Treatment with second line ART is initiated when the first line therapy fails. There is less experience with the immunologic response for second-line ART for adults. Hence this study was done to find out the immunological response after initiation of second line ART by analysis of CD4 counts.This retrospective study was conducted in a tertiary level hospital attached to a medical college that caters to a large number of HIV positive patients. The study population for this analysis included all HIV positive individuals who were undergoing second line ART treatment.The data was collected using semi-structured pre-tested proforma from the hospital records of HIV positive individuals. The immunological response after the initiation of second line ART was analyzed using the CD4 cell counts taken at intervals of 3 months and 6 months after the initiation.Out of the 32 patients studied, 27(84.4%) were males and only 5(15.6%) were females. Mean age of the patients was 40.56±6.78 years. The mean CD4 value at initiation was 152.35±142.89cells/μL, which significantly increased to 324.43 ±163.65cells /μLby 3 months after initiation  and to 348.21± 253.57cells /μLby 6 months after initiation.Around 91.3% of patients had a baseline CD4 T cell count <350 cells L-6. After 3 months of therapy, 65.2% of patients and after 6 months 46.2% had a baseline CD4 T cell count <350 cells /μL.The mean weight at initiation was 50.548± 11.37 kg, which very significantly increased to 53.30± 11.1 kg by 3 months of therapy  and to 54.63 ± 10.29kg at the end of 6 months.The CD4 counts increase very significantly within the first 3 months of initiation of second line therapy.The rise in CD4 count between 3 months and 6 months is not as statistically significant as the earlier one. Also, there is significant gain in weight within 6 months of initiation of second line therapy.


Viral reservoirs that persist in HIV-1 infected individuals on antiretroviral therapy (ART) are the major obstacle to viral eradication. The identification and definition of viral reservoirs in patients on ART is needed in order to understand viral persistence and achieve the goal of viral eradication. We examined whether analysis of episomal HIV-1 genomes provided the means to characterize virus that persists during ART and whether it could reveal the virus that contributes to treatment failure in patients on ART. For six individuals in which virus replication was highly suppressed for at least 20 months, proviral and episomal genomes present just prior to rebound were phylogenetically compared to RNA genomes of rebounding virus after therapy interruption. Episomal envelope sequences, but not proviral envelope sequences, were highly similar to sequences in rebounding virus. Since episomes are products of recent infections, the phylogenetic relationships support the conclusion that viral rebound originated from a cryptic viral reservoir. To evaluate whether the reservoir revealed by episomal sequence analysis was of clinical relevance, we examined whether episomal sequences define a viral population that contributes to virologic failure in individuals receiving the CCR5 antagonist, Vicriviroc. Episomal envelope sequences at or near baseline predicted treatment failure due to the presence of X4 or D/M  viral variants. In patients that did not harbor X4 or D/M viruses, the basis for Vicriviroc treatment failure was indeterminate. Although these samples were obtained from viremic patients, the assay would be applicable to a large percentage of aviremic patients, based on previous studies. Summarily, the results support the use of episomal HIV-1 as an additional or alternative approach to traditional assays to characterize virus that is maintained during long-term, suppressive ART. Infection by HIV-1 and the related effects on human health continue to be a major problem throughout the world. Since the early 1980's, more than 25 million people have died from AIDS and the only treatment option for infected individuals is likely to be life-long treatment with a combination of antiviral drugs. While antiviral drug therapy can reduce viral replication to levels that are undetectable by currently used assays, there is a rapid recrudescence of viremia upon interruption of therapy. This indicates that there are viral reservoirs, undetectable by conventional diagnostic assays that sustain the virus in the face of ART. We have developed an alternative or additional approach to study cryptic viral replication based on episomal HIV-1 genomes. Although HIV-1 episomes are not suitable substrates for integration and thus are dead-end products in the viral life cycle, episomal HIV-1 genomes are useful surrogate markers of viral replication since they are labile and indicative of recent infection events. Here we have used episomal HIV-1 analysis to study the reservoir that fuels viral rebound during treatment interruption and to demonstrate the utility of this approach in guiding the clinical treatment of infected individuals. Despite great advances in the treatment of HIV infection and disease, therapeutic eradication of viral infection is currently not possible. Individuals who respond well to ART and maximally suppress virus replication based on standard assays, harbor viral reservoirs that fuel a rapid rebound in viral replication upon interruption of antiviral treatment. Although the biological relevance of different reservoirs is subject to ongoing debate, there is evidence to support the existence of at least two distinct viral reservoirs that persist during effective, prolonged suppression of HIV-1 replication. One is a latent reservoir that is comprised of lymphocytes that were infected and subsequently reverted to a quiescent state in vitro experiments have suggested that 2-LTR circles are stable in vivo analysis has demonstrated their labile nature, which supports the use of episomal HIV-1 cDNAs as a valid surrogate marker of recently infected cells In addition to latently infected cells, there is mounting evidence to support the existence of a reservoir of low-level ongoing replication in patients on HAART. Evolution of viral envelope sequences in the context of HAART has been documented for HIV-1 infected patients Other groups have also focused on defining the effects of therapy intensification on residual viremia using raltegravir, yielding different conclusions In patients who suppress virus replication to levels below the limit of detection of standard assays, low-level ongoing replication can be detected through the analysis of the HIV-1 episomes. Therefore, in highly suppressed individuals who have undetectable virus in plasma, analysis via episomal HIV-1 cDNA provides the means to monitor and define the nature of actively replicating virus that persists in the face of ART.env) sequences to probe the nature of actively replicating virus that persists during ART and demonstrate the utility of episomal sequence analysis in guiding clinical treatment of infected individuals. Envelope sequences were compared to define the phylogenetic relationships between episomal and proviral genomes prior to rebound in patients on ART to RNA genomes of virus that emerged upon therapy interruption. Additionally, episomal env genes were analyzed from a group of patients undergoing salvage therapy with Vicriviroc (VCV) in order to predict the virologic response to this CCR5 antagonist.Here we have focused on analyzing episomal envelope  and neighbor-joining phylogenetic trees were constructed using the Kimura two-parameter model as implemented in the MEGA Version 4 program env sequences in the rebounding virus were highly similar to episomal sequences present just prior to treatment interruption, but not to proviral sequences obtained from the same sampling time , we selected fourteen patients who experienced protocol-defined virologic failure. Virologic failure for this protocol is defined as failure to experience a log reduction in viral replication from baseline values at or after week 16. Episomal env sequences were generated by PCR from lymphocyte DNA isolated from baseline through week 48 after initiation of VCV therapy. Full-length env genes were amplified from episomal cDNA and cloned into an expression vector. Pseudotyped viruses were generated with patient-derived envelopes and an NL4-3 proviral clone expressing luciferase in place of envelope. Co-receptor dependence was then gauged in an infection assay utilizing U87-based indicator cells. Amplification of full-length env genes was problematic for a small number of samples. Therefore, in order to derive secondary information on virus genotypes from these samples, short fragments spanning the V3 region of env were amplified and sequenced. Tropism predictions were determined using the Geno2Pheno algorithm with a false positive rate of five and ten percent as well as coupled 11/25 and net charge rules. The phenotypic/genotypic data is summarized in env fragments from 74 of 91 (81.3%) PBMC samples. The Trofile assay and phenotypic assay based on episomal env sequences were concordant at all timepoints for 8 of the 14 patients. In these individuals, R5 viruses persisted at all time points based on both phenotypic assays and virologic failure could not be explained by outgrowth of X4 virus .Here we have applied episomal analysis to characterize virus that rebounds upon antiviral treatment interruption. In an analysis of six patients, there was a rapid rebound in plasma viral RNA when therapy was interrupted after a prolonged period of sustained suppression of viral replication. Phylogenetic relationships based on C2 to V4 + T cells and low-level active viral replication have been implicated in the rapid resurgence of viremia following treatment interruption in vivo evidence that residual viral replication persists in patients on highly suppressive therapies and that viral rebound is fueled by this cryptic replication. While the present study supports the hypothesis that HIV-1 rebounds from a reservoir in which replication is ongoing, a recent study by Joos et al. concluded that rebounding virus is derived from the activation of latently infected cells + T cells from the same patient revealed that most sequences generated from plasma were distinct from those in CD4+ T cells, which is consistent with previous studies + T cells. Our analysis focused on genomes that were associated with PBLs, and given the genetic similarity between episomes and rebounding virus, episomal genomes may be derived from a reservoir that is distinct from the source of residual viremia. Although it would be potentially informative to compare env sequences that we generated to env present in patient plasma prior to rebound, it was not possible due to the archival nature of our samples. Additionally, one might predict that the sources of residual viremia and episomes are distinct based on the results of therapy intensification with raltegravir in which episomes increase transiently Previous studies aimed at revealing the origin of virus that rebounds upon therapy interruption have reached different conclusions. Both activation of latently infected CD4env genes from sixty-four percent of the samples and of the samples in which viral plasma RNA copies were below 100, we could amplify env fifty percent of the time. Thus, targeting episomal genomes provides a feasible approach to the study of residual virus replication.To further support the methodology and demonstrate that episomal analysis has the potential to provide information with which to guide clinical decisions regarding patient therapy, we analyzed samples from individuals undergoing salvage treatment with the CCR5 antagonist VCV. Although these patients had varying levels of viremia, we wanted to determine whether episomal HIV-1 cDNA analysis could provide information on the effects of adding VCV to a regimen that was no longer effective in suppressing viral replication. We were able to generate full-length in vitro and in vivoenv genes indicated the presence of dual or X4 viruses while undergoing therapy. For these patients, it is highly likely that failure to respond to VCV therapy was due to the presence of viral variants that were unaffected by the CCR5 antagonist. In all cases, D/M or X4 viruses were present at early time points of the trial, so it is unlikely that selective pressure from VCV therapy resulted in a replacement of R5 with X4 virus. We observed very little variability in the primary V3 loop sequences of the patient samples, so the basis for VCV therapy failure is unclear in the subset of patients that did not harbor X4 or D/M viruses. These results are consistent with previous work studying the same group of patients env sequences that are peripheral to the V3 loop or involve other, as yet undefined determinants. Alternatively, patients who experienced virologic failure with R5 virus may not have been fully compliant with the regimen.Although VCV is considered to be a potent inhibitor of R5 HIV-1 env genes to determine HIV-1 tropism from plasma virions env sequences were highly concordant. The high degree of correlation between the two assays indicates that episomes are not archival, but are evolving and derived from a reservoir of actively replicating virus. While the majority of samples analyzed were from time points in which viral plasma RNA was greater than the limit of detection, we would expect episomal analysis to be a viable approach in patients that suppress virus replication to levels below the limit of detection of standard assays. Having a method to determine viral co-receptor usage in patients with highly suppressed viremia would be advantageous for patient management. It is common for patients to experience toxicity from the drug regimens in use and defining the virus co-receptor dependence would allow a clinician to switch therapy to a better tolerated drug such as a CCR5 antagonist.Since most of the samples were analyzed while patients were experiencing viremic episodes, we were able to compare our assay data to phenotypic data previously obtained via the Trofile assay env from episomes also targets 1-LTR circles that are approximately ten times more abundant than 2-LTR circles resulting in a much greater sensitivity of detection Previous studies have shown that 2- LTR episomes are detectable in a high percentage of patients on long-term highly suppressive antiviral therapy After obtaining written informed consent, cells and plasma were collected by phlebotomy from HIV-1 infected volunteers at the NIAID and research centers involved in AIDS Clinical Trials Group (ACTG) protocol A5211. Eligible participants were age 18 or older and either voluntarily suspended ART or chose to receive a new antiviral compound in the context of salvage therapy. The study was reviewed and approved by both the NIAID and the University of Massachusetts's institutional review boards. Human experimentation guidelines of the US Department of Health and Human Services were followed in the conduct of this research.Six HIV-1 infected individuals who were receiving effective antiviral treatment for approximately 2 years and subsequently suspended therapy were studied. None of the patients exhibited any detectable viremia prior to suspension of therapy, based on plasma viral RNA measurements (limit of detection 50 or 500 copies per milliliter). PBMCs were obtained prior to, and at the time of, therapy interruption via leukapheresis by Ficoll-Hypaque density gradient centrifugation. One milliliter of plasma was obtained 3, 4 or 8 weeks after suspension of therapy at which time viral rebound had occurred. The study was approved by the NIAID's institutional review board and informed consent was obtained in writing from study participants. Human experimentation guidelines of the US Department of Health and Human Services were followed in the conduct of this research.Low molecular weight DNA was purified using a modified plasmid isolation procedure (Qiagen) and chromosomal DNA was subsequently recovered from the SDS-precipitate by extraction with DNazol (Invitrogen). Cryopreserved PBMCs were collected by centrifugation at 4 KRPM for 5 min and DNA was purified according to the manufacturer's modified plasmid protocol for low copy number samples. SDS-precipitates were then thoroughly resuspended in 900 µL DNazol at 50°C and DNA was precipitated by the addition of 500 µL 100% ethanol. DNA was pelleted by centrifugation at 12 KRPM for 8 min, washed in 1.4 ml 70% ethanol and resuspended in 50 µL 8 mM NaOH. After resuspension, lysates were neutralized by the addition of 50 µL 10 mM Tris-HCl, pH 8.0. Virions were pelleted from thawed plasma at 14 KRPM for 2 h at 4°C and RNA was purified using an RNAeasy kit according to the manufacturer's protocol (Qiagen) and eluted in 50 µL water.5905GGGTCACAGTCTATTATGGGGTA and LA17/r, T332CTCCTTCTAGCCTCCGCTAGTCAA) or proviral-specific primers  and LA9/f, A6015CATGCCTGTGTACCCACAGA) and Jumpstart RedAccuTaq (Sigma) in a reaction containing 1X buffer, 250 µM dNTPs, 0.5 µM primers and 1.5 units polymerase. PCR conditions consisted of a denaturation step at 95°C for 1 min followed by 22 cycles of 15 sec at 95°C, 30 sec at 58°C and 5 min at 68°C. Second round products were generated using 40 cycles of amplification as above except with an extension time of 1 min and LA11/f (C6546ACAGTACAATGTACACATGGA) and Env7/r (A7128GGGGCATACATTGCTTTTCCTA) envelope primers. 8 µL viral RNA was converted to cDNA using Superscript RT (Invitrogen) according to the manufacturer's protocol. cDNA was amplified by limiting dilution pcr using the same conditions as the second round reactions above. PCR products were purified and used directly for sequencing with the LA11/f primer. Primers were numbered according to the HIV-1LAI consensus (K02013).A portion of the envelope gene spanning the C2 to V4 region was amplified by nested PCR using limiting dilution replicates of the first round products. First round products were generated using episomal-specific  and phylogenetic relationships were generated using the neighbor-joining method included in the MEGA 4 software package with a Kimura-2 parameters distance matrix. Confidence limits for individual branches were estimated by bootstrap re-sampling of the neighbor-joining trees (1000 replicates).Nucleotide sequences corresponding to the C2 to V4 regions of The study was conducted using archival patient samples that had originally been collected under AIDS Clinical Trials Group (ACTG) protocol A5211, a phase 2b study of the investigational CCR5 antagonist Vicriviroc (VCV). The study was approved by the University of Massachusetts's institutional review board and informed consent was obtained in writing from study participants. Human experimentation guidelines of the US Department of Health and Human Services were followed in the conduct of this research. Ninety-one cryopreserved PBMCs were obtained from fourteen HIV-1-infected, treatment-experienced patients, who experienced protocol-defined virologic failure. PBMCs were collected during the period from study entry through week 48 . PBMCs at study entry (week 0) were available only for six out of fourteen patients.6 cells). PBMCs were collected by centrifugation at 4 KRPM for 5 min. Cell pellets were resuspended in buffer P1 and extrachromosomal DNA was purified using a QIAprep spin miniprep kit (Qiagen) using the modification for the isolation of low copy number plasmids as recommended by the manufacturer. Final elution of the purified DNA was in 120 µl of elution buffer preheated at 70°C.Nucleic acid extractions were performed from patients' cryopreserved PBMCs samples  were amplified from 20–30 µl of extrachromosomal DNA by nested PCR using the Phusion Hot Start High-Fidelity DNA Polymerase . Amplifications were done in a 50-µl reaction containing 1 x Phusion HF Buffer, 200 µM deoxynucleotide triphosphates, 0.5 µM primers, and 1 U of Phusion Hot Start DNA Polymerase. Outer primers were envA  and LA17 , and inner primers were envB  and envM . Primers were numbered according to the HIV-1LAI consensus (K02013).HIV-1 full-length After an initial denaturation at 98°C for 1 min, first round products were generated by cycling 25 times at 98°C for 15 s, 60°C for 30 s, and 72°C for 3.30 min, followed by a final extension at 72°C for 5 min. Two microliters from the first round were then used for nested PCR which was performed with an initial denaturation step of 98°C for 1 min, followed by 25–30 cycles of 98°C for 15 s, 63°C for 30 s, and 72°C for 1.45 min, and a final extension of 72°C for 5 min.env.PCR product DNA was purified over a column using QIAquick PCR Product Purification Kit (Qiagen) and cloned into the pcDNA3.1D/V5-His-TOPO expression vector . Plasmids were isolated with Qiagen miniprep purification kit (Qiagen) and were digested with HindIII and XhoI to confirm insertion of full-length env fragment, encompassing V3 region was amplified within episomal cDNA by nested PCR using envA/LA17 as outer primers and LA12  and LA13  as inner primers. The PCR products were subjected to clonal analysis using StrataClone Blunt PCR Cloning Kit (Stratagene).Short env with LA12 primer using an ABI/Prism 377 DNA sequencer (Applied Biosystems). Sequences were edited and aligned with MacVector software (v.10.0.2). To predict HIV-1 tropism from the V3 genotype we used bioinformatic tool Geno2pheno (http://coreceptor.bioinf.mpi-sb.mpg.de/cgi-bin/coreceptor.pl) with a false positive rate of 5% and 10%, as well as, coupled 11/25 and net charge rules.The V3 region was sequenced from cloned env sequence. Virus particles released into the supernatant after 48 hours were used to infect indicator cells bearing CD4 and CCR5 (U87.CD4.CCR5 cells) or CD4 and CXCR4 (U87.CD4.CXCR4 cells) . Recombinant viruses with known co-receptor usage  were used as controls in each assay. 293T cells transfected with empty vector plasmid were used as negative control. The ability of pseudoviruses to infect target cells was assessed by measuring the luciferase activity in lysed cells  72 h postinoculation with Bright-Glo Luciferase Assay System (Promega). Tested virus was assigned a particular tropism if infection of cells expressing the relevant co-receptor resulted in an RLU reading above the background. The specificity of low-level positive luminescent signals was confirmed using co-receptor antagonists. Bicyclam JM-2987  was used to block CXCR4-mediated entry and TAK-779 to block CCR5-mediated entry . We used three concentrations (1∶3 dilutions) for each component, starting at 1.25 µg/ml for AMD-3100 and 1.5 µg/ml for TAK-779.The phenotype of HIV-1 co-receptor usage was determined using a single-cycle recombinant virus entry assay. Human embryonic kidney 293T cells were co-transfected using Lipofectamine 2000 (Invitrogen) with pNL43-Δenv-Luc vector DNA and expressing vector containing the amplified full-length Figure S1LAI provirus per cell, were combined with a constant 8×106 uninfected PBLs. DNA was purified as is described in the Fractionation of PBL DNA results in minimal proviral contamination of extrachromosomal supernatants. Variable numbers of 8E5 cells, which have a single HIV-1 (3.00 MB TIF)Click here for additional data file.Figure S2LAI and HIV-1ADA laboratory strains (black stars) are included for reference.Phylogenetic tree based on the C2-V4 regions of envelope for patient 1 undergoing interruption of antiviral treatment. Phylogenetic relationships were estimated using the neighbor-joining method to generate best tree with genetic distances for episomal (yellow triangle) and proviral (blue circle) envelope sequences derived at therapy interruption to plasma viral RNA (orange square) envelope sequences obtained several weeks after rebound. Envelope sequences from HIV-1(0.21 MB TIF)Click here for additional data file.Figure S3LAI and HIV-1ADA laboratory strains (black stars) are included for reference.Phylogenetic tree based on the C2-V4 regions of envelope for patient 2 undergoing interruption of antiviral treatment. Phylogenetic relationships were estimated using the neighbor-joining method to generate best tree with genetic distances for episomal (yellow triangle) and proviral (blue circle) envelope sequences derived at therapy interruption to plasma viral RNA (orange square) envelope sequences obtained several weeks after rebound. Envelope sequences from HIV-1(0.32 MB TIF)Click here for additional data file.Figure S4LAI and HIV-1ADA laboratory strains (black stars) are included for reference.Phylogenetic tree based on the C2-V4 regions of envelope for patient 3 undergoing interruption of antiviral treatment. Phylogenetic relationships were estimated using the neighbor-joining method to generate best tree with genetic distances for episomal (yellow triangle) and proviral (blue circle) envelope sequences derived at therapy interruption to plasma viral RNA (orange square) envelope sequences obtained several weeks after rebound. Envelope sequences from HIV-1(0.34 MB TIF)Click here for additional data file.Figure S5LAI and HIV-1ADA laboratory strains (black stars) are included for reference.Phylogenetic tree based on the C2-V4 regions of envelope for patient 4 undergoing interruption of antiviral treatment. Phylogenetic relationships were estimated using the neighbor-joining method to generate best tree with genetic distances for episomal (yellow triangle) and proviral (blue circle) envelope sequences derived at therapy interruption to plasma viral RNA (orange square) envelope sequences obtained several weeks after rebound. Envelope sequences from HIV-1(0.31 MB TIF)Click here for additional data file.Figure S6LAI and HIV-1ADA laboratory strains (black stars) are included for reference.Phylogenetic tree based on the C2-V4 regions of envelope for patient 5 undergoing interruption of antiviral treatment. Phylogenetic relationships were estimated using the neighbor-joining method to generate best tree with genetic distances for episomal (yellow triangle) and proviral (blue circle) envelope sequences derived at therapy interruption to plasma viral RNA (orange square) envelope sequences obtained several weeks after rebound. Envelope sequences from HIV-1(0.21 MB TIF)Click here for additional data file.Figure S7LAI and HIV-1ADA laboratory strains (black stars) are included for reference.Phylogenetic tree based on the C2-V4 regions of envelope for patient 6 undergoing interruption of antiviral treatment. Phylogenetic relationships were estimated using the neighbor-joining method to generate best tree with genetic distances for episomal (yellow triangle) and proviral (blue circle) envelope sequences derived at therapy interruption to plasma viral RNA (orange square) envelope sequences obtained several weeks after rebound. Envelope sequences from HIV-1(0.26 MB TIF)Click here for additional data file.Table S1Tropism assignments based on HIV-1 env sequences derived from episomal cDNA and plasma viral RNA from patients with concordant results between the two assays.(0.08 MB DOC)Click here for additional data file.Table S2Co-receptor use at baseline as determined by the enhanced-sensitivity Trofile assay.(0.04 MB DOC)Click here for additional data file.


Neglected data more or less recent, showed that the voltage-dependent sodium channel (Tran MKG) and the Mannose receptor  are involved in the transmission of HIV. In particular, the mannose Receptor seem essential for contamination since in a discordant couple, the uninfected male partner does not own this receptor . From these facts we have experienced a patient of 68 years, HIV + since 28 years, under active antiretroviral therapy  (another presentation at this conference), the following chemotherapy: Epivir (150mg 2daily), resveratrol (500mg 2daily), D-Mannose (1 g 3daily), Omacor (1 g 2daily). Indeed it is known that omega-3fatty acids bind the Na+ channel. Resveratrol has an anti TaT activity  and a synergy with nucleoside analogues Measurements of viral load , CD4 and CD8 and other usual blood parameters were followed during 9 months, every month.Our results show that over a period of 9 months the patient remains undetectable and CD4 count increases significantly from the one of the previous therapy (557+/-43 vs 478+/- 35). Besides, it seems that the CD4/CD8 ratio tended to increase . Moreover strong activation of the immune system almost always observed among patients HIV+ is standardized at this patient (CD3+/HLADR+ 7%) and the NK strongly increase (24%).Our results show the feasibility of HAART including Epivir, resveratrol, D-mannose, and omega3 fatty acids. In addition control of dyslipidemia induced by orthodoxic anti retrovirals should be unnecessary.These results paves the way for clinical trials with effective, low toxicity and low cost agents. Moreover the fact that mutations on the Mannose receptor and sodium channel (cell structures) are unlikely, make the appearance of resistance to such therapies unlikely.


Data on lipid profile abnormalities among patients receiving highly active antiretroviral treatment in Ethiopia are very limited. The aim of this study was to determine the prevalence of dyslipidemia and characteristics of lipid profiles among patients living with human immunodeficiency virus (HIV) using first-line highly active antiretroviral therapy (HAART) in Southern Ethiopia.This cross sectional comparative group study was conducted between March and May 2012, and included 113 HIV infected patients treated for a minimum of one year with first-line HAART regimens that included Efavirenz and Nevirapine (HAART group) and others 113 who had never received HAART (pre-HAART group). Serum lipid profiles were determined after overnight fasting and dyslipidemia was assessed according to the United State National Cholesterol Education program-III guideline. For statistical analysis Chi-square, student’s t-test, and logistic regression were used using Statistical Package for Social Sciences (SPSS) Version 20.Ninety-three (82.3%) of HAART and 87 (76.9%) pre-HAART patients had at least one laboratory abnormality, which is compatible with a diagnosis of dyslipidemia. Total cholesterol ≥ 200 mg/dl occurred in 43.4% of HAART and 15.9% pre-HAART patients (p=<0.0001), whereas HDL-cholesterol below 40 mg/dl occurred in 43.4% and in 63.7% respectively, (p=0.002). The LDL-cholesterol ≥ 130 mg/dl occurred in 33.6% of HAART and 15% pre-HAART patients (p=0.001), while triglycerides ≥ 150 mg/dl occurred in 55.8% and 31.0% respectively, (p=0.001). Receiving of HAART was significantly and positively associated with raised total cholesterol, LDL-cholesterol, and triglycerides. The adjusted odds ratio (95% CI) of HAART-treated vs. pre-HAART was 3.80 (1.34-6.55) for total cholesterol ≥ 200 mg/dl; 2.64 (1.31-5.32) for LDL- cholesterol ≥ 130 mg/dl and 2.50 (1.41-4.42) for triglycerides ≥150 mg/dl.Use of first-line antiretroviral therapy regimens that contain Efavirenz and Nevirapine were associated with raised total cholesterol, LDL-cholesterol, and triglycerides, an established atherogenic lipid profiles. Lipid profiles should be performed at baseline before commencement of antiretroviral therapy and then periodically through treatment follow-up to monitor any rising trends. In 2011, an estimated 34 million people were living with human immunodeficiency virus /acquired immunodeficiency syndrome (HIV/AIDS) worldwide; of them 22.9 million were living in Sub-Saharan Africa. About 1.2 million people were estimated to be living with HIV in Ethiopia, and proThis was a cross-sectional comparative group study. Subjects were recruited between March 2012 and May 2012 at ART clinic of Hawassa University Referral Hospital. Two groups of participants were selected for this study. The first group included HIV-infected individuals who had been receiving WHO recommended first-line HAART for a minimum of one year (HAART group). Participants used first-line HAART regimens that included nucleoside reverse transcriptase inhibitors (NRTIs): lamivudine (3TC), ZDV, or d4T, and non-nucleoside reverse transcriptase inhibitors (NNRTIs): NVP or EFV. Patients who had had their therapy regimens changed during follow-up were not included. The second group was pre-tested and confirmed HIV-positive individuals who were not yet getting HAART. All participants included were ≥18 years of age and HAART treated group to have a good ART adherence (adherence rate ≥ 95%). A good adherence is defined by missing < 2 dose of 30 doses or < 3 dose of 60 doses; and it was adopted from Ethiopian Federal Ministry of Health, HIV Care/ART follow-up form. Participants receiving lipid altering therapies, pregnant women, known diabetes mellitus patients and renal failures were excluded.+ lymphocyte count was done by using flow cytometry instrument . Blood sample was collected from each participant after a 12 hour overnight fast and centrifuged at 3000 cycles/ minute, and then serum was obtained for lipid profiles. Fasting serum TC and TG were assayed by enzymatic method . Serum HDL-c determined by enzymatic method after a selective precipitation of apolipoprotein containing lipoproteins  by phosphotungstic acid/MgCl2 . LDL-c was assessed by enzymatic method after precipitation of LDL by polyvinyl sulfate  and then LDL-c was calculated by subtracting the supernatant cholesterol fractions from the TC of the sample. And also the TC/HDL-c ratio was calculated.For all participants, data were collected on the socio-demographic informations along with body mass index, medical history including diabetes mellitus, renal failures, use of drugs that alter lipid profiles, and current use of anti-TB drugs. CD4Finally dyslipidemia was defined as TC ≥ 200 mg/dl, HDL-c < 40 mg/dl, LDL-c ≥ 130 mg/dl, TG ≥ 150 mg/dl and TC/HDL-c ratio ≥ 5 by the United States National Cholesterol Education Program, Adult Treatment Panel (NCEP-ATP) III guidelines[, we considered frequencies of LDL-c ≥130 mg/dl, which was 21% for the group of individuals with HIV/AIDS without HAART treatment[The estimation of the sample size was based on the difference between proportions and the following parameters were considered: alpha (α) = 5%, beta (β) = 10 % and power = 90%. Concerning dyslipidemiareatment; and 40.reatment. With thThe study was approved by the ethical review committee of School of Biomedical and Laboratory Sciences, University of Gondar. All individuals participated in the study provided their informed consent.Total number of participants enrolled in the study was 226, with 79 (35%) males and 147 (65%) females. Two groups were studied: the first group 113 HIV-infected patients (38 (33.6%) males, 75 (66.4%) females) were currently receiving HAART treatment; the second group 113 patients (41 (36.3%) males, 72 (63.7%) females) were pre-HAART. The age of individuals who were on HAART was 37.2 ± 8.7 while that of pre-HAART was 33.7 ±8.3 years and the age difference was significantly different (TableNinety-three 82.3%) of HAART treated and 87 (76.9%) pre-HAART patients had at least one laboratory abnormality, which is compatible with a diagnosis of dyslipidemia Figure. The pre.3% of HANo significant difference observed in lipid profile derangements between patients receiving ZDV when compared to those on d4T; and patients treated with EFV when compared to those treated with NVP Table.Multivariate analysis was adjusted for potential confounding factors such as age, sex, current history of smoking, past history of smoking, BMI, CD4+cells, current use of anti-TB drugs, and HAART treatment. Receiving of HAART was significantly and positively associated with raised TC, LDL-c, and TG. The adjusted odds ratio 95% CI) of HAART-treated vs. pre-HAART was 3.80 (1.34-6.55) for TC ≥200 mg/dl; 2.64 (1.31-5.32) for LDL-c ≥ 130 mg/dl and 2.50 (1.41-4.42) for TG ≥ 150 mg/dl. However, antiretroviral therapy was not positively associated with decreased of HDL-cholesterol  among HIV-infected patients receiving first-line HAART that include NNRTIs. We found that CD4 cell count in the HAART group was significantly different from the pre-HAART group, thus showing the effects of HAART in improving the immunological properties of the subjects.In the present study, the majority of HAART patients (82.3%) had at least one laboratory abnormality, which is compatible with a diagnosis of dyslipidemia according to NCEP-ATP III criteria. We found that the proportions of raised TC, LDL-c and TG were significantly higher in HAART group when compared to pre-HAART group. The proportion of HDL-c below 40 mg/dl was significantly higher in the pre-HAART group when compared to those on HAART, thus indicating the effects of HAART treatment in raising the lipid profiles, in addition to the restoration of health. However, there was no significant difference between treated group and non-treated group regarding TC/HDL-c ratio, this because increases in TC were counterbalanced by HDL-c improvement. The described raised lipid profiles , are atherogenic,16,17, aThe association between lipid derangements and HAART has been mainly depicted for regimens that contain PIs,18. In aThe reports from rural South India and Brazil indicated that the proportion of lipid profile derangements were higher among patients received d4T when compared to other NRTIs,22. We fComprehensive cardiovascular risk stratifications were not assessed in this study. However, the increased risk of cardiovascular diseases associated with described lipid derangements is well known-27, and Our study indicates HIV-infected patients receiving WHO-recommended first-line HAART that include Efavirenz and Nevirapine have a high prevalence of lipid profile derangements when compared to those non-treated HIV-infected patients. Uses of first-line HAART regimens are significantly associated with atherogenic lipid profiles. Therefore, the findings indicate that the need to assess lipid profiles at baseline before initiation of HAART treatment and lipid profile monitoring during therapy to monitor any rising trends. Additionally, the results also recommend implementation of well-controlled cohort studies for the evaluation of long-term effects of HAART treatment on lipid profiles.2NN: 2 non-nucleosides; AIDS: Acquired immunodeficiency syndrome; HAART: Highly active antiretroviral therapy; ZDV: Zidovudine; d4T: Stavudine; EFV: Efavirenz; ART: Antiretroviral therapy; HDL-c: HDL-cholesterol; 3TC: Lamivudine; HIV: Human immunodeficiency virus; WHO: World Health Organization; LDL-c: LDL-cholesterol; NNRTIs: Non-nucleoside reverse transcriptase inhibitors; NVP: Nevirapine; PIs: Protease inhibitors; TC: Total cholesterol; TG: Triglycerides; SPSS: Statistical package for social sciences; USNCEP-ATP: United States National Cholesterol Education Program, Adult Treatment Panel.We announce that we have no any competing interests.AT generated and designed the study, performed analysis and interpretation of data including with manuscript drafting, ZA, HA and SB assisted with the design, interpretation of data and the critical appraisal of the manuscript. All authors read and agreed the final manuscript.


Consequences of lack of viral monitoring in predicting the effects of development of HIV drug resistance mutations during HAART in resource-limited settings (RLS) is still a matter of debate.To assess, among HIV+ patients receiving their first-line HAART, prevalence of virological failure and genotypic resistance mutations pattern in a Médécins Sans Frontières/Ministry of Health programme in Busia District (Kenya).Patients with HAART treatment for ≥12 months were eligible for the study and those with HIV-RNA ≥5000 copies/ml underwent genotypic study. Total HIV-1 RNA from Dried Blood Spots was extracted using Nuclisens method.926 patients were included. Among 274 (29.6%) patients with detectable viral load, 55 (5.9%) experienced treatment failure ; 61.8% were female and 10 (18.2%) had clinical failure. Median CD4 cell count was 116 cell/mm3 (IQR: 54–189). Median HIV-RNA was 32,000 copies/ml (IQR: 11000–68000). Eighteen out of 55 (33%) samples could be sequenced on PR and RT genes, with resistance associated mutations (RAMs) in 15 out of 18 samples (83%). Among patients carrying RAMs, 12/15 (81%) harboured RAMs associated to thymidine analogues (TAMs). All of them (100%) showed M184V resistance associated mutation to lamivudine as well as NNRTI's RAMS.Virological failure rate in resource-limited settings are similar to those observed in developed countries. Resistance mutation patterns were concordant with HAART received by failing patients. Long term detectable viral load confers greater probability of developing resistance and as a consequence, making difficult to find out a cost-effective subsequent treatment regimen. Early virological failure on a nucleoside reverse transcriptase inhibitor (NRTI) and nonnucleoside reverse transcriptase inhibitor (NNRTI) regimen is associated with emergence of the M184V mutation and an NNRTI resistance mutation in approximately 50–75% of patients in resource-rich settings Continuation of a failing regimen may be associated with more complex mutation patterns, as has been observed in several studies in developing countries Sub-Saharan Africa need to face major challenges to the scaling up of treatment programs such is scarcity in financial and human resources and inadequate health-care infrastructure. Models of care must be adapted to these circumstances, accounting for the few trained health personnel, different groups of patients compared with high-income countries (increased proportions of children and women of child-bearing age affected), restricted drug availability and procurement, inadequate access to monitoring equipment, and few extra funds.Scaling up antiretroviral (ARV) therapy in RLS requires a simplified approach. Because of inadequate laboratory capacity, many programs have minimized laboratory monitoring in an effort to accelerate widespread availability of HIV treatment If one of the main issues that remain still a matter of debate in low and middle income countries is the suitability for using viral load monitoring, does it make sense to use it in order to predict development of resistance mutations to HAART?The aim of the study was to assess, among HIV+ patients receiving their first-line HAART and with virological failure , the prevalence of genotypic resistance mutations and their pattern. Technology used was dried blood spots filter paper (DBS), due to its simplicity and feasibility in resource limited settings.Médécins sans Frontieres Spain (MSF-S) has provided ARV treatment since July 2003 in Busia District Hospital, in the western region of Kenya (Western province). Busia has a population of approximately 430,000 and HIV prevalence of 5.9% (age 15–49 years). After a period of expansion and scale-up of patient numbers, the project focused on strengthening the care and treatment of patients, having started around 3.500 patients on treatment at district level and rural level through decentralised provision of care by December 2008.The first-line regimen in Kenya when this study was performed consisted in stavudine (d4T), lamivudine (3TC), and nevirapine (NVP). In the event of toxicity, one can substitute zidovudine (ZDV) for d4T or efavirenz (EFV) for NVP, using TDF/3TC or abacavir (ABC)/didanosine (DDI) for patients failing first-line treatment R, a fixed-dose formulation containing a combination of 2 nucleoside reverse transcriptase inhibitors (stavudine and lamivudine) with the non-nucleoside reverse transcriptase inhibitor nevirapine. Triomune can be taken twice daily, irrespective of food.In April 2008, all the adult ART-naïve patients receiving a triple ARV therapy regimen classified as standard first-line  for 12 months or more, who had attended the clinic at least once within the previous 6 months, and had given informed consent to participate, were considered for the study. The most widely ATR drug used is TriomuneAs of mid-April 2008 there were 1037 adult, active, previously treatment-naïve patients who were receiving standard fist line ARV for more than 12 months at start of study period in Busia program. Non-naïve patients as well as those treated with any other ARV regimen were excluded from the study.This study was approved by the Ethical Review Board of Médecins Sans Frontières and the Kenya National Ethical Review Committee in KEMRI . Written inform consent was not obtained because the majority of local people participating in this study were illiterate. Therefore, and after a thorough explanation of the study purposes, and in presence of a third person , we requested to all participants to acknowledge their verbal informed consent. Ethical Review Board of Médecins Sans Frontières as well as Kenya National Ethical Review Committee in KEMRI  approved this consent procedure.On the day of enrolment, a venous blood sample of 10 mL was taken and divided into two parts: one had a CD4 count done at the Busia lab using the FASCOUNT machine using the manufacturer's guidelines, and the other one was sent to Kenya Research Institute Nairobi (KEMRI) in a plasma preparation tube where viral load testing was done using the NucliSENS EasyQ HIV equipment, version 1.2, using the manufacturer's guidelines, with limit of detection of 50 copies/ml.Patients having a viral load >5.000 copies/ml provided an additional sample for preparation in dried blood spot (DBS) Fifty microlitres of whole blood and plasma were used to make each spot respectively. These were dried overnight on special drying racks, packed and sent to the Research Laboratory from Hospital Universitari MutuaTerrassa, in Barcelona, for HIV resistance genotyping tests. Before study initiation, training was implemented among technicians  responsible of sampling procedure.Adherence was measured using a simple questionnaire translated to the local language. The questions were structured based on the WHO adherence criteria. Pill counts for the patients were done by adherence counsellors.For the purpose of our study we performed a viral load test for all patients that were enrolled in the study. Samples with viral loads ≥5.000 copies/ml were selected for genotype testing.3 after 12 months of therapy. Treatment failure based on clinical criteria was defined as the occurrence of either a new or a recurrent disease defining WHO 3 or stage 4 at 12 months from the start of medication Treatment failure based on CD4 test criteria was defined as either a CD4 below the baseline or less than 50% of peak at 12 months or CD4 <100 cells/mmCurrently, WHO defines virological failure as plasma HIV-1 RNA level >5.000 copies/ml after 6 months of treatment. However, at the time of the study (2008) the programme was following WHO 2006 recommendations where the cut-off for virological failure was >10.000 copies/ml,. Notwithstanding, in our study the aim was to assess the level of HIV drug resistance mutations and to describe and rank specific HIV-Drug Resistance mutations and mutation patterns among patients not achieving virological suppression. Therefore, all patients with VL >5.000 had their sample collected for genotype testing.TM tubes using universal phlebotomy practices, and briefly gently inverted to mix. Of this sample, 400 ul were spoted on the filter paper in the 5 pre-marked circles (50 ul each). The remaining sample was centrifuged at 3000 rpm for 10 minutes to obtain plasma. This plasma was collected into vacutainer tubes for viral load assay. Filter papers were leaved to dry on drying racks overnight in a horizontal position at a room temperature. They were then packed in glycine envelopes with desiccants to keep them dry and shipped to the genotyping laboratory in Barcelona following the laid down procedures for sample packaging and shipment. Samples were kept during the whole procedure and transportation at room temperature. In our laboratory, total HIV-1 RNA from DBS was extracted using Nuclisens method . A unique fragment of 1023 bp of pol gene containing the major part of PR and RT was amplified using in-house RT-PCR and nested PCR previously described in the literature Patients having a viral load ≥5.000 copies/ml provided an additional blood sample to prepare dried blood (DBS-FP) on Schleicher and Schuell 903 filter paper to send to Hospital Universitari MutuaTerrassa, AIDS research Unit, in Terrassa, for genotyping test. The BD PPT™ tubes (plasma preparation tubes) were used to separate plasma, following the manufacturer's instructions. Briefly, 5 ml of whole blood were collected into the PPTGenotyping was validated using the ViroScore Suite .Qualitative and quantitative variables were described using percentages and 95% confidence intervals (CI), means and standard deviation or median and interquartile ranges (IQR), depending upon distribution was normal or not. Comparisons between patients with or without virological failure were examined using Pearson's chi square test or Fisher's exact test for qualitative variables and t-test or U Mann-Whitney test for quantitative variables. Previously, Levene test was performed to determine homogeneity of variance.In addition, proportion comparison test for independent samples through Z statistic were performed for comparison of adherence results.P values less than 0.05 were considered statistically significant. Analyses were conducted using the statistical package StataSE vs9.0 .Among 1037 recruited patients under HAART during ≥12 months, 926 were included in the analysis . Out of Among the whole population studied, 274 patients  had detectable viral load (≥50 copies/ml) and 55  had a viral load ≥5.000 copies/ml, thus underwent genotypic study. Among 55 patients of genotypic study, mean age was 37.9 years (range: 18.7–72.5) and 34 (61.8%) were females. Only 10 out of 55 patients (18.2%) experienced clinical failure,  defined as the occurrence of either a new or a recurrent disease defining WHO 3 or stage 4 at 12 months from the start of medication. The median CD4 cell count and HIV RNA were 116 cell/mm3 [interquartile range (IQR) 54–189] and 32,000 copies/ml respectively (IQR: 11.000–68.000) . ART regOf the 55 samples, only 18 (33%) could be amplified and evaluated by genotyping. Based on Stanford University HIV Drug Resistance Database, we inferred subtypes from entering FASTA sequence information: the majority of samples were subtype A , subtype D  and G .Among 18 samples sequenced on PR and RT genes, 15 out of 18  showed RT resistance associated mutations (RAMs). The median number of resistance mutations (any class) was 5 . Three samples had no mutations identified .Fifteen out of 18 patients (83.3%) carried NNRTI resistance associated mutations, although there were no patients with only NNRTI mutations. The median number of NNRTI mutations was 1 (IQR: 1–2). The most frequent NNRTI mutations were K103N/S (27.8%), followed by Y181C (22.2%), V108I and G190A (16.7%), Y181V (11.1%), K101E and Y188L (5.6%) .Prevalence of NRTI resistance associated mutations are depicted in Among patients carrying RAMs, 12/15 (80.0%) harboured RAMs associated to thymidine analogues (TAMs)  . All of In our study all failing patients with HIV-1 RNA >5.000 copies/ml harboured TAMs. Among the three patients having samples with no mutations identified, viral load were high .According to Stanford HIV database, 10 out of 15 (66.7%) patients were resistant to both ARV families used, that are stavudine, zidovudine, lamivudine and nevirapine/efavirenz.Regarding samples sequenced on protease gene, we have found only common polymorphisms not associated with decreased PI susceptibility. We did not find primary RAMs to protease inhibitors .Adherence questionnaire did not discriminate among those patients harbouring some resistance mutations and those without any resistance at all. In contrast, 30,99% of patients having viral load >5000 copies/ml (n = 55) vs. 44.2% of patients with viral load <5000 copies/ml (n = 871) reported “never missed medications” .Despite earlier doubts, evidence is available that even in countries with very limited resources, ART programmes based on the public health approach have shown effectiveness equal to that seen in clinical cohorts in USA and Europe using similar regimens Also virological failure rate in our study is similar than those reported so far in developed countries. Likely, results from several operational research performed in different RLS yielded the same results than our study, with a longer follow-up period .The ability to collect blood samples on filter paper represents an advantage for HIV drug resistance surveillance and monitoring, particularly in areas that lack the appropriate infrastructure for plasma processing and transport. We used DBS method for genotyping purposes due to its simplicity and the fact that filter paper technology allows shipment with minimal biohazard risks. One of the crucial points for the use of dried fluid spots for drug resistance genotyping in the field is the stability of RNA over time. Dried blood spots can be kept for long periods when refrigerated or frozen in hermetic bags with desiccant but exposure to high temperatures for extended periods seems to be associated with degradation. Few studies have reported on long-term storage or testing under field temperature conditions, and contradictory percentage of genotyping efficiency of blood and plasma sample collection devices such DBS results have been reported, compared with gold standard plasma pol in DBS among Cameroonian HIV infected people that were stratified according different plasma viral load (from 5.000 to >50.000 copies/ml) In our study, we could amplify to evaluate by genotyping only one third of samples. Such a low percentage of positive genotypic results among failing patients might be explained by different factors, mainly related with the obstinate real life in Operational Research in RLS that make difficult the optimisation of sampling procedure: storage conditions, transportation and cold chain preservation as it was mentioned in methods, in our study samples were kept during the whole procedure and transportation at room temperature. Also, combination of insufficient concentration of viral RNA and decreased quality of RNA in DBS specimens can explain our low PCR amplification rate. Nonetheless, in our study, there were no statistically differences in viral load between amplificated and non amplificated samples. McNulty et al. demonstrated also the same rate of amplification of HIV-1 Although resistance mutation patterns were not available among the whole samples sequenced, genotypic results were concordant with ARV treatment received by failing patients, which is either nevirapine/d4T/3TC or nevirapine/AZT/3TC . ResistaThe absence of any relevant resistance mutation among three patients probably corresponds to patients who did not take medication at all, reinforcing the importance of treatment adherence in the context of lack of regular access to VL monitoring and genotype. Intensive therapeutic education, counselling and psychosocial support might explain the very low rate of lost-to follow-up patients observed, thus emphasizing that human resources remain a key factor in resource-limited settings.The mutations observed were those expected under the first-line regimen used, and the great majority of patients were resistant to nevirapine/efavirenz (100%), lamivudine (100%) and stavudine (83%). Overall, these observations are similar to other first-line cohort studies in RLS describing HIV-1 mutation patterns The accumulation of thymidine analogue and other resistance mutations can confer cross-resistance to many, and possibly, to all NRTIs that might be used in a second line-regimen. This phenomenon may be less likely when tenofovir is included in the first-line regimen. However, despite WHO 2010 recommendations, tenofovir is still commonly unaffordable in RLS. This data, together with our findings showing high TAMS mutation rate in our cohort, as well as the high rate of d4T-related polineuropathy–data not shown-, suggest that in RLS we desperately need other changes to make treatment more comparable to that in industrialised countries. We should consider moving away from d4T, rarely used in the North, to the easier tenofovir, although it would also cost more money (tenofovir currently costs four times more).UNAIDS 2010 global report have included important key elements to improve the efficiency and quality of HIV treatment and care: a) start ARV therapy earlier, b) use less toxic and more patient-friendly options (fixed dose combinations) and c) use of laboratory monitoring tools such as CD4 and viral load counts. Nevertheless, introduction of safer but currently more expensive first-line ARTs, as well as viral load monitoring, needs to be phased-in as currently they may not be feasible or affordable in many high-burden settings with low coverage, less developed health systems, limited laboratory capacity, finite budgets and competing health priorities.Newer guidelines Although recent modelling study support little added survival benefit to the addition of laboratory viral load monitoring compared with clinical monitoring alone In this regard, recently, some authors have presented data from an operational research trial in Zambia, aiming to ascertain the added value of routine viral load testing. This cluster randomized trial of routine versus discretionary  viral load monitoring among adults starting ARV therapy, demonstrated that routine viral load testing clearly limits the amount of time spent on a failing regimen in RLS. Time between virological failure and change of the regimen to second line treatment among routine viral load testing group was 168 days compared with 560 days among control group, thus with substantially less time (>1 year) on a failing regimen and higher rate of change to a second line regimen We have long known of increased mortality in African patients on ART compared with outcomes elsewhere. A conclusion would seem that if the future is to be different, we have to intervene earlier. Current worldwide expansion of ARV therapy for HIV in resource-limited countries entails new challenges to be considered. More than 6,5 million people are receiving ART, with the majority of patients on treatment worldwide now being from resource limited settings. Important treatment-related issues identified by several scientific communities, such as when to switch therapies, long-term consequences of virological failure after current first ART strategy, toxicity, adherence, drug-drug interactions or optimal treatment of pregnant women, need to be addressed immediately.Moreover, controversial issues about comprehensive laboratory monitoring such as routine monitoring viral load for safety and efficacy in these settings should be elucidated. VL testing could increase in importance as a guide for clinical decisions on when to switch to second-line treatment and on how to optimize the duration of the first-line treatment regimen, minimizing the impact of ARV resistance mutations on treatment. Therefore, now we must pay attention to monitoring to limit the costs associated with widespread use of expensive second-line therapy, ensuring that VL testing becomes affordable, simple and easy to use in RLS.


Two phase IV trials, ARTEN and NEWART, compared virologic safety/efficacy of NVP vs ATV/r on a background of TDF/FTC in HIV+ patients within these CD4 ranges. NEWART (US) was designed to be confirmatory of ARTEN and to supplement ARTEN data [Until recently, few prospective data existed to indicate safety/efficacy of NVP with entry CD4 count restricted to <250 (women) or <400 (men) cells/mmTEN data .In NEWART, treatment naïve patients were randomized to open-label NVP 200mg BID (lead-in dose 200mg QD x 14 days) or ATV/r (300/100mg) plus TDF/FTC (300/200mg). Women had CD4 counts (cells/mm3) <250; men <400. Primary endpoint was virologic response prior to and at week 48 defined as confirmed HIV viral load (VL) <50 copies/mL without subsequent rebound or therapy change. A point estimate of -6.5% or higher for difference in proportion of responders (NVP-ATV/r) was considered consistent with a successful ARTEN study.152 patients were treated: 89% male, 68% White, 31% Black. At baseline, mean log10 VL was 4.9 and median CD4 count (cells/mm3) was 176 (NVP) and 193 (ATV/r). Table NVP + TDF/FTC was noninferior to ATV/r + TDF/FTC. Although trial discontinuations were greater in the NVP arm, VR was similar because of less documented VF. HDLc increased and TC/HDLc decreased significantly more for NVP than for ATV/r.


The subjects for this study comprised twenty (20) non-HIV infected volunteers as control and one hundred (100) HIV infected subjects. Forty nine (49) infected patients have not been on treatment, while fifty one (51) were at various stages of treatment. There was a negative correlation between the serum malondialdehyde concentration and CD4 count . There was also a negative correlation between serum malondialdehyde concentration and extracellular superoxide dismutase activity  and Vitamin A concentration . Conversely a positive correlation was observed between the CD4 counts in HIV infected patients and activity of extracellular superoxide dismutase . Similarly there was a positive correlation between CD4 count and serum vitamin A concentration (Pearson r=0.89 p<0.01). The possibility remains for using these indicators to monitor HIV patients not eligible for therapy in resource constrained facilities of our rural areas.This study looked at the profile of plasma extracellular superoxide dismutase (SOD3) activity, malondialdehyde (MDA) vis-à-vis that of CD Human Immunodeficiency Virus (HIV) Infection induces a wide array of immunological alterations resulting in a slow but progressive development of opportunistic infections and malignancy, culminating in a disease condition known as acquired immunodeficient syndrome (AIDS) due the collapse of the immune system . The mec4 T-cells. Thus, the progressive loss of CD4 lymphocytes in HIV-infected patients may be related to an oxidative stress induced apoptosis , and 4-hydroxynonenal , 3, 6. USeveral lines of evidence suggest that HIV infected patients are under chronic oxidative stress. Oxidative stress occurs in HIV infection even at the early stages of disease . This prAntiretroviral treatment was embarked in Nigeria as a self sponsored program in 1990s, available to only those who could afford. Subsequently government intervention became evident in 2002, with the ambition of providing drug to 10,000 adult and 5,000 children within a year at the cost of $7 per person  of an avAll chemicals were purchased form Sigma Aldrich USA unless otherwise stated.This study was conducted according to the guidelines laid down in the Declaration of Helsinki  and all Forty-Nine HIV infected patients who have not been placed on Anti retroviral therapy and fifty one HIV infected patients who are receiving anti retroviral therapy were recruited for the study. Twenty HIV negative volunteers were included as control.3) of blood sample was collected into EDTA vacutainer tubes from the subject, attending Prof. S. S. Wali centre of Human Virology, Aminu Kano Teaching Hospital, Kano, Nigeria. The whole blood was used to determine CD4 count, while the remaining sample was centrifuge to obtain the plasma at 300 rpm for 5 minute. The plasma in sample bottle was wrapped with Aluminum foil and kept at −80°C before analysis.Five milliters .Determination of Serum Vitamin A Level: The method of Carr-Price (3 plasma was added followed by the addition of 0.5 cm3 ethanolic potassium hydroxide (containing 1 M potassium hydroxide and 90% absolute ethanol) to precipitate protein. Light petroleum 3.0 cm3 of petroleum ether was added and shaken vigorously for 10 min, then centrifuged at a low speed (1000 rpm) for about 1 min. 3.0 cm3 of petroleum ether layer was pipitted into a 10 cm3 test-tube and evaporated to dryness at 60°C in a water bath with the addition of 0.1 cm3 acetic anhydride to remove traces of water. 1 cm3 of chloroform was added to the residue, followed by the addition of 3 cm3 of antimony trichloride (250 g/L in chloroform). The colour developed rapidly reaching a clearly defined maximum in 5 to 15 seconds and faded quickly. The spectrophotometer was set to zero and reading taken immediately at 620 nm.rr-Price  was used3 of chloroform, 0.1 cm3 acetic anhydride, followed by the addition of 3 cm3 of antimony trichloride (250 g/L in chloroform).Blank was prepared by pipetting 1 cmVitamin A. Vitamin A as Retinoyl palmitate 1,600,000 usp units per gram purchased from sigma was used to construct a calibration curve. Both working and standard solutions were prepared in absolute ethanol. The standards were treated as test samples. Beckman coulter DU 720 UV/Vis Sepectrophotometer was used to measure absorbance at 620 nm.et al. .Extracellular Superoxide Dismutase (ESOD): The method of McCord and Fridrich  → O2 + NBT (purple)OIn the absence of SOD, the radical reacts with the detector. In the event that SOD is present, it competes with the detector for superoxide radical.−3 MNitroblue tetrazolium salt (NBT) 1 × 10−5 M and EDTA 0.11 mM (Prepared together for test and reference standard), 5.56 × 10−5 moles of oxygen was bubbled into the the combined solution of FAD and EDTA. Oxygen was prepared by the decomposition of 0.038g of Potassium trioxo chlorate (V) in the presence of 0.01g of Manganese (IV) dioxide as a catalyst).Flavine adenine dinucleotide (FAD) 5.56 × 10−5 M (without EDTA for blank)Flavine adenine dinucleotide (FAD) 5.56 × 10All stock solutions were prepared in 50mM potassium phosphate pH7.8.−5 M FAD (without EDTA) and 50 μl of 1 × 10−3 M NBT to give a final concentration of 5.0 × 10−5 M FAD and 5.0 × 10−5 M NBT.Blank: 50 μl of 50 mM potassium phosphate pH7.8, was added to 900 μl of 5.56 × 10−5 M FAD (with EDTA containing bubbled oxygen) and 50 μl of 1 × 10−3 M NBT to give a final concentration of 5.0 × 10−5 M FAD and 5.0 × 10−5 M NBT.50 μl of 50 mM potassium phosphate pH7.8, was added to 900 μl of 5.56 × 10The blank was used to set double beam spectrophotomer to zero and change in absorbance at 550 nm was recorded.−5 M FAD (with EDTA containing bubbled oxygen) and 50 μl of 1 × 10−3 M NBT were to give a final concentration of 5.0 × 10−5 M FAD and 5.0 × 10−5 M NBT. The blank was used to set double beam spectrophotometer to zero and change in absorbance at 550 nm was recorded. Amount of enzyme protein to inhibite the reaction between NBT and superoxide radical by 50% was determined. Activity per mg protein of ESOD was then calculated.Graduated volume of the sample was added to give 50 μl with 50 mM potassium phosphate pH7.8 Then 900 μl of 5.56 × 102− and NBT by 50%.DEFINITION OF A UNIT: A unit is defined as the quantity of SOD required to inhibit reaction between OEstimation of protein concentration was carried out by the method of Bradford . Bradforp<0.05 were used for all tests.Data are expressed as mean ± SD. In preliminary analyses, we used one-way analysis of variance (ANOVA) (SSPS 14.2 software) for continuous variables, to compare data from the three groups analyzed. Statistical significance levels of at least 4 status of the subjects studied. The F values for MDA, vitamin A and ESOD were 43.16, 40.88 and 101.60 respectively (p<0.000).Tables 4 between 200−400 cells/mm3 on treatment compared with CD4 below 200 cells/mm3 naïve patients  respectively. However there was no statistical difference between MDA plasma level between patients with CD4 count below 200 for treated and untreated groups  respectively. Figure 4 count and plasma MDA concentration. There was a negative correlation between CD4 count status and serum MDA concentration . Plasma MDA concentration increased with the decrease in CD4 count.The Plasma levels of MDA expressed as μmol/L are shown in Table 4 count . There was a positive correlation between serum vitamin A concentration and ESOD activity .The serum levels of vitamin A expressed as IU are shown in Table t Tables  and 2. T4 counts. There was a positive correlation between extracellular superoxide dismutase and CD4 count in this study . The change in extracellular SOD activity was not statistically significant for naïve and treated HIV subject with CD4 ≤200 . There was very slight significant increase in ESOD activity between HIV1 treatment-naïve and treated HIV 1 patients with CD4 count 200−400 . There was a significant increase in ESOD activity with CD4 count 200-400 over patients with CD4 count ≤200 for either HIV treatment naïve or treated HIV subjects (p<0.001).The levels of extracellular superoxide dismutase activity are shown in Table 01] Fig. . There wThe success of HIV infection management does not only rely on effective treatment, but also on efficient and cost effective monitoring tests. In Nigeria the cost of drugs per patients was about $368, while monitoring was about $170, representing 46% of drug cost in 2004. Though government subsidized ARV drugs, patients paid about $86 annually as a contribution to the drug cost  respectively  (Table ARVs were being administered in only 25 treatment centers across the country which was a far from adequate attempt at helping the estimated 550,000 people requiring antiretroviral therapy . As a reely Fig. . Increastus Fig. . Conversunt Fig. . Though unt Fig. , in this4 count in HIV infection. Thus the use of oxidative stress and antioxidants indicators can be used as surrogate markers of CD4 count in the laboratory for monitoring of HIV treatment-naïve patients in laboratory with facilities constrains.We have shown that oxidative stress increases with depression of CD


Switch to once daily unboosted (u) atazanavir (ATV) is an attractive option for HIV-infected patients with undetectable viral load, due to its convenience and favorable metabolic profile. However, due to a large interindividual variability, increased risk of inadequate ATV plasma concentrations (ATVc) and virological failure (VF) may be observed. Dividing the daily dose in a twice a day (BID) regimen could increase ATVc and improve virological success.In a prospective observational cohort of HIV-infected patients, all individuals with undetectable viral load who were switched to uATV during at least one month were retrospectively selected. ATVc were measured by a validated HPLC in all patients at least two weeks after the initiation of ATV. ATVc was considered inadequate when trough concentration was below 0,150 mg/L. Patients with once (400 mg QD) versus twice daily (200 mg BID) uATV were compared.From 2002 to 2009, 58 patients who received a total of 69 uATV based-regimens (27 QD and 42 BID regimens) were included. At the start of uATV, patients received a median duration of 9 years (IQR 4-11) of antiretroviral therapy. The mean exposure time of uATV was 16 months. Clinical characteristics and comedications (including tenofovir) were similar in the two groups. ATVc was inadequate in 17 (63%) patients in the QD group versus 4 (9%) patients in the BID group (p<0.001). During the follow up, VF happened significantly more frequently in the QD group than in the BID group . VF or low ATVc lead to treatment discontinuation in10 (37%) QD regimens versus 4 (9%) BID regimens . No other significant differences were detected in the two groups.Despite possible bias due to the observational study design, strong differences were detected in plasma drug exposure and virological outcome in our study. When a switch to uATV is proposed in HIV-controlled patients, BID would be preferred to QD.


During followup, 152 (20%) patients died, with the majority of deaths occurring within 12 months of ART initiation. Mortality was associated with advanced immunodeficiency or the presence of incurable disease at baseline. Among patients remaining on treatment, the median CD4 gain was 216 cell/mm3 and 86% of patients had viral load <400 copies/ml at the last clinical visit. The Georgia ART program has been successful in treating injection drug users infected with HIV.Since 2004, Georgia achieved universal access to free antiretroviral therapy (ART). A retrospective cohort study was conducted to evaluate the outcomes of Georgia's ART program. The study included adult patients enrolled in the ART program from 2004 through 2009. Of 752 patients, 76% were men, 60% were injection drug users (IDU), 59% had a history of an AIDS-defining illness, and 53% were coinfected with hepatitis C. The median baseline CD4 cell count was 141 cells/mm The advent of highly active antiretroviral therapy in the mid 1990s fundamentally altered the natural history of HIV infection in industrialized countries, resulting in dramatic reduction in AIDS-related morbidity and mortality . HoweverGeorgia, Armenia, and Azerbaijan make up a group of former Soviet republics known as the South Caucasus Republics. With the fall of the Soviet Union and the independence of all these republics in 1991, much of the social structure supporting health care became increasingly dysfunctional and the system of national healthcare that held a high standard for all Soviet citizens fell into disarray as economies crashed and conflicts within and between the countries disrupted services and infrastructure. Each one of these countries has its unique set of problems and issues, but, in general, their declining economic situations, coupled with rising drug use and commercial sex work, and their geographic proximity to Russia and Ukraine, countries with emerging epidemics, make the South Caucasus a region ripe for the spread of HIV. Georgia is located at the juncture of Eastern Europe and Western Asia and is bordered by Russia, Azerbaijan, Armenia, and Turkey. In 2009, the population of Georgia was estimated to be 4,615,807 in a territory of 26,900 sq miles. The GDP per capita in 2009 was approximately $4,300 US ranking 103 in the world, and the GINI coefficient was 40.8 ranking 59th in the world. Georgia regained independence in August of 1991, followed by a two-year civil war that led to large population migration with internal displacement, severe poverty, a dysfunctional economy, and disruption of many national services including healthcare. The country was stabilized in 1995, but the dire economic position of the Georgian government necessitated conversion of the state-funded health care system to a market-driven system. Nearly half of all Georgians are now forced to forego professional medical care when sick, opting instead for advice from friends and relatives, traditional medicine, or no care .The first case of HIV in Georgia was reported in 1989. As of December 31, 2009 a cumulative 2,236 HIV cases had been reported. Among them 1,151 persons developed AIDS and 479 died. Males constitute the majority (74%), of the reported HIV/AIDS cases with injection drug use (IDU) responsible for HIV acquisition among nearly 60% of all reported cases. The estimated HIV prevalence in Georgia is less than 0.1%. Antiretroviral drugs have been available in the country since 1990s, but the access was limited only to those able to afford them. Since 2004, through support from the Global Fund, Georgia became the first country among the newly independent states (NIS) of Eastern Europe to achieve universal access to antiretroviral therapy (ART). This major achievement has been acknowledged in the joint WHI/UNAIDS/UNICEF report on universal access [A retrospective cohort study was conducted at the Infectious Diseases, AIDS and Clinical Immunology Research Center (IDACIRC), in Tbilisi, Georgia, which is the country's referral institution for HIV/AIDS diagnosis, treatment, and care. The study population included all adult (age ≥18 year) patients enrolled in the ART program from 2004 through September 30 2009, who started therapy for at least six month prior to April 1, 2010.The National ART Program is coordinated by the IDACIRC, and antiretroviral drugs are dispensed in the IDACIRC clinic in the capital city of Tbilisi as well as at three affiliated regional centers in the cities of Kutaisi, Batumi, and Zugdidi. HIV infected persons are identified through state and donor funded HIV testing and counseling (HTC) services and screening programs. All persons with positive screening test results are referred to IDACIRC for confirmatory testing and if confirmed are initially assessed at IDACIRC. Patients have the option to continue clinical care either at central or regional levels.3 or if the patient had an AIDS defining illness. ART was also recommended at CD4 cell count of ≤350/mm3, based on the CD4 cell decline rate, a high HIV-1 viral load, and coinfection with viral hepatitis. Currently, steps are being taken towards implementation of the recommendation to initiate treatment in all patients with CD4 cell count of ≤350/mm3. Provision of therapy is governed by the National HIV/AIDS Treatment and Care guidelines developed based on the protocols of WHO as well as the guidelines of major Western countries –16. The Identification of patients in need of treatment is based on following those who don't qualify for therapy every 3-4 months and monitoring the CD4 cell count and HIV viral load. This allows for the timely identification of those in need of treatment.The recommended initial regimen consists of two nucleoside reverse transcriptase inhibitors (NRTI) and one nonnucleoside reverse transcriptase inhibitor (NNRTI). A ritonavir (r) boosted protease inhibitor (PI) is recommended in cases when an NNRTI cannot be prescribed. Currently tenofovir (TDF) + emtricitabine (FTC), zidvudine (AZT), or abacavir (ABC) + lamivudine (3TC) are used for the NRTI component of initial regimen. Stavidine (d4T) is no longer recommended after the 2007 revision of National Guidelines; however the drug is reserved for short-term use for situations when AZT, ABC, or TDF cannot be used because of severe toxicity. Since 2008, patients are tested for HLA B∗5701 before starting on an ABC-containing regimen. Efavirenz (EFV) is the preferred NNRTI with nevirapine (NVP) being recommended as an alternative to EFV. Selection of subsequent regimens in treatment-experienced patients is based on the drug resistance profile with the goal of providing patients with at least two, and preferably three, fully active drugs. In addition to boosted PIs , new classes of drugs are also now available for highly treatment-experienced patients, such as the integrase strand transfer inhibitor raltegravir (RAL), the fusion inhibitor enfuvirtide (ENF), and the CCR5 antagonist maraviroc (MVC). Recently, the new NNRTI etravirine (ETV) has also become available.As per the Georgian National guidelines, the standard of ART monitoring relies upon laboratory monitoring of CD4 count, HIV-1 viral load, and development of resistance based on a resistance-genotype detection when indicated. Virological failure is defined as confirmed plasma HIV-1RNA >400 copies/ml 6 months after starting therapy or plasma HIV-1 RNA >50 copies/ml 12 months after starting therapy in a patient who is on potent ART. Georgia was the first NIS country to introduce genotypic resistance testing in 2005 into routine clinical practice. HIV drug resistance testing is used to guide treatment decisions during virologic failure and to make decisions as to the most effective subsequent regimen.Special attention is paid to adherence to therapy as an important determinant of treatment success. A program to promote and maintain antiretroviral adherence has been developed that includes maintenance of an adherence diary, pill identification by shape and color, patient self-report about the medication intake in the preceding 3- to 7-day period and medication refill using pharmacy records. In addition, to improve adherence, mobile units to deliver home-based adherence support operate countrywide.Plasma HIV-1 RNA levels were initially measured using Amplicor HIV-1 Monitor test, version 1.5 , with lower limit of detection of 400 copies/ml. Since 2006, the real-time PCR assay COBAS TaqMan HIV-1 test  has been in use with a lower limit of detection of 40 copies/ml.Determination of CD4+ cell count is based on the single-platform immunophenotyping technique using the FACSCalibur flow cytometer  with four-color direct immunofluorescence reagent MultiTEST CD3/CD8/CD45/CD4.For genotypic resistance testing, the TruGene HIV-1 Genotyping Kit was employed according to the manufacturer's instructions using OpenGene DNA Sequencing System . The Guidelines Rules version 14.0 and Stanford University algorithm (http://hivdb.stanford.edu/) were used for resistance interpretation. Mutations listed by the International AIDS Society-USA Panel were considered .Data were obtained from the National HIV/AIDS electronic database, operated by IDACIRC. The database contains information on all reported HIV cases, including demographic, epidemiological, clinical, and laboratory data. Information on all patients initiating HAART from 2004 through 2009 was extracted. Observations were censored as of April 1, 2010. Descriptive statistics were performed to assess distribution of covariates. Normality of continuous variables was evaluated using Q-Q plots. Kaplan-Meier product-limit estimator method was used to assess probability of survival and probability of virological failure. Predictors of mortality were evaluated in multivariate Cox proportional hazards model. The proportional hazards assumption was tested and was met for the final model. All tests were two-sided at a significance level of 0.05. Statistical analyses were performed using SAS v 9.2 .Study was approved by Institutional Review Board (IRB) of the Infectious Diseases, AIDS and Clinical Immunology Research Center. The study was based on information routinely collected as part of the standard of clinical care of HIV infected individuals.Since 2004, over 1,800 patients have been seen for HIV clinical care at the IDACIRC and of these, 841 adults met the treatment initiation criteria and were enrolled in the HAART program (through December 2009). This analysis includes 752 adult patients who had started therapy at least six months prior to inclusion in the study .3 (IQR 76–208), with approximately one third of patients having a CD4 cell count less than 100 cells/mm3. Median viral load was 5.4 log10 copies per ml (IQR 4.8–5.8). Fifty-nine percent of patients had a history of an AIDS defining illness (ADI), with 32.4% having a history of active tuberculosis. More than half of the patients had antibodies against Hepatitis C virus (HCV), 8% of patients had evidence of chronic Hepatitis B infection, and 6% had dual infection with HCV and HBV. Fourteen percent of patients had cirrhosis. Almost 3% of patients had a malignancy.All but 5 patients were started on an NNRTI-based regimen, most frequently with EFV (82.9%). AZT + 3TC was the most common NRTI component of the first ART regimen (53.0%), followed by ABC + 3TC (27.2%) and d4T + 3TC (18.3%). After the 2007 revision of the national HIV/AIDS Treatment and Care guidelines, all patients on d4T were switched to AZT, ABC, or TDF.The median duration of followup was 24 months (IQR 10–45 months). During followup, 152 (20.2%) patients died and 34 (4.5%) patients self-discontinued ART/were lost to follow-up, with overall retention rate of 84%. Kaplan-Meier estimates of survival probability were 0.84 (95% CI: 0.82–0.87), 0.80 (95% CI: 0.77–0.83), 0.78 (95% CI: 0.75–0.81), and 0.77 (95% CI: 0.74–0.80) at 12, 24, 36, and 48 months, respectively . Of 152 3 , history of an AIDS-defining illness , cirrhosis . Other covariates did not show statistical significance in multivariate analysis , CD4 cell count <100 cells/mmanalysis . Presenc3 (IQR 112–348) and 487 (86%) patients had an HIV-1 viral load measurement of <400 copies/ml at their last clinical visit. Data on medication refill adherence were available starting from 2007; the refill adherence rates were 85% in 2007, 83% in 2008, and 92% in 2009.Among 566 patients still on ART, the median gain of CD4 cells was 216 cells/mmEighty-two patients (10.9%) experienced virological failure. All of them have been tested for the presence of drug resistance virus while on a failing regimen. Median time to virologic failure was 16 months (IQR: 10–28 months). Kaplan-Meier probability of failure at 12, 24, 36, and 48 months were 0.04, 0.11, 0.16, and 0.19, respectively . Of 82 pThe frequency of drug resistance mutations in the reverse-transcriptase (RT) gene is shown in All patients with drug resistant viruses were switched to second line regimens with boosted PIs. At present, of 56 patients remaining on second-line therapy, 61%, 21%, 11%, and 7% receive LPV/r, ATV/r, DRV/r, and FPV/r based regimens respectively. Three patients are on salvage regimens, consisting of ENF plus optimized background therapy.We report the outcomes of the universal ART access program in Georgia. The program builds upon the successful training of young clinicians and scientists who started in their training in the care of HIV/AIDS patients in the 1990s with the goal of providing high quality clinical care in an environment that promotes research. The capacity to provide universal access to ART has been further strengthened through the support of the Global Fund. Under this framework, the approach to ART provision taken by Georgia has been designed to maximize effectiveness of the intervention.The IDACIRC mandate—as the lead agency for the provision of all HIV care activities in the country—ensures universal access and high retention on therapy, with less than 5% drop-out rate. In addition, a coordinated approach to ART monitoring, including availability of laboratory monitoring , and adherence counseling contributes to the achievement of optimal outcomes. Establishment of mobile units for providing home-based adherence support and monitoring resulted in improvement of medication refill adherence from 83% in 2008 to 92% in 2009. Good immunological and virologic responses seen in our cohort also serve as an evidence of good overall adherence. The probability of virological failure in our study was similar to that reported from developed countries , 19. In Interestingly, the most common NNRTI mutation detected in our study was G190S/A but not K103N. It is possible that G190S/A is the favoured NNRTI mutation in HIV subtype A1—which is the most common circulating strain in Georgia . FurtherAn important challenge that remains to be addressed and that has been identified through this study is the early mortality of patients who start ART in Georgia. As shown in our analysis, the highest mortality rate was observed within the first year of ART initiation, with 75% of all deaths occurring in the first 12 months. Our results are consistent with findings from other resource-constrained countries reporting increased risk of mortality early after starting ART –26. In oThe most common causes of death in our cohort were TB and end stage liver disease, together accounting for more than 40% of all deaths. Multivariate analysis showed a strong association between cirrhosis and death, emphasizing the problem of coinfection with viral hepatitis, especially with hepatitis C. Dually infected patients virtually have no access to anti-HCV therapy because of prohibitively high costs. Due to this fact, HIV/HCV coinfected patients die from hepatic disease despite the successful antiretroviral therapy. The high prevalence of coinfection with HCV among patients with HIV in Georgia may prevent the full realization of the benefits of ART, and may compromise the cost-effectiveness of ART. Recent studies have shown that the availability of ART does not decrease the risk of mortality among patients with a dual infection with HIV and HCV . MoreoveAnother important challenge is HIV/TB coinfection. The high prevalence of this coinfection in our cohort was not a surprise given the overall high burden of TB in the country . HoweverAs with all studies, limitations should be mentioned. First, the study was based on data available in national HIV/AIDS database. Adverse events of ART are carefully monitored, the data are not entered into electronic database, and therefore we were not able to determine contribution of adverse events to outcomes of interest. A further limitation is that the only baseline information on drug abuse was available and this precluded us from assessing the impact of current drug use on outcomes.In summary, the HIV epidemic in Georgia has entered a new phase. ART has been successfully introduced in Georgia, including in IDU population. The next stage is to maximize and sustain the benefits of ART. Mortality can be substantially reduced by improving earlier HIV diagnosis and initiation of ART. The comprehensive program for the identification and treatment of patients with HIV/AIDS in Georgia, a relatively low income country, has been quite successful in limiting premature mortality and morbidity from this epidemic disease. Increased efforts and new strategies are needed to effectively address the intersecting HIV/TB and HIV/HCV epidemics and their resulting mortality.


Second line antiretroviral therapy (ART) was rolled out in India in December 2008. However, there has been no study to assess the response to second line ART in India. This study aims to see the response to the NACO recommended second line therapy over 24 week period.Study was conducted in ART centre, IMS, BHU. Patients who failed on first line ART (NACO criteria) were included in the study after consent. Virological, immunological and clinical response was assessed at 24 weeks. Independent t-test /Mann-Whitney U test was used for making comparison among the independent groups. Chi-square test was used to test the association between categorical variables. Spearman correlation coefficient was used to measure the degree of association between two variables.Out of 78 patients, only 39 (50%) had virological suppression  and 20 (25.6%) had partial response , 10 (12.8%) had viral load >1000 copies/ml and 8 (10.3%) patients died before 6 months and 1 patient was lost to follow up. Median increase of CD4 count was 133 cells /ml (IQR -46-498). Among patients who expired 90% had clinical failure at baseline, 50% and 18.75% were in stage IV and III respectively.Currently recommended second line regimen appears to be inadequate as large proportion of patients failed to achieve the desired virological suppression. Our finding implies a relook at the present second line regimen recommended by NACO.