg twice daily with 600 mg peramivir once daily; one patient (2.56 %) changed oseltamivir dosage to 75 mg twice daily after 6 days of 75 mg once daily in the local hospital with 600 mg peramivir once daily.p >0.05). The median time from antiviral treatment to viral-negative in oseltamivir monotherapy group was 6.50 days , and oseltamivir-peramivir combination therapy group was 7.00 days   patients and the oseltamivir-peramivir group had 32 (82.05 %) patients. The median time from NAIs therapy to negative viral RNA shedding of the remaining 70 patients was 7.00 days . The median duration of viral shedding in the whole 82 patients was 12.00 days , in the oseltamivir monotherapy group was 11 days , and in the oseltamivir-peramivir combination group was 13 days . Baseline information of the two groups is listed in Table p >0.05; Table Among the 82 confirmed patients, there were 60 patients with both Day 0 and Day 2 viral data, comprising 31 patients in the oseltamivir-peramivir combination therapy group, and 29 patients in oseltamivir monotherapy group. The median viral load decrease of Day 2 to Day 0 was 0.69 log10 copies/μl  in the oseltamivir-peramivir combination therapy group, and 0.00 log10 copies/μl  in the oseltamivir monotherapy group  patients had ARDS before receiving NAIs treatment in the oseltamivir monotherapy group, and 3 (9.10 %) patients in the oseltamivir-peramivir combination therapy group. These 10 patients were therefore excluded from this portion of the study, and 36 patients remained in each treatment group. The incidence of newly-developed ARDS during NAI therapy was 23/36 (63.89 %) in oseltamivir monotherapy group and 28/36 (77.78 %) in the oseltamivir-peramivir combination therapy group pdm09 virus [Most in vitro studies that assess the efficacy of combination NAIs therapy have shown synergism and additive effects. The combination of antivirals not only decreases the emergence of resistant strains, but also ensures antiviral effect –6. There9 season , and a r09 virus . Oseltam09 virus , and antInfluenza A/NWS/33 (H1N1) virus [The patients initiated NAI therapy a relatively long time after the onset of influenza symptoms . Wiku Adisasmito et al. found that H5N1 patients could still benefit when initiated oseltamivir up to 6–8 days after onset of symptoms . Here, tIn the current study, most patients in the oseltamivir monotherapy group  were enrolled in the first H7N9 wave, while all the patients in the oseltamivir-peramivir combination group were enrolled in the second H7N9 wave. It was observed that H7N9 virus in the second wave to infect younger patients with less concomitant disease. The second wave had higher mortality. The influenza virus is an RNA virus, with a high error rate during transcription . The oseInfluenza A (H1N1) virus in vitro and in mice showed additive to synergistic effects [In the current study, the dosage of NAI varied widely. The majority of the patients  received the standard or double dosing of NAIs. Two oseltamivir dose-comparison studies in influenza showed no difference from standard dosage (75 mg twice daily) or double dosage (150 mg twice daily) in virological clearance or clinical outcome , 26. OneThe current study suggested that the median duration of viral shedding of H7N9 virus was 12 days . The viral load of the seasonal influenza usually shed for 5 days in adults , 28. TheThere are several limitations to the current study. First, it was a retrospective study and the sample size was too small for robust investigation in subgroups of patients. Despite the use of a standardized case-report form, not all information was available for all 82 patients. Secondly, the H7N9 viral RNA of these 82 cases was not sequenced for analysis of antiviral resistance. It was not possible to determine whether there were any mutations associated with resistance to NAIs in either group, especially the NA-R292K. Third, the dosage of oseltamivir and peramivir varied widely. The heterogeneity may influence the antiviral effect of two groups. Fourthly, most people in our study received NAIs more than 5 days, not the optimal time. It may influence the effect of the NAIs. Further prospective clinical studies should be performed to compare the effect between NAI monotherapy and combination therapy with two NAIs with standardization NAI administration. And the NAI-resistance should be analyzed.In conclusion, our study has shown that oseltamivir-peramivir combination was not superior to oseltamivir monotherapy for treating H7N9 influenza; therefore the use of the combination therapy may not be useful when treating critically ill patients.


Plasma HIV-1 RNA monitoring is one of the standard tests for the management of HIV-1 infection. While HIV-1 RNA can be quantified using several commercial tests, no test has been commercialized for HIV-2 RNA quantification. We studied the relationship between plasma HIV-2 viral load (VL) and CD4 count in West African patients who were either receiving antiretroviral therapy (ART) or treatment-naïve.A cross sectional survey was conducted among HIV-2-infected individuals followed in three countries in West Africa from March to December 2012. All HIV-2 infected-patients who attended one of the participating clinics were proposed a plasma HIV-2 viral load measurement. HIV-2 RNA was quantified using the new ultrasensitive in-house real-time PCR assay with a detection threshold of 10 copies/ mL (cps/mL).3 and 43 (46.7%) of these patients had a detectable VL (≥10 cps/mL). Among the 220 patients receiving ART, the median CD4-T cell count rose from 231 to 393 cells/mm3 (IQR [259-561]) after a median follow-up duration of 38 months and 145 (66.0%) patients had CD4-T cell count ≤ 500 cells/mm3 with a median viral load of 10 cps/mL (IQR [10-33]). Seventy five (34.0%) patients had CD4-T cell count ≥ 500 cells/mm3, among them 14 (18.7%) had a VL between 10-100 cps/mL and 2 (2.6%) had VL >100 cps/mL.A total of 351 HIV-2-infected individuals participated in this study, of whom 131 (37.3%) were treatment naïve and 220 (62.7%) had initiated ART. Among treatment-naïve patients, 60 (46.5%) had undetectable plasma HIV-2 viral load (<10 cps/mL), it was detectable between 10-100 cps/mL in 35.8%, between 100-1000 cps/mL in 11.7% and >1000 cps/mL in 6.0% of the patients. Most of the treatment-naïve patients (70.2%) had CD4-T cell count ≥500 cells/mmThis study suggests that the combination of CD4-T cell count and ultrasensitive HIV-2 viral load quantification with a threshold of 10 cps/mL, could improve ART initiation among treatment naïve HIV-2-infected patients and the monitoring of ART response among patients receiving treatment. Infection due to Human Immune deficiency Virus type 2 (HIV-2) is predominantly found in West Africa and in countries with historical ties to this part of the world . This in3 had detectable HIV-2 viral load and 38% of patients with CD4 count <300 cells/mm3 had detectable HIV-2 viral load ) and there were 202 (57.2%) women. The main baseline and follow-up characteristics of the study population are summarized in 3 and did not vary much over time, moving to 619 cells/mm3 at the time of blood collection, after a median follow-up duration of 27 months (3 (IQR [131–395]) and 393 cells/mm3 at the time of sampling. The first-line ART regimen was mainly based on an association of 2 NRTI and a boosted PI (79.0%). Lopinavir/ritonavir was the most prescribed boosted PI (initiated in 66.1% of patients), followed by Indinavir/ritonavir (33.9%). Fourteen patients (6.4%) received 3 NRTI combinations, mainly AZT+3TC+abacavir (86.0%) and 14.6% of the patients in the cohort initiated NNRTI–based regimen.Among the 131 treatment-naïve patients, the median CD4-T cell count at enrolment in the cohort was 601 cells/mm7 months . Among tWith a threshold of 10 cps/mL the viral load was detectable in 146 (41.6%) of the 351 samples tested. When thresholds of 50 and 100 cps/mL were used, the viral load was detectable in only 65 (18.5%) and 59 (16.8%) samples respectively.3 and within them 43 (46.7%) had a detectable viral load (≥10 cps/mL) including 35 (81.5%) between 10 and 100 cps/mL. For the 39 (29.8%) treatment-naïve patients with CD4-T cell count <500 cells/mm3, the viral load was detectable in 70.0% of them. The proportion of patients with detectable viral load decreases when CD4-T cell count increases. Sixty (46.5%) treatment-naïve patients had undetectable plasma HIV-2 viral load (<10 cps/mL), 35.8% had it between 10–100 cps/mL, 11.7% between 100–1000 cps/mL and 6.0% >1000 cps/mL . The med3 at ART initiation to 393 cells/mm3 (IQR [259–561]) after a median duration on ART of 38 months (IQR [40–54]) and 145 (66.0%) patients had a CD4 count < 500 cells/mm3. The viral load was undetectable (<10 cps/mL) for 144 (65.5%) patients, 40 (18.8%) other patients had a viral load between 10–100 cps/mL, 14 (7.3%) between 100–1000 cps/mL and 22 (10.0%) >1000 cps/mL  had a viral load between 10–100 cps/mL and 2 (2.6%) had viral load > 100 cps/mL. Among the 75 (34.0%) patients with CD4-T cell count ≥ 500 cells/mm3 have a viral replication. For patients receiving ART, we described the viral suppression over time and highlighted the fact that one patient out of five with CD4-T cell count >500 cells /mm3 had a detectable viral load, indicating treatment failure. This study confirmed the low viral replication already reported in treatment naïve HIV-2-infected individuals [This study quantified plasma HIV-2 viral load in 351 plasma samples from West African HIV-2-infected individuals using an ultra-sensitive procedure (threshold 10 cps/mL) of the new HIV-2 quantification assay developed by the ANRS . We firsividuals ,14,28, aThe use of viral load to monitor ART response among HIV-1-infected patients is a common recommendation of WHO already implemented in resources constrained settings . Regardi3 according to 2013 WHO guidelines [3 according to 2013 French and US guidelines [3 had a detectable viral load (≥10 cps/mL) and among this group of patients, 81.5% had viral load between 10 and 100 cps/mL, making them undetectable with less sensitive assays. Using an ultrasensitive assay could help detect viral replication in these patients who will therefore be eligible for ART according to the latest French guidelines [The successful ART scaling up  and the idelines  or in alidelines . The aimidelines ,32,33 anidelines . The dec2E collaboration [3 and a detectable viral load after treatment, since our study was a cross-sectional survey, we did not perform a CD4-T count control.Despite the clinical importance of these results among HIV-2 infected individuals, our study presents some weaknesses. The genetic diversity of HIV-2 with 9 groups ,34 was nboration . Thus thThe main lesson learned from this study is that the clinical use of viral load is clearly different in treatment-naïve and in ART-receiving patients. In treatment naïve patients, the detection of viral load improved knowledge on the natural history of the infection, showing that more treatment-naïve patients have a viral replication. This finding is important because it could help guiding treatment initiation in these patients. Regarding patients on ART, viral load measurement could help to monitor the treatment efficacy, detect treatment failure and help to reinforce observance. Our results also emphasize the need of an ultrasensitive assay to quantify HIV-2 viral load.This study highlights the relationship between plasma HIV-2 viral load and CD4-T cell count in HIV-2-infected individuals. It suggests that the combination of CD4-T cell count and ultrasensitive HIV-2 viral load quantification with a threshold of 10 cps/mL, could improve both ART initiation decision for HIV-2 treatment-naïve patients and the monitoring of ART response among HIV-2-infected individuals.S1 Dataset(XLSX)Click here for additional data file.


The growing numbers of HIV-infected patients requiring second-line antiretroviral therapy (ART) in Vietnam make essential the evaluation of treatment efficacy to guide treatment strategies.We evaluated all patients aged ≥15 years who initiated second-line ART after documented failure of first-line therapy at the Hospital for Tropical Diseases in Ho Chi Minh City. The primary outcome was time from second-line ART initiation to death, or to a new or reoccurrence of a WHO-defined immunological or clinical failure event, whichever occurred first. Risks of treatment failure and death were evaluated using Cox proportional hazards modeling.Data from 326 of 373 patients initiating second-line ART between November 2006 and August 2011 were included in this analysis. The median age was 32 years (IQR: 28–36). Eighty one percent were men. The median CD4 count was 44 cells/μL (IQR: 16–84). During a median follow-up of 29 months (IQR: 15–44), 60 (18.4%) patients experienced treatment failure, including 12 immunological failures, 4 WHO stage IV AIDS events, and 44 deaths (13.5%). Sixty percent of deaths occurred during the first 6–12 months. The Kaplan–Meier estimates of treatment failure after 1, 2, 3, and 4 years were 13.1% (95% CI: 9.2–16.8), 18.6% (95% CI: 14.0–23.1), 20.4% (95% CI: 15.4–25.1), and 22.8% (95% CI: 17.2–28.1), respectively. Older age, history of injection drug use, lower CD4 count, medication adherence <95%, and previous protease inhibitor use independently predicted treatment failure.While treatment efficacy was similar to that reported from other resource-limited settings, mortality was higher. Early deaths may be averted by prioritizing second-line therapy for those with lower CD4 counts and by improving treatment adherence support. Despite generic production for resource-limited countries, a second-line regimen containing ritonavir-boosted lopinavir (LPVr) costs 6 times that of a first-line regimen.5 In most low and middle-income countries, second-line therapy is the last option for patients failing treatment with drug resistance. As third-line therapy is forbiddingly expensive and is unavailable in resource-limited countries, it is imperative for national programmes in these settings to maximize the efficacy and durability of second-line therapy.The availability of low-cost fixed-dose combination antiretroviral drugs has enabled rapid scale-up of antiretroviral therapy (ART), resulting in substantial reduction in morbidity and mortality due to HIV in resource limited countries.6 Nearly 90,000 people were on ART as of 2014, and an estimated 3% were on second-line therapy.6 The HIV system in Vietnam is undergoing a critical transition from an international-donor to a national-funding approach that integrates with the national health insurance programme.7 Outcome data on second-line therapy in Vietnam are lacking, but are important for the national programme to devise treatment strategies and to forecast treatment options beyond second-line therapy. In this study, we investigate second-line therapy outcomes and factors that determine therapy failure and death at the largest HIV referral centre in southern Vietnam.Vietnam is among the countries with the highest HIV burden in Asia with an estimate of 280,000 people living with HIV.8This was a retrospective analysis of adult patients who switched to second-line therapy in a cohort of over 4000 patients on the national ART programme at the Hospital for Tropical Diseases (HTD) in Ho Chi Minh City (HCMC). This is the largest primary and referral center for HIV care in southern Vietnam (population around 45 million). The national ART programme began providing free antiretroviral drugs through international funding support in 2003. First-line therapy consisted of zidovudine (AZT) or stavudine (d4T) in combination with lamivudine (3TC) and nevirapine (NVP). Prior to the availability of efavirenz in 2004, cases of NVP-related toxicity were switched to indinavir (IDV). Second-line therapy became available in 2006 initially including abacavir (ABC), didanosine (ddI), and nelfinavir (NFV). In 2007, LPVr replaced NFV, and in 2009 tenofovir (TDF) and 3TC replaced ABC and ddI as the nucleotide reverse transcriptase inhibitor (NRTI) backbone.10 Patients were required to come to the clinic monthly for clinical evaluation and medication pick-up. CD4 count was measured every 6 months. HIV viral load was tested at the time patients were diagnosed with immunological or clinical failure and was confirmed with repeat testing. HIV viral load was performed using a generic real-time PCR assay  with a limit of detection of 250 copies/mL.11 Virological failure was defined as confirmed HIV RNA levels ≥5000 copies/mL as per WHO guidelines during the study period.10 HIV genotyping was performed to evaluate for drug resistance prior to therapy switch using a published in-house assay12 on the Beckman Coulter CEQ 8000 platform. Both HIV viral load and genotyping tests were performed at the Pasteur Institute, a WHO-accredited HIV reference laboratory, in HCMC.ART was monitored using immunological and clinical failure criteria based on the WHO's guidelines for settings without routine viral load monitoring.We included all HIV-infected patients aged ≥15 years who initiated second-line therapy due to documented immunological and/or clinical failure of first-line therapy. Patients who were alive and well but had been on second-line therapy for <6 months, and those without documented treatment failure to first-line therapy, were excluded. The study was approved by the Scientific and Ethical Committee of the HTD.10 The secondary outcome was time to death.The primary outcome was treatment failure and was defined as time from second-line ART initiation to death, or to a new or reoccurrence of an immunological or a clinical failure event, whichever occurred first. Immunological failure was defined by the WHO as a decrease of CD4 count to or lower than baseline, a decrease of >50% of peak CD4 value while on treatment, or a persistent CD4 count of <100 cells/μL after at least 6 months of continued ART. Clinical failure was defined as new occurrence or reoccurrence of a WHO stage IV disease.Routinely collected clinical and laboratory data were recorded on a standardized form and included demographic information, history of injection drug use (IDU), antiretroviral drug timeline, WHO stage 4 AIDS events, 6-month serial CD4 counts, HIV viral load, and HIV genotype when these were available, ART adherence evaluation, deaths, and causes of death.13 Additionally in patients who were in active follow-up, adherence was prospectively evaluated using a simple self-reported Visual Analogue Scale (VAS).14 This VAS has been shown to be as reliable as other methods such as pill-count and 3-day recall self-report, yet much simpler to administer.15ART adherence counseling was provided to patients pre- and post-second-line ART initiation according to standard of care. Adherence was routinely assessed by the clinicians according to the MOH guidelines and was recorded either as an estimated percentage of pills taken, or as a qualitative assessment of “good,” “average,” or “poor,” corresponding to ≥95%, 80% to 94%, or <80% adherence, respectively.For analysis, suboptimal adherence was defined as having at least 1 adherence score of <95% by pill count, by the VAS, and/or receiving at least 1 qualitative adherence assessment of “average” or “poor” over a 6-month period preceding an outcome event or preceding the time of study assessment in patients who had not had an event.The cumulative incidence of treatment failure and failure rates after 1, 2, 3, and 4 years and corresponding 95% confidence intervals were calculated using the Kaplan–Meier method. The Cox proportional hazards model was used to analyze the time to treatment failure (composite primary endpoint) and the time to death (secondary endpoint). Patients who were transferred to other provincial or district clinics while on second-line therapy had been judged by doctors to be clinically and immunologically stable before the transfer. For the analysis, event-free transferred patients were censored at the time of transfer . Alternatively, assuming the transferred patients were doing well clinically and immunologically on therapy, we treated them as censored at the time-point where their last monthly follow-up visit would have been had they not been transferred .21 Although recommended by the WHO guidelines,10 not all patients in the cohort had a viral load performed to document virological failure to first-line therapy prior to second-line therapy initiation. We performed a sensitivity analysis including only patients with documented virological failure to first-line therapy (confirmed HIV RNA levels ≥5000 copies/mL). The proportional hazards assumption was assessed by examining plots of weighted Schoenfeld residuals and by formal testing. There was strong evidence of nonproportional hazards for the effect of CD4 cell count at second-line therapy initiation on treatment failure . To account for this, we decided to model a time-varying effect on the hazard of treatment failure with separate effects for the first year of follow-up and subsequently. There was no clear evidence for nonproportional hazards between any other covariates and the primary or secondary endpoint . Both univariate and multivariable Cox regressions were performed.The following predefined covariates were included in the model: age at second-line therapy initiation, history of IDU (yes/no), CD4 cell count, and (log10-transformed) HIV RNA viral load at second-line therapy initiation, second-line therapy delay , history of protease inhibitor (PI) use, and an overall measure of therapy adherence (<95% vs. ≥95%). The chosen covariates have been shown to be associated with poor ART outcome.T and Nelson-Aalen estimator H(T)].22 All reported confidence intervals are 2-sided 95% intervals and analyses were performed with the statistical software R version 2.15.0,23 and the companion R package mice version 1.2.5 (for multiple imputation).24Data were analyzed based on multiple imputations of missing data and on a complete-case analysis. To avoid bias, the imputation algorithm included the endpoints  and were included in this study. Approximately 50% of patients came from HCMC; the rest from the remaining 17 southern provinces of Vietnam. The median duration of first-line ART treatment was 33 months (IQR: 21–44). The characteristics of the 326 patients at the time of initiation of second-line therapy are summarized in Table HIV genotyping was performed for 246 of 326 (75.5%) patients who failed first-line therapy. Mutations conferring high-level resistance to NRTIs were detected in 238 of 246 patients (96.7%), to nonnucleotide reverse transcriptase inhibitors (NNRTIs) in 229 of 246 (93.1%), and to PIs in 6 of 246 (2.4%). Resistance mutations to both NRTIs and NNRTIs were present in 226 of 246 patients (91.9%) and to all 3 drug classes in 5 of 246 patients (2.0%). The most common NRTI mutations were M184I/V (85.4%), thymidine analog mutations (TAMs) M41L, D67N, K70E/R, T215F/Y, and K219E/Q (30–55%), Q151 M (21.1%), and K65R (14.6%). Two patients had a T69 insertion mutation. The most common NNRTI mutations were Y181C/I/V (45.5%), G190A/S (41.9%), and K103N (31.3%). The most common PI mutations were I54 V (2.4%), M46I/L (2.8%), V82A (2.0%), and L90 M (1.2%).The predicted susceptibility to the national second-line regimen containing TDF, 3TC, and LPVr was evaluated for the 246 patients who had genotype results using the Stanford HIV Drug Resistance Database . Intermediate to high-level resistance to TDF was present in 161 of 246 (65.4%), to 3TC in 230 of 246 (93.5%), and to LPVr in 5 of 246 (2.0%).In total, 320 patients (98.2%) received LPVr in combination with 2 NRTIs and 6 patients received NFV with 2 NRTIs Table . One hunP values: 2.34 (1.53–3.59), P < 0.001; 3.09 (1.50–6.38), P = 0.002; 0.54 (0.32–0.89), P = 0.016); 3.07 (1.52–6.22), P = 0.002; and 2.22 (1.06–4.67), P = 0.035), respectively.The 7 covariates entered in the Cox model are listed in Table P = 0.715).The cumulative incidence of treatment failure among patients according to IDU, treatment adherence, and PI is shown in Figures Pneumocystis jiroveci pneumonia , candida esophagitis , cryptococcal meningitis (2), Penicillium marneffei infection (2), herpes simplex (2), cytomegalovirus (2), toxoplasmosis (2), hepatitis C-related liver failure (2), renal failure of unclear etiology (2), nontyphoid salmonella sepsis (1), and AIDS-associated wasting . Univariate and multivariate Cox regression analyses were performed to determine predictors of death. Consistent with the results of the primary outcome analysis, lower CD4 count  and suboptimal adherence  predicted death in both univariate and multivariate analyses. Age and history of IDU did not predict death in the univariate analysis, but in the multivariate analysis older age and history of IDU became significant predictors of death .Deaths occurred in 44 patients (13.5%) and accounted for 73.3% of failure events; 39 were AIDS-related deaths, and 5 were unknown or non-AIDS-related deaths. The median time to death was 9 months (IQR: 3–22), with 26 deaths (59.1%) occurring within the first 6 to 12 months. The causes of AIDS-related deaths included microbiologically confirmed tuberculosis , 25 A 27-cohort study comprising 632 patients from Africa and Asia reported a failure rate of 28% over 2 years.16 Treatment failure in that study was defined as the first diagnosis of clinical, immunological or virological failure, or death. Only 4 of these 27 centers had routine viral load monitoring. These failure rates reflect the reality of HIV care in resource-limited settings. Our treatment failure rates are lower when compared with studies that use virological failure as the measure of outcomes. A meta-analysis of 2035 patients from 19 cohorts across low-income and middle-income countries reported failure rates of 22% to 38% after 6 to 36 months on second-line ART.25 Although an outcome measure relying on CD4 count and clinical evaluation can underestimate virological failure rates, the long-term clinical and health economic benefits of routine viral load monitoring still need to be determined in resource-poor settings.To our knowledge, this is the first study to systematically evaluate the outcomes of second-line ART in patients who fail first-line therapy in Vietnam according to the WHO's immunological and clinical criteria. Despite the profound level of immune deficiency in this patient cohort (median CD4 count: 44 cells/μL), high viral replication (median HIV RNA: 5.1 log copies/mL), and extensive resistance to second-line NRTI backbone  at the time of treatment switch, treatment failure rates are similar to studies in comparable settings.16 The median time to death was shorter, 9 months (IQR: 3–22) versus 15 months (IQR: 12–26).16 Death accounted for 73% of failure events, and 90% of deaths were due to AIDS-related infections. Using multivariate analysis, death during the first year was predicted by lower CD4 counts at second-line ART initiation. The median CD4 count and HIV viral load at second-line ART initiation were 44 cells/μL and 5.1 log copies/mL, respectively. This compares with median CD4 counts >100 cells/μL and HIV viral load ranging from 3.9 to 4.8 log copies/mL in other African and Asian cohorts.27 The lower CD4 count likely explains the higher mortality observed in our patients and suggests second-line therapy delay plays a role. When comparing to studies from other resource-limited countries, therapy delay in our cohort was longer, median 9 (range: 5–15) versus 5 (range: 1–8) months.26While the overall treatment failure rates were similar to equivalent settings, the mortality in our cohort was higher: 13.5% versus an estimate of 5% in the 27-cohort study from Africa and Asia.20 The longer a nonsuppressive ART regimen is given, the higher the chance of developing accumulation of drug resistance mutations, impairing the efficacy of current as well as future ART options.Therapy delay can be explained by programmatic reasons not unique to Vietnam. Without viral load monitoring, it can take months to years for a patient who fails treatment virologically to manifest failure immunologically and clinically. The process of defining treatment failure in Vietnam may be too conservative. Immunological failure is usually confirmed with a repeat CD4 measure in 3 to 6 months. Cases of confirmed immunological and/or clinical failure are referred to a panel of experts located in centers that have access to viral load and drug resistance testing. The referral process, albeit necessary in some circumstances, further delays the onset of therapy switch. When the relationship of treatment delay and CD4 count was assessed with respect to therapy failure and death in our study, only lower CD4 count predicted therapy failure and death. The likely explanation for this is that these 2 variables are interdependent or have a causal relationship, that is, treatment delay directly results in a decline in CD4 count. In our cohort, the CD4 count is likely the stronger variable that drives the outcomes. When these same variables were evaluated by Levison et al. in a study using virological failure as outcome, they found that therapy delay and not CD4 count predicted virological failure, and for every month a patient remained on a failing first-line ART regimen there was a 7% increase in risk of lack of virological suppression.34 Older age at ART initiation has been linked to poor immunological recovery, loss to follow-up, and death in ART cohorts in Zambia and South Africa.31 CD4 count and treatment adherence are established risk factors for first-line ART failure and are also found to be independent predictors of second-line ART failure in the analysis of 27 ART programmes across Africa and Asia.16 PI exposure has been associated with risks of virological failure in Cambodia and India.36 Treatment adherence in our study was assessed at multiple time points, suggesting that suboptimal adherence at anytime during therapy predicts treatment failure. This finding is similar to other second-line cohorts from South Africa, Malawi, and Thailand,21 highlighting the importance of ongoing patient education and adherence support in improving treatment outcomes. The history of IDU predicted treatment failure independently from treatment adherence. This effect is likely multifactorial and likely involves cofactors that are not measured in this study including nutritional status, social economic status, and hepatitis B and/or C coinfection.In our cohort older age, history of IDU, PI exposure, lower CD4 count, and suboptimal adherence were independent predictors of treatment failure in both complete-case and multiple imputation analyses. With the exception of PI exposure, these variables remained independent predictors of death. These findings are consistent with the published literature on first-line ART failure worldwide.37 The safety data of this super-boosted ritonavir regimen in HIV-associated tuberculosis are very limited,39 and significant side effects and hepatic toxicity have been reported in healthy volunteers.40 Therefore, many clinicians in developing countries defer second-line therapy until the rifampicin-containing phase of tuberculosis treatment is complete. This delay may explain the significantly higher treatment failure rate (51%) in patients with tuberculosis coinfection in our cohort.Coinfection with tuberculosis in patients failing ART is common in countries where tuberculosis is highly endemic. In our study, microbiologically confirmed tuberculosis accounted for the majority of AIDS-related deaths (30%). HIV and tuberculosis coinfection is a common reason for clinicians to delay second-line ART initiation because of the rifampicin and PI drug–drug interactions via CYP450 metabolism pathway. The WHO recommends rifabutin in place of rifampicin for patients who need tuberculosis treatment while on PI therapy. Unfortunately rifabutin is not yet available in Vietnam. A super-boosted dose of ritonavir (400 mg twice daily) is therefore recommended.Our study has limitations. This is a single-center study; therefore, the criticism is that the patients may not be representative of the entire adult population on second-line therapy in Vietnam. However, the HTD is the largest center for HIV in Vietnam and is the primary provider of second-line therapy for patients in southern Vietnam. Half of the patients in this cohort come from the 17 southern provinces of Vietnam, representing a wide selection of patients. Further this study is the largest second-line therapy cohort in Southeast Asia, allowing for robust analyses of clinical outcomes. Another limitation is that clinical data are not prospectively collected. However, all study variables were consistently assessed in a standardized way in accordance with the national guidelines. We expect the quality of data is approaching that of a prospective study. Lastly, the attrition rate was high (16%); however, this is largely due to transfer of care (52 patients). Only 1 patient was lost to follow-up.In summary, this is the first study to report the outcomes of second-line ART with a LPVr-based regimen in Vietnam. The overall treatment failure rate using immunological and clinical criteria is 18.4% after a median follow-up of 29 months. Early AIDS-associated death is the main result of treatment failure and is predicted by older age, history of IDU, lower CD4 count at therapy switch, and medication adherence levels <95%. In the absence of routine virological monitoring, interventions to prioritize timing of second-line ART based on CD4 counts and to support medication adherence will improve the treatment outcomes of patients on second-line ART in Vietnam.


We evaluated a low-cost virological failure assay (VFA) on plasma and dried blood spot (DBS) specimens from HIV-1 infected patients attending an HIV clinic in Harare. The results were compared to the performance of the ultrasensitive heat-denatured p24 assay (p24). The COBAS AmpliPrep/COBAS TaqMan HIV-1 test, version 2.0, served as the gold standard. Using a cutoff of 5,000 copies/mL, the plasma VFA had a sensitivity of 94.5% and specificity of 92.7% and was largely superior to the VFA on DBS  or to the p24  when tested on 302 HIV treated and untreated patients. However, among the 202 long-term ART-exposed patients, the sensitivity of the VFA decreased to 72.7% and to 35.7% using a threshold of 5,000 and 1,000 RNA copies/mL, respectively. We show that the VFA (either on plasma or on DBS) and the p24 are not reliable to monitor long-term treated, HIV-1 infected patients. Moreover, achieving acceptable assay sensitivity using DBS proved technically difficult in a less-experienced laboratory. Importantly, the high level of virological suppression (93%) indicated that quality care focused on treatment adherence limits virological failure even when PCR-based viral load monitoring is not available. In 2011, 34 million people were estimated to be living with HIV/AIDS of whom 69% reside in sub-Saharan Africa  was used to define the optimal p24 cutoff value to determine virological failure at 5,000 or 1,000 RNA copies/mL as per VLref assay. Differences between groups were calculated using the Student's t-test for numerical values and the chi-square test for categorical data. The level of significance of α was set at 0.05. Agreement between quantitative results of VFA, the p24, and VLref was evaluated using a Bland Altman analysis.Data analysis was performed using both the previous  and current  cutoff values to define virological failure according to WHO guidelines . IndividData were analyzed using SPSS version 20.0  and Graph Pad Prism version 6.00 for Windows .μL) as compared to the ART-naive group (17.1% versus 9%). Thirty-eight patients (18.8%) of the treatment-exposed group had immunological failure.One hundred ART-naive and 202 ART patients treated for at least 36 months were included in the study. There were no significant differences in age and gender between the two patient groups . Most paref > 5,000 copies/mL and 92 a VLref > 1,000 copies/mL. Among the ART-treated patients these figures were 11 (5.4%) and 14 (6.9%), respectively.Among the 100 ART-naive patients, 81 had a VLThe standard curve was built from testing serial dilution of one well-characterized plasma aliquots (data not shown). Mean Ct-values and SD corresponding to 5,000 and 1,000 RNA copies/mL were extrapolated from the equations of three individual standard curves. Thresholds Ct-values defined as [mean ± 2SD] were calculated and equaled to 36.65 ± 0.04 for a cutoff of 5,000 copies/mL and 38.56 ± 0.35 for a cutoff of 1,000 copies/mL. The same threshold Ct-values were used for both plasma and DBS.ref results. Three hundred one plasma specimens were tested with the p24 assay and 300 with the VFA on plasma. Two hundred ninety-nine DBS specimens were tested with the VFA.A total of 302 specimens had VLref was the highest in the plasma VFA  followed by the DBS VFA .Area Under the ROC Curve  indicated that the capacity to predict virological failure as per VLOverall, the percentage of samples correctly classified was the highest with plasma VFA,  or VLref > 5,000 copies/mL (sensitivity = 49.0%) with more than a quarter of the samples being misclassified using either threshold . The Youden index indicated that the optimal cutoff to identify virological failure using the p24 assay was 3 pg/mL for the threshold of 5,000 RNA copies/mL and 2.65 pg/mL for the threshold of 1,000 RNA copies/mL as per VLhreshold . Sensitihreshold .ref . Additional disagreements were observed with 4 samples giving false positive values using plasma VFA as compared to VLref.Although the VFA is not designed for quantitative VL determination, we conducted a Bland Altman analysis to further explore the agreement between measurements with plasma VFA and the VLref . The gran = 106), using a threshold of 1,000 RNA copies/mL, indicated an overrepresentation of treated patients amongst the false negative . No differences in gender, age, or CD4 count were observed between correctly classified and misclassified virological failures. Findings were similar when the analysis was done using a threshold of 5,000 RNA copies/mL to define virological failure (data not shown).Using a threshold of 5,000 RNA copies/mL, the plasma VFA misclassified 5 samples as false negative and 15 samples as false positive see . Using aref ≤ 5,000 RNA copies/mL, VLref values were significantly higher in the 15 samples classified as false positive using the plasma VFA as compared to the 195 correctly classified samples .Conversely, among a total of 210 samples with VLref and plasma VFA as well as levels of p24 concentration between groups of patients categorized as a function of WHO-defined immunological failure. Both VLref and plasma VFA quantitative measurements were significantly higher in patients with immunological failure  as compared to immunological responders. In contrast, there was no significant difference in p24 concentration in immunologically failing patients as compared to the other participants .In order to explore the utility of the three assays beyond the identification of virological failures, we compared levels of viral load as per VLThe cost of the VFA in plasma was calculated based on one run of 19 plasma samples, including positive and negative controls. Labor costs were based on an average of 2.5 hours spent by one laboratory technician per test run. Overall, the cost of the VFA in Zimbabwe (US$ 30.31) was very similar to South Africa  in plasma and DBS as compared to the p24 assay, using the commercial viral load assay from Roche (CAP/CTM) as the gold standard for the identification of virological failure in a group of treated and ART-naive HIV-1 infected patients. The evaluation was done in a local setting and included samples from patients treated for at least 36 months, in whom the performance of an alternative viral load assay is most relevant to study.ref. AUC, sensitivities, and specificities of the plasma VFA were comparable to previous reports on similar panels of samples [The data indicate that the plasma VFA showed the best performance in identifying virological failures as compared to the DBS VFA and the p24 assay, using a cutoff of either 5,000 or 1,000 RNA copies/mL as per VL samples , 24. In  samples , 24. The samples . However samples , 31. In  samples , 24, theref . Despite initial observations indicating that heat-denatured p24 is a good alternative to HIV-1 RNA load [ in vivo. This translates into p24 reactivity being measured in samples from patients receiving long-term ART where HIV-1 RNA is undetectable [Overall, the p24 assay had a poor capacity to classify samples into virological or nonvirological failure, regardless of the cutoff used. More than a quarter of the samples tested were misclassified mostly due to overcalling of the VLRNA load , subsequRNA load . SeveralRNA load , 33. OurRNA load , 35 in vtectable , presumaThe sensitivity of VFA in plasma and DBS decreased substantially in the group of long-term treated patients. A plausible explanation is that VL associated with virological failure might be relatively lower than those measured in ARV-naive individuals, with greater odds of being classified as false positives by the VFA. Previous evaluations of the VFA were conducted on panels of samples artificially composed to cover very high to low levels of viral load, regardless of ART exposure , 24. OurThe present findings do not support the use of the VFA (either on plasma or on DBS) or the p24 for the reliable monitoring of long-term virological response to ART. To date, no consensus has emerged on acceptable rates of misclassification for the field use of alternative VL assays. However, it is our feeling that missing more than 20% of patients failing their treatment is unacceptable, given the fact that false positive patients are not eligible for a retesting before the next (bi-)annual visit , in the ref and VFA, there was no association between higher p24 levels and more advanced disease stages as defined by CD4 count. Although intrapatient longitudinal changes of p24 concentration have been shown to have some value in identifying patients at risk of disease progression [Interestingly and in contrast with VLgression , 37, ourOur observations indicate that the VFA technology transfer to a less-sophisticated laboratory was feasible in this setting. Procurement of reagents and consumables is one of the main barriers limiting adequate operation of medical laboratories in sub-Saharan Africa. Hence, careful consideration and planning are needed when implementing a new assay in the field; especially when this assay is based on an open platform. In this case, instruments and reagents were procured relatively fast and at a reasonable price, albeit with significant logistical and administrative assistance from the reference laboratory in South Africa. Importantly, this report indicates that the price of US$ 30.31/test is only 12% cheaper than the discounted NucliSENS EasyQ HIV-1 v1.2 previously reported for a reference laboratory in South Africa . IrrespeThe duration of the training was brief (two weeks), since the operator had received previous training in molecular biology techniques, which significantly alleviated the learning curve. At the end of the training, it took 2.5 hours for the laboratory technician to process one test run of 19 samples, which is the average time required for this type of assay.The high level of viral suppression rate at month 36 on treatment of PASER patients (93%) was achieved in the absence of any sensitive virological monitoring and underscores the excellent clinical practice provided by the site. In addition, the high percentage of patient retention (82%) and the previously reported low level of acquired HIV-DR at month 12  demonstrThis report demonstrates the feasibility of implementing a PCR-based VL assay in a less-experienced, low throughput laboratory, with the plasma VFA results comparable to those obtained in accredited reference laboratory in South Africa and in The Netherlands. Adequate performance of the VFA on DBS was however not achieved in this setting, indicating that using this type of specimen for molecular assays requires more technical proficiency as compared to liquid plasma.This study indicates that neither the VFA nor the p24 are reliable tools to identify virological failure to ART in long-term treated patients, who are mostly virologically suppressed. The routine use of affordable VL testing is largely advocated to preserve the cost-efficiency of ART programmes in resource limited settings , 43. It The good clinical outcome of treated patients at this site suggests that despite the absence of conventional virological monitoring, quality clinical care, focused on patient support for treatment adherence, contributes to limit virological failure, thereby preventing the emergence of HIV drug resistance.


First-line antiretroviral therapy (ART) failure often results from the development of resistance-associated mutations (RAMs). Three patterns, including thymidine analogue mutations (TAMs), 69 Insertion (69Ins) and the Q151M complex, are associated with resistance to multiple-nucleoside reverse transcriptase inhibitors (NRTIs) and may compromise treatment options for second-line ART.We investigated patterns and factors associated with multi-NRTI RAMs at first-line failure in patients from The TREAT Asia Studies to Evaluate Resistance – Monitoring study (TASER-M), and evaluated their impact on virological responses at 12 months after switching to second-line ART. RAMs were compared with the IAS-USA 2013 mutations list. We defined multi-NRTI RAMs as the presence of either Q151M; 69Ins; ≥2 TAMs; or M184V+≥1 TAM. Virological suppression was defined as viral load (VL) <400 copies/ml at 12 months from switch to second-line. Logistic regression was used to analyze (1) factors associated with multi-NRTI RAMs at first-line failure and (2) factors associated with virological suppression after 12 months on second-line.p=0.004) and ART duration >2 years . Among 87/105 patients with available VL at 12 months after switch to second-line ART, virological suppression was achieved in 85%. The median genotypic susceptibility score (GSS) for the second-line regimen was 2.00. Patients with ART adherence ≥95% were more likely to be virologically suppressed , p=0.001). Measures of patient resistance to second-line ART, including the GSS, were not significantly associated with virological outcome.A total of 105 patients from 10 sites in Thailand, Hong Kong, Indonesia, Malaysia and Philippines were included. There were 97/105 (92%) patients harbouring ≥1 RAMs at first-line failure, 39/105 with multi-NRTI RAMs: six with Q151M; 24 with ≥2 TAMs; and 32 with M184V+≥1 TAM. Factors associated with multi-NRTI RAMs were CD4 ≤200 cells/µL at genotyping (OR=4.43, 95% CI [1.59–12.37], Multi-NRTI RAMs at first-line failure were associated with low CD4 level and longer duration of ART. With many patients switching to highly susceptible regimens, good adherence was still crucial in achieving virological response. This emphasizes the importance of continued adherence counselling well into second-line therapy. In resource-limited settings, first-line antiretroviral therapy (ART) for HIV-positive patients consists of nucleoside and non-nucleoside reverse transcriptase inhibitors (NRTIs and NNRTIs). Protease inhibitors (PIs) are usually the core component of second-line therapy . First-lThree drug resistance patterns including thymidine analogue-associated mutations (TAMs), 69 Insertion (69Ins) and Q151M complex are associated with multi-NRTI drug resistance . TAMs, oThe presence of TAMs, 69Ins, Q151M and M184V at first-line ART failure can compromise treatment options for second-line therapy. A case study on one patient from Sierra Leone treated with stavudine, lamivudine and nevirapine reported extensive mutations to all NRTIs and NNRTIs . A studypol genotype (FASTA file) within six months prior to switch to second-line. First-line failures were obtained from the ART stop reasons reported in the database, and included treatment failure, clinical progression or hospitalization.Patients were selected from The Therapeutics, Research, Education and AIDS Training in Asia (TREAT Asia) Studies to Evaluate Resistance – Monitoring Study (TASER-M). TASER-M began recruitment in 2007 and included 12 clinical sites in Thailand, Hong Kong, Malaysia, Philippines and Indonesia. Patients were either enrolled in TASER-M as treatment naïve initiating first-line ART or treatment-experienced switching to second-line ART due to failure . PatientData transfers from clinical sites were aggregated at The Kirby Institute, UNSW Australia , Sydney, Australia. Ethics approvals were obtained from UNSW Australia Ethics Committee and institutional review boards at the participating clinical sites and coordinating centre . Written informed consent was obtained from participants prior to enrolment.Genotyping was performed at laboratories certified by TREAT Asia Quality Assessment Scheme (TAQAS). TAQAS was developed as a quality assessment program aimed at standardizing HIV genotyping among laboratories in the Asia-Pacific region and Africa , 18. FASThis study consists two analyses:Variables adjusted in the regression model were age, sex, mode of HIV exposure, failing NRTI backbone categorized as lamivudine+stavudine, lamivudine+zidovudine and other combinations, viral load (VL) and CD4 count measured within six months prior to switch to second-line ART closest to the date of genotyping, time since ART initiation, prior AIDS-defining illnesses at time of genotyping and HIV-1 subtype. Sensitivity analysis was performed by including K65R in our definition of multi-NRTI RAMs as this mutation is associated with reduced virological response to tenofovir, didanosine, abacavir and stavudine .We analyzed the effects of second-line treatment susceptibility on VL suppression by using the genotypic susceptibility score (GSS) for the second-line regimen, calculated based on the drug resistance scores extracted from the Stanford HIVdb. Each antiretroviral drug (ARV) was assigned a score according to the five-level Stanford HIVdb interpretation: 1.00, 0.75, 0.50, 0.25 and 0.00 for susceptible, potential low-level resistance, low-level resistance, intermediate resistance and high-level resistance, respectively . The GSSFor sensitivity analyses, we replaced GSS with other resistance scores such as the sum of the actual resistance scores from the Stanford HIVdb, continuous genotypic susceptibility score (cGSS) , optimizp<0.10 in the univariate model were chosen for inclusion in the multivariate model. Covariates with p<0.05 in the final multivariate model were considered statistically significant.Logistic regression models were used in both analysis (1) and (2). In analysis (2), we attempted to categorize GSS as either above or below the optimal cut-off point calculated from the Youden Index . The YouAll data management and statistical analyses were performed using SAS software version 9.3  and STATA software version 12.1 .Of 2023 TASER-M patients, 105 patients fit inclusion criteria and were included in the analysis. Of these, 28 patients were enrolled in the treatment-naïve group and 77 were enrolled in the treatment-experienced group. There were 97 patients 92%) with ≥1 RAMs at first-line failure: 90 patients with NRTI RAMs, 92 with NNRTI RAMs and two with PI-major RAMs. A total of 86 patients (82%) had multi-class mutations. % with ≥1p<0.10 were failing NRTI backbone (p=0.035), VL (p=0.093), CD4 count (p=0.009), time since ART initiation (p<0.001) and previous AIDS-defining illnesses (p=0.005). In the multivariate model, low CD4 count of ≤200 cells/µL was associated with the presence of multi-NRTI RAMs  compared to higher CD4 count at genotyping. Patients who have been on ART for more than two years were more likely to harbour these mutations compared to those who have been treated for a shorter period of time . All other covariates were not significant in the final multivariate model at p<0.05; however, VL and previous AIDS-defining illnesses were borderline significant at p=0.060 and p=0.059, respectively. When K65R was included in the sensitivity analysis, a total of 46/105 (44%) patients were classified as harbouring multi-NRTI RAMs. Variables remaining significant in the final multivariate model were CD4 ≤200 cells/µL  compared to CD4 >200 cells/µL, and time on ART >2 years , compared to ≤2 years. As the results of this sensitivity analysis are similar to those presented in In the univariate analysis , variablOut of 105 patients, 87 had VL testing at 12 months after switching to second-line therapy. The proportion of patients who achieved VL suppression was 74/87 (85%). Second-line regimen consisted mainly of NRTI+PI (98%). The NRTI backbones were lamivudine+tenofovir (32%), zidovudine+tenofovir (16%) and other combinations (52%). The proportions of individual NRTI combinations included in the other category were <10% each. The second-line PI component consisted of boosted lopinavir (86%), atazanavir (7%), indinavir (6%) and darunavir (1%).The GSS was calculated from the Stanford HIVdb resistance interpretations for the second-line regimen. GSS had a range of 1.00–4.00, with a median of 2.00 and IQR (1.50–3.00). The GSS cut-off point obtained from the Youden Index was 1.25 . Using tp=0.001). The GSS variable was not significantly associated with VL suppression in both univariate and multivariate models.In both the univariate and multivariate analyses , the onlp=0.868). Further analyses were also performed using cGSS, OBS, and multi-NRTI RAMs binary covariate. These variables were also not associated with virological outcome . The wGSS had a median of 2.6 with IQR (2.10–3.60). There was no difference between wGSS scores above and below the median in their association with VL suppression .In the sensitivity analyses, we first replaced the GSS with the sum of the actual resistance scores for each ARV obtained directly from the Stanford HIVdb. The resistance scores ranged from −20 to 200, with a median of 45 and IQR (0–75). The resistance scores were not significantly associated with VL suppression . Multi-NRTI RAMs were found in 37% of the patients. Six patients had Q151M, 24 had ≥2 TAMs and 32 had M184V+≥1 TAM. Low CD4 count and longer duration on ART were linked to the presence of multi-NRTI RAMs at first-line failure. However, these mutations and the resulting treatment susceptibility did not have a significant effect on the virological response of second-line therapy. Adherence level was the only significant factor associated with achieving VL suppression 12 months after switching to second-line ART.A high prevalence of drug resistance mutations is commonly seen in patients failing first-line ART. Our findings were consistent with previous reports which showed that NNRTI RAMS were the most common, followed by NRTI and PI RAMs , 29, 30.p=0.060, it is known that viraemia can be associated with the accumulation of drug resistance mutations, in particular, NRTI RAMs. A study in Africa has found that NRTI cross-resistance was associated with higher VL in the univariate analysis and was borderline significant (p=0.052) in the adjusted model [Results from analysis (1), which investigated factors associated with multi-NRTI RAMs at first-line failure, indicate that the two significant factors were low CD4 count and longer time on ART. Although our study found that VL was only marginally significant at ed model . Anothered model . VL monied model , 34. TheThe association of low CD4 level with RAMs was consistent with a Chinese study  which alThe second-line virological outcome analysis  showed The Department of Health and Human Services Antiretroviral Guidelines for Adults and Adolescents (February 2013) recommend that after first-line failure has occurred, the new ARV regimen should contain at least two fully active drugs . The susAmong the TASER-M sites, selection of the second-line regimen after first-line ART failure varied across different clinical settings. Sites with relatively quick sequencing turn-around time were able to select drugs and start patients on second-line regimen after the genotypic resistance profile had been completed. However, at some sites, patients were switched to second-line ART prior to completion of sequence amplification, either due to the long sequencing time, or limited options available for second-line ART. The WHO guidelines recommend selection of second-line ARVs based on the combination of ARVs received during first-line therapy, rather than the presence of drug resistance results showing mutations at failure . BecauseThe limitations of the study were the limited sample sizes of patients who failed first-line ART and those with VL available at 12 months after switching to second-line. We were also unable to adjust for site or country differences due to relatively few data points in some of the categories. Adherence to first-line ART was not available for the majority of patients. This is because more than 70% of patients included in the analysis were enrolled in TASER-M in the treatment-experienced group. For this group, prospective adherence data was collected at time of switch to second-line onwards. Since no retrospective self-reported adherence was available, it was not possible to determine the confounding effects of first-line ART adherence on the presence of multi-NRTI RAMs.TASER-M patients failing first-line ART reported high prevalence of multi-NRTI RAMs with many switching from a NNRTI-based regimen to a PI-based regimen. Although multi-NRTI RAMs can compromise treatment choices of second-line therapy, it has been shown that virological suppression can be achieved if high level of adherence was maintained. ART adherence proved to be a significant contributor to the success of second-line therapy in this patient group who mostly switched to a highly susceptible treatment regimen. This emphasizes the importance of appropriate adherence counselling and intervention which should be continued well beyond first-line therapy. Further studies with larger sample sizes and longer follow-up time are required to determine if high ART adherence level can maintain its positive effects over time in the presence of drug resistance mutations during second-line therapy.3TC+d4T+NVP:KJ868870,KJ868873,KJ868881,KJ868888,KJ868889,KJ868890,KJ868891,KJ868892,KJ868894,KJ868901,KJ868902,KJ868908,KJ868909,KJ868912,KJ868914,KJ868915,KJ868917,KJ868919,KJ868921,KJ868922,KJ868927,KJ868928,KJ868929,KJ868932,KJ868933,KJ868934,KJ868935,KJ868940,KJ868947,KJ868950,KJ868951,KJ868975,KJ868978,KJ868986,KJ868993,KJ868994,KJ868995,KJ868996,KJ868997,KJ868999,KJ869006,KJ869007,KJ869013,KJ869014,KJ869017,KJ869019,KJ869020,KJ869022,KJ869024,KJ869026,KJ869027,KJ869032,KJ869033,KJ869034,KJ869037,KJ869038,KJ869039,KJ869040,KJ869045,KJ869052,KJ869055,KJ869056;3TC+AZT+NVP:KJ868850,KJ868864,KJ868871,KJ868882,KJ868883,KJ868885,KJ868898,KJ868900,KJ868904,KJ868905,KJ868916,KJ868918,KJ868920,KJ868930,KJ868938,KJ868939,KJ868943,KJ868945,KJ868946,KJ868948,KJ868949,KJ868952,KJ868955,KJ868969,KJ868976,KJ868987,KJ868988,KJ868990,KJ869003,KJ869005,KJ869009,KJ869010,KJ869021,KJ869023,KJ869025,KJ869035,KJ869043,KJ869044,KJ869048,KJ869050,KJ869051,KJ869053,KJ869054,KJ869057;3TC+d4T+EFV:KJ868849,KJ868857,KJ868862,KJ868868,KJ868899,KJ868907,KJ868910,KJ868911,KJ868925,KJ868936,KJ868937,KJ868942,KJ868954,KJ868962,KJ868967,KJ868973,KJ869004,KJ869012,KJ869015,KJ869016,KJ869030,KJ869041,KJ869042,KJ869047;3TC+AZT+EFV:KJ868851,KJ868865,KJ868866,KJ868887,KJ868893,KJ868896,KJ868897,KJ868924,KJ868944,KJ868956,KJ868970,KJ868971,KJ868992,KJ868998,KJ869001,KJ869002,KJ869029,KJ869049;3TC+TDF+EFV:KJ868875,KJ868878,KJ868879,KJ868880,KJ868884,KJ868886,KJ868913,KJ868980,KJ868983,KJ868984,KJ868985,KJ868989,KJ868991,KJ869018;3TC+ABC+EFV:KJ868853,KJ868854,KJ868856,KJ868859,KJ868860,KJ868861,KJ868958,KJ868959,KJ868961,KJ868964,KJ868965,KJ868966;3TC+TDF+NVP:KJ868872,KJ868874,KJ868895,KJ868923,KJ868931,KJ868977,KJ868979,KJ869000,KJ869028,KJ869036;3TC+d4T:KJ868867,KJ868926,KJ868941,KJ868972,KJ869031,KJ869046;FTC+TDF+EFV:KJ868876,KJ868877,KJ868906,KJ868981,KJ868982,KJ869011;3TC+AZT:KJ868852,KJ868863,KJ868957,KJ868968;3TC+ABC+NVP:KJ868855,KJ868960;3TC+AZT+TDF+NVP:KJ868903,KJ869008;3TC+DDI+EFV:KJ868858,KJ868963;AZT+DDI+NVP:KJ868869,KJ868974;FTC+TDF:KJ868953,KJ869058.The Genbank accession numbers obtained from the sequences used in this study are KJ868849-KJ869058. The accession numbers categorized according to the failing first-line NRTI+NNRTI combination are as follows: