All of these markers, with the exception of sTNF-R1, were also elevated in patients failing HAART (P < 0.05). The persistently elevated levels of CXCL9, CXCL10, and β2M in patients failing therapy in the setting of a marked reduction in these markers in patients on successful HAART suggest that they may be useful not only to monitor immune activation during HAART, but also to distinguish between good and poor responders. In the case of sCD14 and TGF-β1, the levels of these biomarkers remained persistently elevated despite HAART-induced virological suppression, a finding that is consistent with ongoing monocyte-macrophage activation, underscoring a potential role for adjuvant anti-inflammatory therapy.Few studies have examined immune activation profiles in patients with advanced HIV-1 subtype C infection or assessed their potential to predict responsiveness to HAART. BioPlex, ELISA, and nephelometric procedures were used to measure plasma levels of inflammatory biomarkers in HIV-1 subtype C-infected patients sampled before and after 6 months of successful HAART (n = 20); in patients failing HAART (n = 30); and in uninfected controls (n = 8). Prior to HAART, CXCL9, CXCL10,  In addi (TGF)-β –13. In t (TGF)-β –23. The ll death , 21, 22.ll death . While htivation .Immune activation in HIV infection is associated with the presence of circulating proinflammatory/anti-inflammatory and antiviral cytokines/chemokines, as well as with other biomarkers of immune activation, which vary qualitatively and quantitatively with disease progression –31. Howe+ T-lymphocyte counts (CD4+) (Beckman Coulter SA (Pty) Ltd.) and HIV-1 RNA (VL)  were measured by standard flow cytometric and PCR-based procedures respectively, according to manufacturer's instructions.Black, adult (≥18 years) participants attending the Antiretroviral Clinic at a district hospital in Pretoria, South Africa, were included in this study. Ethics approval was granted by The Research Ethics Committee, Faculty of Health Sciences, University of Pretoria . All participants gave informed consent and whole blood samples were collected in EDTA vacutainers, processed within 24 hours to separate the plasma component by centrifugation, and stored at −70°C for up to 37 months. CD4+ counts ≤200 cells/μL blood or WHO stage 4 disease. Twenty patients were randomly selected from those who started HAART, were clinically stable, did not develop clinical signs of IRIS during the first six months of treatment, and were virologically suppressed (VL < 50 copies/mL plasma) at approximately 6 months of HAART (suppressed group). Drug regimens consisted of two nucleos(t)ide reverse transcriptase inhibitors (NRTIs) (stavudine (d4T) + lamivudine (3TC), n = 18, or tenofovir (TDF) + 3TC, n = 2) and one nonnucleoside reverse transcriptase inhibitor (NNRTI) (efavirenz (EFV), n = 14 or nevirapine (NVP),  n = 4). Two patients were started on ritonavir-boosted lopinavir (LPV/r) for clinical reasons.Sixty HIV-infected participants were followed from pre-treatment to approximately 6 months on HAART as part of a larger study on immune reconstitution inflammatory syndrome (IRIS). Pre-treatment samples were taken prior to the initiation of HAART in patients presenting with CD4n = 23 or zidovudine (AZT) + 3TC, n = 7) and one NNRTI . Participants had been referred for drug resistance testing and study samples were taken at the time of referral. They had been on HAART for a median time of 30 months (range 9–97 months) and had been failing treatment for a median of 15.5 months (range 5–38 months). Five patients (17%) had been referred from peripheral clinics and the duration of treatment failure could not be determined. Three patients (10%) had experienced treatment interruptions at some time before treatment failure and 13 (43%) never had a suppressed VL while on HAART. All patients with CD4+ ≤200 cells/μL (n = 21) were on cotrimoxazole or dapsone prophylaxis.A second group consisted of 30 participants failing HAART as evidenced by two successive VL results of >1000 copies/mL plasma at least eight weeks apart despite intensive adherence counselling (failing group). Drug regimens consisted of two NRTIs  of black, HIV-uninfected, healthy control subjects was also included in the study. The median ages of the control, suppressed, and failing groups were 29 (range 24–49), 41.5 (25–63), and 40.5 (27–55) years, respectively, and the corresponding male : female ratios were 1 : 0.6, 1 : 4, and 1 : 4.A third group  or (ii) conventional ELISA, namely, IFN-α ; TGF-β1 total ; CXCL9/MIG and sTNF-R1 ; and sCD14 . C-reactive protein (CRP) and β2-microglobulin (β2M) were assayed by nephelometry . Previously published ranges for each of these parameters together with supporting references are shown as supplementary data .Circulating cytokines/chemokines were measured using (i) the Bioplex suspension bead array system  , as well as for this group combined with the group failing HAART (n = 50). Statistical significance was set at P ≤ 0.05.As participant groups consisted of ≤30 individuals, data were considered to be nonparametric and distribution-free statistical tests implemented in Stata v11.2 (StataCorp). Continuous data were analysed according to the median, minimum, and maximum concentrations. Median concentrations of each parameter were compared between cohorts using the Wilcoxon Mann-Whitney test for independent groups and Wilcoxon signed rank sum test for matched groups. Correlations between parameters were determined using the Spearman correlation test for the HIV-infected pre-HAART group  in the suppressed group. With respect to the circulating biomarkers of immune activation, CXCL9, CXCL10, TGF-β1, sTNF-R1, β2 M, and sCD14 were significantly elevated (P < 0.03) and CCL4 significantly decreased (P = 0.04) in the pre-HAART group relative to the control group, while IFN-γ was moderately increased but not significantly so (P = 0.07). Following 6 months of HAART, CXCL9, CXCL10, β2 M, IFN-γ, IL-6, TNF-α, and sTNF-R1 were significantly decreased (P < 0.01), CCL4 increased (P < 0.001), while TGF-β1 and sCD14 also remained elevated despite undetectable plasma VL. It is difficult to attribute major significance to the decreases in TNF-α and IL-6 as the pretherapy values for both were low. No difference was observed in IFN-α, CCL3, and CRP either between the HIV-infected and uninfected control group or the virologically suppressed group pre- and post-HAART.CD4β1, β2M, and sCD14) were also significantly elevated compared with the control group (P < 0.02), the values for CXCL10 and β2M being somewhat lower than those of the pre-HAART group (P < 0.03), while those of CXCL9, TGF-β1, and sCD14 were essentially comparable (P > 0.5). Although the value for CCL2 was significantly lower and that of IL-10 higher than the corresponding values of the control group, interpretation is difficult as these values were low in both groups.In the failing group, the same 5 biomarkers  were generally comparable, albeit weaker, with the exception of CD4+ count with VL , while the following modest correlations were found: (i) CCL4 with CXCL9, IL6, CCL3, and IFN-γ ; and (ii) β2M with CD4 counts, IL-6, and IFN-γ .Correlations between CD4γ, probably originating from CD4 and CD8 T cells, underpins the increases in CXCL9 and 10, a contention supported by the strong, positive intercorrelation between IFN-γ and CCL10, as well as that of CCL9 with CCL10. Other cell types such as dendritic cells and monocytes may also contribute to the increases in these cytokines pre-HAART following exposure of the cells to alternative activators such as IFN-λ1 [α, as well as those of CCL2 and 3, may be due to advanced immunosuppression in the setting of high levels of TGF-β1 in this group of patients [β2 M, CXCL 9 and 10, and TNF-R1 (a surrogate for TNF), HAART-associated decreases most likely reflect efficient viral suppression and consequent decreased turnover and reactivity of both CD4+ and CD8+ T cells.Our findings in patients infected with HIV-1 subtype C are consistent with the coexistence of distinct mechanisms of immune activation, which appear to be differentially affected by successful HAART , 25. Alts IFN-λ1 , 32, 33.patients . In the The absence of effects of HAART on plasma sCD14, as previously reported by us and others , 21, as β1 appearing to play a pivotal role. Notwithstanding platelets, plasmacytoid dendritic cells, macrophages of the M2 phenotype, and immunoregulatory CD8+ T cells, immunosuppressive and profibrotic TGF-β1 is likely to originate predominantly from regulatory T cells [In addition, supported by the findings of the current study, endogenous, monocyte/macrophage-targeted, anti-inflammatory mechanisms are also likely to contribute to ongoing immunosuppression with TGF- T cells , 39. In  T cells .β1 and sCD14, even in the setting of ostensibly successful HAART, may therefore identify a subset of patients at highest risk of a poor outcome.Persistently elevated plasma levels of TGF-β2M were also significantly higher than those of the control group and, with the exception of CXCL9, significantly lower than the pre-HAART values for the suppressed group. The circulating concentrations of sCD14 and TGF-β1 in the failing group were comparable to those of the suppressed group both before and after therapy. Persistent elevations, or a rebound following an earlier decrease, in plasma CXCL9, CXCL10, and β2M appear to be associated with a poor response to HAART, suggesting that serial measurement of these biomarkers may be a useful adjunctive strategy. Nevertheless, measurement of VL clearly remains the definitive strategy in the clinical setting.In the group of patients failing HAART, the circulating concentrations of CXCL9, CXCL10, and + T cells and VL but failed to show a correlation between these disease markers and CXCL10 in the pre-HAART group. However, this correlation was detected when the pre-HAART and failing groups were combined, most likely due to increased statistical power. As mentioned above, and in agreement with Kamat et al. [γ was evident as can be expected in conditions of chronic inflammation.With respect to previous studies, our findings are generally in agreement with a recent study by Kamat et al. in which elevated circulating concentrations of CXCL9, CXCL10, sCD14, and soluble IL-2 receptor (sIL-2R) represented a profile which distinguished viremic and aviremic subjects infected with HIV-1 subtype B from uninfected, healthy control subjects . In agret et al.  we also β2M or TGF-β1, was not measured serially in a single cohort of patients pre- and post-HAART as done in the current study, which may account for the observed lack of effect of HAART on IFN-γ in the former study.Notwithstanding the different viral types investigated, several other important differences underscore the strengths of the current study. Most importantly, the profile of biomarkers of immune activation measured by Kamat et al. , which dLimitations, however, are (i) small sample sizes; (ii) measurement of circulating biomarkers at a single time point (6 months) following initiation of HAART in the suppressed group; and (iii) no pretherapy measurement of circulating biomarkers prior to initiation of therapy in the failing group. Nonetheless, the general agreement with previous studies, predominantly in the setting of HIV-1 subtype B infection, supports the reliability of our findings.γ, CXCL9, CXCL10, sTNF-R1, and β2M). Serial measurement of 3 of these  may represent a useful adjunct to measurement of viral loads in monitoring responses to HAART. In addition, persistently elevated levels of sCD14 and TGF-β1, despite successful HAART, are consistent with chronic activation of monocytes/macrophages and possible risk of a poor outcome, underscoring the adjunctive therapeutic potential of monocyte/macrophage-targeted anti-inflammatory chemotherapy in patients with advanced HIV infection.In conclusion, successful administration of HAART to patients with HIV-1 subtype C infection is accompanied by significant decreases in circulating biomarkers associated with T-cell activation and turnover (IFN-This is a list of previously published studies documenting normal ranges for the various circulating biomarkers of immune activation assayed in the current study.Click here for additional data file.


This study aims to describe the virological, immunological and clinical efficacy of protease inhibitor (PI)-based second-line antiretroviral therapy (ART) in rural South Africa.An observational cohort study was performed on 210 patients (including 39 children) who initiated PI-based second-line therapy at least 12 months prior to data collection. Biannual clinical, immunological and virological monitoring was performed. Primary endpoints were adequate virological response (plasma HIV-1 RNA<400 copies/ml), full virological suppression (plasma HIV-1 RNA<50 copies/ml) and treatment failure  or on-going viremia (plasma HIV-1 RNA never<400 copies/ml for more than six months)). Data were analyzed by an on-treatment (OT) and intention-to-treat (ITT) approach. Analyses were primarily performed on the group of patients who switched following first-line virological failure.Median duration of follow-up after switch to second-line treatment was 20 months [IQR 11–35]. 191 patients had switched to second-line ART due to first-line virological failure. 139/191 of them  were in care and on treatment at the end of follow-up and 11/191  had died. After twelve months, an adequate virological response was seen in 92/128 patients , of which 78/128  experienced full virological suppression. Virological response remained stable after 24 months. Virological efficacy was similar amongst adult and pediatric patients. As in first-line ART, we observed a lack of correlation between virological failure and WHO-defined immunological failure.Good virological outcomes following first-line failure can be achieved with PI-based, second-line antiretroviral therapy in both adult and pediatric patients in rural South Africa. Retention rates were high and virological outcomes were sustainable during the two-year follow-up period, although persisting low-level viremia occurred in a subset of patients. The observed viro-immunological dissociation emphasizes the need for virological monitoring. Most of the people who are HIV-infected globally, reside in sub-Saharan Africa. The massive roll-out of antiretroviral therapy (ART) that has taken place in this region since 2003, resulted in a stark increase in the number of HIV-infected patients receiving treatment. Despite these impressive achievements, treating HIV-infected people in resource-limited settings (RLS) remains challenging.The World Health Organization (WHO) recommends combining two nucleoside reverse transcriptase inhibitors (NRTIs) and one non-nucleoside reverse transcriptase inhibitor (NNRTI) as first-line ART in RLS. Second-line antiretroviral therapy in RLS generally consists of a ritonavir-boosted protease inhibitor (PI), mostly lopinavir, combined with a dual NRTI backbone. The efficacy of second-line ART programs in RLS may be different from that previously described for western countries for several reasons. First, HIV subtypes circulating in African countries vary, with subtype C being most frequent, whereas subtype B is most prevalent in western countries. Data on the efficacy of second-line ART in RLS are rare; the few existing studies are limited by cohort size and follow-up duration and are mainly based in urban areas. Early reports generally show good short term outcomes with high rates of survival, immune reconstitution and virological suppression on second-line therapy. www.ndlovucaregroup.co.za) that provides “paid for service” primary health care, as well as prevention-, tuberculosis- and HIV/AIDS-programs which are donor-funded and free of charge for the patient. It serves a population of 120,000–140,000 people and around 4,000 HIV-infected patients are in care at the clinic. Patients are seen by doctors and HIV-counselors during each clinic visit in order to provide information and to stimulate treatment adherence. In a previous study, genotyping in this cohort showed that all viruses were of HIV-1 subtype-C origin. Elandsdoorn is a township situated in a poor, rural area in Limpopo, a province in the northeast of the Republic of South Africa. HIV prevalence among antenatal clinic attendees in 2010 was 21.9% in this province. Medical ethics review was not required for this observational cohort study that was a follow-up of previous studies conducted on this cohort. For this study, all patients who initiated (standard public-sector) PI-based second-line therapy at the Ndlovu Medical Centre at least 12 months prior to data extraction (between March 2004 and October 2010) were included. All patients had been previously treated with a WHO-recommended, NNRTI-based first-line ART regimen. Tenofovir was not prescribed for children, as current guidelines do not encourage its use in prepuberty. For second-line regimens, the NNRTI was replaced by ritonavir-boosted lopinavir in all patients, and combined with various NRTI-backbones. Pediatric dosages of lopinavir/ritonavir were supplied as oral solution, while for adults, the capsule formulation (which needs to be refrigerated) was replaced by tablets in 2008.Virological failure on first-line treatment was the main reason to switch to second-line therapy. In certain cases, there were other reasons to switch, such as adverse drug events while using first-line treatment. Virological failure on first-line ART was defined as a plasma HIV-1 viral load rising to over 1000 copies/ml after initial virological suppression , or continued detectable plasma HIV-1 viral loads over 400 copies/ml, after at least six months of ART.Plasma HIV-1 viral loads  and CD4 counts  during treatment were measured biannually in the laboratory on site. Shortly before and three months after switch to second-line therapy, additional plasma HIV-1 viral load measurements were performed.Data were retrospectively collected from the medical charts, using routinely collected follow-up data. Baseline information regarding previous first-line therapy, social status, age and gender were collected from the charts, as was the duration of first-line therapy. Initial first-line regimens were described. A drug switch within the NRTI class because of adverse drug events was not considered to be a switch to second-line treatment. A change to at least one new antiretroviral drug class was considered to define a switch to second-line treatment.The primary endpoints were achieving virological success , both at fixed time points after start of second-line treatment and at end of follow-up, and experiencing treatment failure during second-line therapy. Full virological suppression was defined as a plasma HIV-1 RNA<50 copies/ml and the definition of an adequate virological response was a plasma HIV-1 viral load<400 copies/ml. A plasma HIV-1 viral load>1000 copies/ml confirmed by a subsequent measurement after an initial adequate viral response was used to define virological failure. However, if no subsequent measurements were available, and the last plasma HIV-1 RNA was above 1000 copies/ml, patients were also considered to experience virological failure. The continued (more than six months) detection of a plasma HIV-1 RNA>400 copies/ml was considered to represent on-going viremia. Both virological failure and on-going viremia were regarded as second-line treatment failure. Virological responses were determined both for the entire group, and separately for adults and children (age<15 years).3 or below the pre-therapy count, or a 50% decline from the on-treatment peak CD4 count value. Secondary endpoints were both clinical parameters  and immunological parameters (absolute change in CD4 counts). A patient was considered lost to follow-up if he or she had not shown up at the clinic for six months or longer before the moment of data extraction and if there was no available information on death or transfer out to another clinic. Immunological responses were only analyzed for the adult patients and children of 5 years and older, as CD4 percentages, the preferred immunological parameter for HIV-infected children younger than five, were not available. Immunological failure for adult patients was defined according to the WHO guidelines as either a CD4 count after six months of therapy below 100 cells/mmAll analyses on second-line ART outcome were primarily performed on the group of patients who had switched following first-line virological failure and were repeated for the total group of patients.P<0.50) in univariate analysis. A P-value ≤0.05 was considered statistically significant. Data were processed and statistical analyses were done using SPSS version 17.0.Data were analyzed by an intention-to-treat (ITT) and on an on-treatment (OT) approach at fixed time points, for which a window period of three months prior to and following that time point was accepted for data collection. The end of follow-up for virological or immunological response was defined as the most recent date with available results on either plasma HIV-1 RNA or CD4 cell count, respectively, before the time of data extraction, death, transfer out or being lost to follow-up. Continuous data were compared with the Student's t-test or the paired t-test as appropriate. Proportions were compared with the chi-square-test and data that were not normally distributed were analyzed via the Mann-Whitney U or Wilcoxon rank test. Kaplan-Meier survival analyses were used to estimate the time from switch to second-line ART to full virological suppression. To define predictors of treatment failure, univariate analyses were performed with the following determinants: age, gender, being a child, first-line ART regimen (zidovudine versus stavudine as NRTIs and nevirapine versus efavirenz as NNRTIs), change of CD4 count during first-line treatment, CD4 count at the moment of switch to second-line ART, calendar year of switch, duration of first-line treatment, duration of documented first-line treatment failure, and duration of second-line treatment. A multivariable analysis was performed by using the logistic regression model, including all variables that were associated with the outcome  the reason to switch to second-line therapy was first-line treatment failure. The other 19 patients (9.0%) mainly switched for reasons of toxicity. Only eight of those had an undetectable plasma HIV-1 viral load at the moment of switch.Most patients were female (69.0% (145/210)) and 18.6% (39/210) of patients were children under 15 years. 27 of these children were 5 years or older at the time of switch. Unemployment amongst adults was high (68.4% (117/171)).3 [IQR 18–139]) at treatment start. First-line ART consisted of lamivudine (3TC) with stavudine (d4T) or zidovudine (AZT) as NRTIs and efavirenz (EFV) (96/210 (45.7%)) or nevirapine (NVP) (112/210 (53.3%)) as NNRTI. The median duration of first-line treatment was 19 months [IQR 11–31 months]. The median duration between first documented virological failure on first-line ART and the moment of switch to second-line treatment was 6 months [IQR 3–11 months].Patients were severely immune compromised at initiation of first-line ART, as shown by the low median CD4 counts (62 cells/mmP<0.05). During the first-line treatment period, stavudine had often been replaced by zidovudine or another NRTI, decreasing stavudine usage from 69.5% in first- to 4.8% of patients in second-line regimens. In contrast, tenofovir was prescribed to only 0.6% (1/171) of adult patients during their first-line regimen, as compared to 19.3% (33/171) for second-line ART. The median duration of follow-up after switch was 20 months [IQR 11–35 months].The median log plasma HIV-1 viral load in the total group at time of switch to second-line therapy (4.01 [IQR 3.40–4.48]) was significantly lower than that at time of ART initiation (4.94 [IQR 4.44–5.35]) (Baseline characteristics were similar for the entire group and the group of patients who had switched due to first-line treatment failure (data not shown). Baseline characteristics at start of first- and second-line treatment are summarized in Retention rate at the end of follow-up (whilst on second-line ART) was 72.8%, with a total of 139/191 patients still being in care and on treatment among the 191 patients who had switched to second-line ART following first-line virological failure. 7.9% (15/191) of them were known to be transferred out to other clinics. Approximately one eighth of the cohort  was lost to follow-up at the moment of data extraction. 5.8% (11/191) of patients were known to have died during the observational period; half of them (6/11 (54.5%)) died within the first year after the switch to second-line ART.3 [IQR 194–508] and 641 cells/mm3 [IQR 459–1245], based on 91 adult and 21 pediatric patients with data available at that time point, respectively. Among adults, the CD4 count showed a median increase of 152 cells/mm3 [IQR −4–398] from the time of switch until the end of follow-up, resulting in a median CD4 count of 384 cells/mm3 [IQR 204–586] at end of follow-up. The CD4 count change in 27 children was 27 cells/mm3 [IQR −172–210] and median CD4 count at end of follow-up was 696 cells/mm3 [IQR 357–1189]. At the end of follow-up, immunological failure according to WHO criteria was seen in twenty-four percent of adult patients. Clinical and immunological outcomes are summarized in Among these patients who had switched due to first-line failure, a cross-sectional on-treatment analysis at 12 months after start of second-line therapy showed a median CD4 count of 354 cells/mmCross-sectional analyses showed an adequate virological response (plasma HIV-1 RNA<400 copies/ml) in 71.9%  of patients after one year, and in 75.0%  after two years of second-line ART; 60.9%  and 64.3%  experienced full virological suppression (plasma HIV-1 RNA<50 copies/ml) after one and two years respectively. Of the 191 patients who had started second-line ART following first-line treatment failure, 67.5%  showed an adequate virological response and 58.1%  full virological suppression at the end of follow-up. No differences in virological response between age groups were observed. Virological results are summarized in P = 0.15). No risk factors were found to be significantly correlated with failure on second-line treatment in univariate analysis. However, an association with the outcome (P<0.50) was found for the following risk factors: gender, CD4 count at moment of switch and duration of second-line ART, and were thus included in multivariable analysis. After multivariable analysis none of these factors were found to be independently associated with treatment failure.Treatment failure was seen in 48/191  patients of which 14.7% (28/191) showed virological failure and 10.5% (20/191) on-going viremia. Consecutive measurements of plasma HIV-1 RNA above 1000 copies/ml were available for almost half (13) of the 28 patients who experienced virological failure; in the remaining 15 patients such a high plasma HIV-1 RNA was only seen once. Median duration of follow-up for the patients with treatment failure was 20 months [IQR 13–30] compared to 23 months [IQR 16–39] for the patients achieving virological success, which was not significantly different  adult patients experiencing virological failure or on-going viremia showed immunological failure. Reversely, an adequate virological response was seen in 13 of the 37 adults (35.1%) with immunological failure according to the WHO-guidelines.All analyses were repeated on the group of 210 patients including the 19 patients who had switched to second-line ART for other reasons than first-line treatment failure. These analyses generated similar results for virological, clinical and immunological efficacy (data not shown).In this cohort in rural South Africa, good virological response on second-line antiretroviral therapy following first-line failure was seen during a long period of follow-up. This observational study reflects the actual efficacy of PI-based second-line ART in our cohort, in which regular virological monitoring is routine clinical practice.Only few other reports on virological efficacy of PI-based second-line ART in sub-Saharan Africa have been published, most of them from urban areas. One group from Malawi reported on second-line treatment in a prospective observational study, where virological monitoring was performed every three months. Results of this cohort showed higher numbers of virological suppression after twelve months of second-line therapy. In most of the ART efficacy studies in RLS, a cut-off point of plasma HIV-1 RNA<400 copies/ml was used to define an adequate virological response. However, in developed countries, a more strict value (plasma HIV-1 RNA<50 copies/ml) is generally used to define virological suppression. Virological efficacy of second-line PI-based ART in children in the current study was similar to adult patients. This finding was remarkable, considering the first-line (NNRTI-based) regimen outcomes in this cohort, where treatment failure occurred more often in children than in adults. These analyses were performed amongst patient groups that were comparable to the subjects included in the current study. Treatment failure  had occurred in a quarter of patients at the end of follow-up, numbers being comparable to the earlier mentioned studies on PI-based second-line ART in sub-Saharan Africa. Mortality within twelve months after treatment switch and at the end of follow-up was low, in contrast to the previously described high early mortality rates after initiation of first-line ART in RLS. The median CD4 count increase from start of second-line ART until end of follow-up among adults was substantial and similar to that reported by other observational studies. In this study, the correlation between immunological and virological failure on second-line ART seemed to be minimal. As virological failure precedes immunological and clinical failure, patients experiencing treatment failure were probably frequently detected prior to overt CD4 count decreases and clinical deterioration had occurred, because of regular virological monitoring. There are some limitations to this study. First, as it is a retrospective observational study, there may be unmeasured underlying determinants influencing results. Second, there were some missing data, mainly due to patients being lost to follow-up. Unfortunately, causes of attrition in these patients were unknown. Third, adherence was not systematically measured in our cohort. Further, analyses on immunological response were only possible for adult patients and children of 5 years and older, as CD4 percentages were not available for the small group of pediatric patients under the age of 5. Last, there was a considerable diversity in time of follow-up after start of second-line treatment, limiting the number of patients with prolonged follow-up times (>18 months).In summary, this observational cohort study shows that second-line antiretroviral therapy in a rural area in South Africa can result in an adequate and sustained virological response in a significant number of patients, in both adults and children, following first-line treatment failure. Unfortunately, full virological suppression is seen in only about sixty percent of patients. As a result, persisting low-level viral replication may occur in some patients. It remains to be seen whether this is of clinical relevance in PI-based regimens. Regular virological monitoring is necessary to detect treatment failure before immunological deterioration occurs.


The role of financial incentives in HIV care is not well studied. We conducted a single-site study of monetary incentives for viral load suppression, using each patient as his own control. The incentive size ($100/quarter) was designed to be cost-neutral, offsetting estimated downstream costs averted through reduced HIV transmission. Feasibility outcomes were clinic workflow, patient acceptability, and patient comprehension. Although the study was not powered for effectiveness, we also analyzed viral load suppression. Of 80 eligible patients, 77 consented, and 69 had 12 month follow-up. Feasibility outcomes showed minimal impact on patient workflow, near-unanimous patient acceptability, and satisfactory patient comprehension. Among individuals with detectable viral loads pre-intervention, the proportion of undetectable viral load tests increased from 57 to 69 % before versus after the intervention. It is feasible to use financial incentives to reward ART adherence, and to specify the incentive by requiring cost-neutrality and targeting biological outcomes. Financial incentives may influence behaviors that are resistant to change by providing positive reinforcement . EffectiAccordingly, we sought to conduct a pilot study to evaluate the feasibility of using a financial incentive to improve ART adherence. Our study was motivated by several design goals that are unusual in studies of financial incentives –9: FirstOur study intervention consisted of a monetary payment $100) contingent on an “either/or” reward criterion: patients needed to either (1) suppress their plasma HIV RNA below the lower limit of detection of the assay used in our clinic , or (2) demonstrate a viral load that is at least one log10 lower than their prior lowest viral load in the past year  rather than the behavior itself (ART adherence) because we sought a target that was closely linked to preventable morbidity and mortality, and because viral load is a more objective measure than most measures of adherence . Additionally, it can be argued that the vast majority of variability in viral load is attributable to variations in patient adherence, and much of the variability that is not attributable to patient adherence is attributable to ineffective ART regimens, which can also typically be affected through patient behavior . For these reasons, we thought it was reasonable to link the incentive to the clinical outcome rather than to the behavior itself. However, we noted that this approach may not be suited to individuals who have multidrug resistance.We chose an “either/or” reward criterion . Feasibility outcomes were (1) impact on clinic workflow, (2) patient acceptability, and (3) patient comprehension. We assessed impact on clinic workflow by interviewing clinicians who had patients that participated in the study. We assessed patient acceptability by recording instances when patients were unsatisfied or disappointed as a consequence of the study intervention . We assessed patient comprehension by asking the study coordinator about whether patients understood the incentive system, as well as whether they could correctly predict whether they qualified for it based on their most recent lab result.Although the study was not powered for effectiveness, we performed exploratory analyses of viral load suppression and ART adherence using each patient as his own control, comparing the intervention year with the prior year. We compared (1) proportion of viral load measurements that were undetectable, (2) area under the curve (AUC) viral load during the last year, and (3) ART adherence, assessed by number of ART medication fills during the period of analysis.HIV-1 RNA tests done during the two years were used to assess the proportion of tests with undetectable virus. The limit of detection during the entire study was 48 copies/ml. Because results were not always available for the exact start and end dates, additional viral load tests 90 days beyond the start and end of the two year period . No imputation was performed for patients who were lost-to-follow up or who died in the intent-to-treat analysis.All eligible patients were included in “intent-to-treat” analysis. Only those that completed the entire intervention year were included in “as-treated” analysis. Proportions were compared using the χN = 40) had ≥1 detectable viral load during the year prior to the incentive. Most patients (59 %) were on PI-based regimens, with a minority on NNRTI-based regimens (29 %) or other regimens (12 %). Patients were treatment experienced, with 60 % on third or later ART regimens. Of the 51 patients (74 %) with genotypic resistance testing, 33 (65 %) had wild-type virus, 9 (18 %) had one-class resistance, and 9 (18 %) had two or more-class resistance. All but 5 patients were on ART 2 years prior to enrollment and all but 1 patient was on ART 18 months prior to enrollment.Of 80 eligible patients, 77 consented to participate in the incentive program Table , and 69 The intervention did not seem to have an adverse effect on clinic workflow. Patients were able to learn whether they qualified for the incentive either before or after their clinician visit. Time required for the qualification updates was approximately 5–10 min. No clinician expressed frustration about the impact of the study on workflow as the research activity occurred after and separate from the patient visit. The clinic administrator was moderately concerned at the beginning of the study about workflow challenges, but by the end of the study, she no longer had concerns.The incentive was acceptable to all patients except for one, who expressed frustration after his first quarter of participation because he did not qualify for the incentive. His frustration was exhibited verbally, but without any threatening words or actions. However, he chose to continue participating in the study, and during the subsequent quarter he qualified for the incentive, and became pleased with the intervention. Occasionally patients had to wait to be seen by study personnel after their clinician visit, but this was generally not an issue of concern.Patients generally appeared to understand the incentive system, and could correctly predict whether or not they qualified for it based on their current and prior laboratory results. There were no complaints about unfairness or lack of transparency. However, during the first quarter, there were some confusion about the timing of, and eligibility for the incentives. Some patients who did not qualify for incentive expressed concern that although they were adherent with all antiretroviral therapy, clinic appointments, and study requirements, they were disappointed that they were not able to receive at least a portion of the incentive . These concerns were generally resolved by the second quarter through education by the research staff.No patients expressed concerns that the incentive targeted a clinical outcome rather than a behavior itself, and there was acceptance of the premise that they could control the viral load by taking their medications with greater regularity, and/or working with their clinician to find a more effective drug regimen.2 = 4.7, p = 0.03) for intent-to-treat analyses, and 59 to 71 % in the as-treated patients . Patients without any undetectable viral loads did not appear to be impacted by the intervention, whereas patients with occasionally undetectable viral loads appeared to have greater proportions of undetectable viral load tests after the intervention , the proportion of undetectable viral load tests increased from 57 to 69 % before versus after the intervention  and from 2.1 (1.7–2.6) to 1.9 (1.8–1.9) in the as-treated , but did not reach statistical significance with the number of patients available. ART adherence increased from a mean of 18.8 fills to 20.4  before versus after the intervention for intent-to-treat, and 19.3–21.0  as-treated. The correlation between increase in fills and decrease in AUC/day viral load was r = −0.16 (p = 0.21) in intent-to-treat analysis and was r = −0.21 (p = 0.10) in as-treated patients.AUC/day viral load decreased from 2.2  to 1.9 (1.8–2.0) for intent-to-treat  or AUC viral load, . Among the subgroup of patients who had no detectable viral loads prior to the study period , 85 % of viral loads were undetectable during the study period. There was no evidence that patients on different ART regimens were differentially impacted by the intervention. Among patients on PI-based regimens, 50 % had undetectable viral loads after the intervention, whereas among patients on NNRTI-based regimens, 48 % had undetectable viral loads after the intervention.This single-site study demonstrates the feasibility of implementing financial incentives aimed at reducing viral load in HIV patients in care. While other studies have investigated financial incentives in substance-using populations, this is the first study of contingency management in an HIV population that is not selected on the basis of active or prior substance abuse. Our study demonstrates the feasibility of designing and implementing a financial incentive system in which the size of the incentive was designed to be cost-neutral or cost-saving, the primary target of the incentive was the clinical outcome of interest  rather than the behavior itself , and the incentive algorithm rewarded improvement as well as achievement yet was difficult to “game.”While our study was not powered for effectiveness outcomes, it raises the possibility that the incentive payments contributed to a 12 % increase in undetectable viral loads (from 57 to 69 %) among the target population. However, there are many other possible explanations for the finding, especially given the limitations of this small, single-site study, such as regression-to-the-mean. Follow-up studies can use more robust experimental designs to assess effectiveness. Additional questions that may be addressed by follow-up studies include whether the incentive can be implemented in different settings, and whether the incentive could be implemented using different amounts, schedules, and levels of certainty  –22. AddiOne of the novel features of this study is that the size of the incentive is linked to future costs averted by preventing HIV infections due to greater viral load reduction. While the size of our incentive may seem large ($100 per quarter), it is important to note that this cost-savings calculation was based on extremely conservative assumptions, in particular, substantial viral load reduction from partial ART adherence prior to any intervention, risk behaviors typical of individuals who know their serostatus, and modest pre-ART viral loads. An intervention targeted at one or more high risk subgroups, in particular individuals with high pre-intervention viral loads and/or prevalent risk behaviors could pay up to $2,241 per quarter (Table We report that it is feasible to use financial incentives with the aim of reducing viral load among HIV patients in care, and to specify the incentive by requiring cost-neutrality, based on the avoided costs from downstream infections averted. Future studies are needed to assess its effectiveness, scalability, and sustainability.


Background. The effectiveness of ART interventions is only realized in maximal levels of adherence. A near perfect adherence level of >95%is required for the effective suppression of HIV/AIDS virus. The main objective of this study was to identify the sociodemographic and socioeconomic factors that facilitate adherence to antiretroviral therapy among HIV/AIDS patients. Methods. This descriptive cross-sectional study was conducted between March and May 2013 at the Upper West Regional Hospital, Wa. A total of 201 confirmed HIV 1 seropositive subjects (mean age 36.6 ± 9.9 years) receiving antiretroviral therapy were interviewed using a structured questionnaire. The collected data was analyzed using GraphPad Prism version 5. A P value of <0.05 was considered statistically significant for all statistical analyses. Results. Overall lifetime adherence was found to be 62.2% while medication adherence in the last six months, last three months, last month, and last week were 73.6%, 87.1%, 91.0%, and 86.0%, respectively. The study revealed a positive association between adherence to ART and immunological success, with nonadherence increasing the risk (OR (95% CI): 9.2 (3.2–26.9)) of immunological failure. Univariate logistic regression analysis of the data showed that other ailments and side effects of drug were negatively associated with adherence to ART whereas self-perceived wellness, family support, and regular followup were positively associated with adherence to ART. Conclusion. Regular attendance at followup and family support are vital factors for 100% lifetime medication adherence. Effective counseling sessions on adherence for patients on antiretroviral therapy are paramount for the realization of the purpose of antiretroviral therapy programmes in Ghana. The effect of HIV infection at the individual level is the continued breakdown of the immune system of the host which ultimately results in the onset of AIDS. All infected persons are at risk of illness and death from opportunistic infections and neoplastic complications . InfectiSub-Saharan Africa bears the greatest burden with more than two-thirds (68%) of all persons infected with HIV. An estimated 1.8 million adults and children became infected with the disease in Sub-Saharan Africa. It is recorded that out of 260,000 child deaths that occurred globally from HIV/AIDS in 2009, 88% occurred in Sub-Saharan Africa .The HIV/AIDS epidemic in Ghana continues to be a generalized epidemic with a prevalence of more than 1% in the general population. Promising developments have been seen in recent years in global efforts to address the AIDS epidemic, including increased access to effective treatment and prevention programmes .The number of HIV patients receiving ART in Ghana increased more than 200-fold from 197 in 2003 to over 45,000 in 2010. Some regions report ART enrollment lower than their percent share of number of HIV infected persons in the country .Combination therapies of ARV drugs are the treatment of choice in HIV, and nonadherence is a major, if not the most important, factor in treatment failure and the development of resistance. 100% medication adherence is paramount for the effective management of HIV  and provIn view of the changing trend in prevalence of HIV in Ghana and the lack of data surrounding medication adherence in this population, this study therefore proposed to assess the level of and validate (using CD4 results) self-reported adherence and its predictors among patients attending the HIV Clinic of Upper West Regional Hospital, Wa.This descriptive cross-sectional study was conducted at the Upper West Regional Hospital situated in the southern part of the capital Wa. It is the only specialized referral hospital in the region and the HIV unit currently provides service to over 1600 registered HIV/AIDS patients on antiretroviral drugs. The study was conducted between the months of March and May 2013. A total of two hundred and one (201) confirmed HIV 1 seropositive subjects receiving antiretroviral therapy were interviewed using a structured questionnaire with both open and close ended questions. Primary data was also obtained from their medical records after obtaining permission from the health facility administrators and consent from patients. The Committee on Human Research, Publications and Ethics, School of Medical Sciences, Kwame Nkrumah University of Science and Technology, gave ethical approval for the study to be conducted. Confidentiality, anonymity, and privacy were guaranteed.A structured questionnaire which was developed from different literatures was used for the data collection. The dependent variable was adherence to highly active antiretroviral therapy (HAART) among PLWHA. The independent variables were sociodemographic , socioeconomic variables (income), psychosocial , disease characteristics (duration of HIV infection), regimen related variables , CD4 at diagnosis and current value, followups, adherence to treatment information and symptoms associated with treatment.lifetime self-recall adherence,last 6 months' self-recall adherence,last 3 months' self-recall adherence,last month's self-recall adherence,last week's self-recall adherence.Many researchers who have conducted studies in this area found that there is no existing gold standard by which adherence can be quantified and many predictors have been reported to influence it. The study therefore chose five measurement tools to quantify adherence from self-recalled report data collected from participants at exit face-to-face interviews:Participants were asked if they had ever missed medication in their lifetime beginning from the time s/he was put on antiretroviral therapy. Self-reported adherence was classified as “adherent” when not a single dose was missed or nonadherent if the patient admitted having missed at least one dose. They were asked about adherence to medication since initiation of ART as listed above. This means that patients' memory of medicine intake was likely to be good.However, in such face-to-face interviews patients might feel ashamed to report missed medications. Hence participants were assured of confidentiality for their response and they were made aware that, even though medically precarious, every individual could miss medication for one reason or the other.P < 0.05).Data collected was sorted, coded, and entered into an Excel spreadsheet for analysis using GraphPad Prism for Windows version 5.0 . Descriptive statistics such as mean, frequencies, and percentages were used to summarize the data. Overall lifetime adherence to medication was determined from the data and predictors determined. Analysis of contingency tables was done and Fisher's exact test and the chi-square test were used where necessary to compare proportions. Logistic regression analysis was used to determine the relationships between adherence and other variables  than the females (35.1 ± 9.0 years) although the latter constituted the majority (79.1%) of the population. Whereas only 4.5% had attained higher (tertiary) education, 52.2% were married and as many as 52.7% were living with their nuclear family. Monthly income not exceeding GH¢ 100.00 was earned by as many as 92.0% of the patients.The cross-sectional study included 201 individuals diagnosed with HIV, who had since been receiving ART at the Regional Hospital at Wa in the Upper West Region of Ghana between 2003 and 2012. The patients' demographic and clinical data are presented in Tables n = 125). Of the respondents who had participated in the study, as many as 73.6%, 87.1%, 91.0%, and 86.0% had adhered to medication in the last six months, last three months, last month, and last week, respectively (Most patients (80.1%) were diagnosed during the last four years , with 158 patients attending regular followup, that is, attending greater than 90% of 3 monthly appointments. The overall lifetime medication adherence  since initiating ART was found to be 62.2%   . BesidesAll except four subjects were on NNRTI and NRTI or in combination with septrin prophylaxis. The four exceptional subjects were on septrin prophylaxis only. As many as 41.8% were on nevirapine-based combination therapy and 32.8% on efavirenz-based combination therapy as shown in P > 0.05) with medication adherence. Although not statistically significant, positive trends for medication adherence were seen for increasing age (P = 0.3401) and monthly income (P = 0.6238). However, as shown in the univariate analysis (P < 0.0001), perceiving oneself as very healthy , and family support  were the most significant positive factors associated with increased medication adherence. On the other hand, adherence to medication was significantly reduced in patients who suffered other ailments  and those who suffered side effects of drug . Some of the patients perceived their ART regimen as simple (n = 94) and moderate (n = 100). Although self-perceived well-being was generally high  and very healthy (n = 54)), 17.4% of patients reported having side effects of the drugs, 14.4% presented with some other ailments, whilst about 91.5% were currently placed on food/substance restriction. Also, some participants (79.0%) reported having disclosed their HIV status to at least one person and 69.2% were satisfied by their family support.Univariate analysis of individual patient factors associated with medication adherence is recorded in analysis , regularP = 0.7683) retained their significance. In this adjusted model, patients who suffered other ailments  improved slightly on adherence to medication although the association maintained its negativity; side effects of drug  maintained a constant negative association with adherence as was in the univariate analysis, while regular followups  and perceiving oneself as very healthy  slightly decrease adherence  albeit maintaining their positive association with medication adherence.To construct a predictive model of the determinants of adherence, the significant independent factors were entered simultaneously into a multivariate logistic regression model. The results are presented in Antiretroviral therapy adherence levels of ≥95% optimize outcomes and minimize viral resistance . The oveP > 0.05) in levels of adherence in relation to the duration a patient had been taking ART as well as food/substance restriction as shown in Tables There are concerns that patients on ART have the tendency to be nonadherent once they feel physically better . The curP = 0.0401). This is consistent with a study report from Southwest Ethiopia where participants who had family support were two times more likely to adhere than respondents on the other side of the coin [In univariate analysis, regular followup, family support, and perception of well being were positively associated with adherence to medication as shown in the coin . This stthe coin .The majority of patients (79%) in our study had disclosed their status to at least one relative, a reflection that the study site is complying with the national guidelines for managing HIV/AIDS patients on ART. Besides that, about 69.2% of study participants reported they had support from family members in the form of emotional/psychological, physical, and food provision. These could be the promoters of optimal adherence among these participants as family members could remind them to take the ART, give comfort, and provide food to take ART with. The findings of this study suggest that a poor psychological state could be associated with poor adherence. Other studies have also shown a significant association between poor psychological states and nonadherence .P < 0.0001). This finding is consistent with the numerous studies from both developed and developing countries [Moreover, regular followup is another positive predictor of adherence to ART per the results of our study  compared to their adherent counterparts as depicted in The study results demonstrated positive correlation between adherence to ART and immunological success; nonadherent subjects were nine times (OR (95% CI): 9.2 (3.2–26.9)) at risk of progressing to immunological failure  of people in this subset cited forgetfulness as reasons for missing therapy and 42.1% said they missed medication due to the fact that they ran out of drugs. Of those who did not adhere, 9.2% said they had no food to take with the drug and 15.8% said they were away from home as reasons for missing ART treatment. These findings were similar to works done in South West Ethiopia , India , Kenya .1% of pe, and SouP < 0.001) in respondents who suffered side effects of the drugs (17.4%). This finding is consistent with the study report by [Some of the respondents (14.8%) suffered other ailments and adherence to ART was significantly reduced in this category of respondents. The most common ailments mentioned were coughing, hernia, diabetes, high blood pressure, chest pains, ulcer, rashes, general weakness, and skin itching. Our study found that adherence was negatively affected (eport by .All study participants were on the standard first-line regimen proposed by WHO (2NRTI+1NNRTI) and all patients were managed on the three combinations as found in other study reports in Africa , 13. TheThe findings of the study show that the lifetime adherence was suboptimal. Factors such as regular followup and psychological and physical support were found to be positive promoters of ART adherence. However, other ailments and side effects of the drugs had a negative association with adherence to ART.


To assess the positive predictive value (PPV) of a clinical score for viral failure among patients fulfilling the WHO-criteria for anti-retroviral treatment (ART) failure in rural Lesotho.Patients fulfilling clinical and/or immunological WHO failure-criteria were enrolled. The score includes the following predictors: Prior ART exposure (1 point), CD4-count below baseline (1), 25% and 50% drop from peak CD4-count (1 and 2), hemoglobin drop≥1 g/dL (1), CD4 count<100/µl after 12 months (1), new onset papular pruritic eruption (1), and adherence<95% (3). A nurse assessed the score the day blood was drawn for viral load (VL). Reported confidence intervals (CI) were calculated using Wilsons method.Among 1'131 patients on ART≥6 months, 134 (11.8%) had immunological and/or clinical failure, 104 (78%) had blood drawn . From 92 (88%) a result could be obtained . Out of these 92 patients 47 (51%) had viral failure (≥5000 copies), 27 (29%) viral suppression (<40) and 18 (20%) intermediate viremia (40–4999). Overall, 20 (22%) had a score≥5. A score≥5 had a PPV of 100% to detect a VL>40 copies (95%CI: 84–100), and of 90% to detect a VL≥5000 copies (70–97). Within the score, adherence<95%, CD4-count<100/µl and papular pruritic eruption were the strongest single predictors. Among 47 patients failing, 8 (17%) died before or within 4 weeks after being switched. Overall mortality was 4 (20%) among those with score≥5 and 4 (5%) if score<5 .A score≥5 among patients fulfilling WHO-criteria had a PPV of 100% for a detectable VL and 90% for viral failure. In settings without regular access to VL-testing, this PPV may be considered high enough to switch this patient-group to second-line treatment without confirmatory VL-test. Identifying patients failing on first-line treatment is a major challenge in anti-retroviral treatment (ART) programs in resource-limited settings. As a result patients who are failing on a first-line regimen are often not or not timely switched to a second-line regimen Lynen et al. published in 2009 a clinical predictor score to identify patients in need for targeted VL testing based on the score's post-test probability for viral failure The score is based on reported adherence, prior ART exposure, clinical observation of papular pruritic eruption, and trends in CD4-count and haemoglobin-level.They recommended that only patients with a score 2–4 needed VL testing, as the post-test probability for viral failure in individuals with a score<2 was very low and in case of ≥5 very high. However, the score was derived from a cohort in Cambodia, where only three patients had a score≥5. In a first validation study conducted in Uganda, only 1 patient had a score≥5 The objective of this study is to assess the PPV of a score≥5 among patients fulfilling the WHO-criteria for treatment failure in a rural cohort of Lesotho, with the aim to reduce the delays in switching to second-line treatment when needed.The study protocol was approved by the Ethical Committee of the Ministry of Health and Social Welfare of Lesotho. All patients gave oral and written consent. In the case of children <16 years of age, the care-taker gave oral and written consent.The primary objective of this cross-sectional study was to determine the PPV of a clinical score for viral failure among patients on first-line ART who fulfill the clinical and/or immunological WHO-criteria for treatment failure in rural Lesotho. Secondary objectives were to assess the overall failure-rate among patients fulfilling the WHO-criteria, the PPV of each single predictor within the score and the mortality among patients with confirmed viral failure before or within 4 weeks after switch to second-line ART.The study was conducted between October 2010 and April 2011 in the catchment area of Seboche Hospital in northern Lesotho. Seboche Hospital and its 5 affiliated nurse-led health centers serve an estimated catchment population of 55’000. The whole catchment area benefits from support of SolidarMed, a non-governmental organization based in Switzerland who supported the provision of ART since 2005. In October 2010, 1’301 patients were on ART. Of these, 691 (53%) were followed at one of the five health centers, the remaining at the hospital. ART is available free of costs at health facilities in Lesotho. Clinical monitoring is usually done on monthly basis, CD4 counts are determined every 6 months. In case of suspected treatment failure, clinicians have to write a second-line request to a second-line committee headed by the Ministry of Health and Social Welfare of Lesotho. The committee replies within one to two months. Usually, for the request to be approved, confirmatory VL testing is required. At the time the study was conducted, no VL testing was available within Lesotho. However, in case of immunological or clinical failure, the Ministry provides funds for a limited number of VL tests to be performed in a laboratory in South Africa. For this purpose, blood samples have to arrive before 2 pm at the central laboratory in Maseru in order to be further transported to South Africa. All VL obtained in this study followed this procedure.All patients on first-line ART since at least 6 months, aged ≥10 years, who fulfilled clinical and/or immunological WHO-criteria for treatment failure and who were followed within the study area, were eligible for the study. Clinical failure was defined as a new or recurrent WHO-stage 4 event after at least 6 months of ART. Immunological failure was defined as a drop of the CD4 count back to or below baseline or a ≥50% drop of CD4-count from the on-treatment CD4-peak or persistent CD4 levels <100 cells/µl after ≥12 months on treatment All patient-data to build the score were collected the same day the blood sample for VL was taken. Data were entered anonymously using EpiData 3.1® and stored on a password-protected computer. An undetectable VL (<40 copies/µl) was considered as viral suppression, a VL between 40 and 5’000 as indeterminate result and ≥5’000 as viral failure. Based on the clinical score, two subgroups of patients were formed: those with a score<5 and those with a score≥5. The PPV for a VL>40 or a VL≥5’000 copies was calculated as the proportion of individuals with a VL>40 or a VL≥5’000 respectively out of the subgroup of all patients who have a score≥5. Confidence intervals (CI) for the PPV were calculated using Wilsons score method. To assess the risk of mortality among patients continuing first-line ART while waiting for the VL-result and the approval from the MoHSW to be switched to second-line, outcomes of all patients were determined eight weeks after the blood draw for VL. In case of patients with viral failure, their status  was checked the moment they were supposed to start second-line ART . In patients who were able to start second-line ART at that moment, the status  was again determined 4 weeks after the switch. Risk of mortality during this time-period between blood-draw for VL and 4 weeks after switch to second-line was assessed using univariate logistic regression, with a score≥5 as predictor. All analyses were run on STATA 10.1™.st 2010, 1’131 were on ART since at least 6 months, and 134 (11.8%) fulfilled WHO-criteria for immunological and/or clinical failure. Of those, 104 (78%) had blood drawn on one of the scheduled days, 13 had died, 10 were lost to follow-up and 7 did not manage to come at several occasions  elapsed. Among those patients with viral failure, a median time of 55 days (39–74) elapsed between the day they were scheduled for VL blood-draw and the day a response from the second-line committee was obtained.Out of the 92 patients with a VL result, 47 (51%) had viral failure (VL≥5000 copies), 27 (29%) viral suppression (<40) and 18 (20%) an intermediate result (40–4999).Overall, 20 (22%) had a clinical score≥5. A score≥5 had a PPV of 100% to detect a VL>40 copies (95%CI: 84–100), and of 90% to detect a VL≥5000 copies (95%CI: 70–97) . The 72 The probability of viral failure (≥5’000 copies) in patients who only had immunological WHO criteria was 44% (95%CI: 33–55) as compared to 81% (95%CI: 57–93) in patients with immunological and clinical WHO criteria combined . HoweverAmong the 47 patients with viral failure, 8 (17%) died before or within 4 weeks after being switched. Among the patients with VL<5000 all 45 were retained 8 weeks after blood was drawn for VL. The overall mortality among the 92 participants was 4 out of 20 (20%) in the group with score≥5 and 4 out of 72 (5%) among those with a score<5 .This cross-sectional study assessed the PPV of a clinical score for viral failure among patients on first-line ART who fulfilled the immunological and/or clinical WHO-criteria for treatment failure in rural Lesotho. The score applied in this study was originally derived from a cohort in Cambodia with 764 individuals This study has several limitations. First, we only measured one viral load per patient. A paper by Castelnuovo et al. showed that a majority of patients who have intermediate or high viremia achieve viral suppression at a second measurement Future studies on scoring systems in low-resource settings should look at criteria that do not depend on previous CD4 counts. Our study only looked at the PPV of the score≥5 in addition to WHO criteria. Looking at the individual PPV of the criteria that build the score, the strongest predictors are VAS<95%, a new onset papular pruritic eruption and a CD4-count <100 cells/µl after ≥12 months on ART . As in sThe CD4-cell count alone, as it is used in the WHO definition of immunological failure, has shown limited accuracy in predicting viral failure A clinical score≥5 in patients fulfilling the clinical and/or immunological WHO treatment failure criteria results in a high probability of viral failure. For these patients, especially in settings with limited access to VL-testing, we may consider switching patients to second-line treatment without prior VL-confirmation. This approach may save money and valuable time for the patient, thereby reducing mortality and lost-to follow-up. However, a lower score cannot be used to exclude viral failure. These patients require a confirmatory VL measurement prior to switch to second-line.


Even with the introduction of once daily antiretroviral (ARV) regimens of low pill burden, long-term adherence remains a challenge, particularly in subgroups, such as patients with drug addiction. We aimed to determine levels of adherence and identify factors associated with suboptimal adherence in treated, HIV-infected patients attending a busy European inner-city HIV outpatient clinic.In a prospective cohort study, adherence was assessed in HIV-infected patients on antiretroviral therapy by self-report (ACTG adherence questionnaire). Relationships between suboptimal adherence (defined as <95%) and 49 covariates, including demographics, treatment factors, Centre for Epidemiological Studies Depression (CES-D) score and comorbidities were assessed using simple regression. Variables significant (P < 0.05) in univariate analyses were evaluated using multivariate logistic regression.130 subjects  were recruited. 83% were on once daily ARV and 16%, 34%, 48% and 2% were on regimens comprising one, two, three and four ARV medications respectively. Median CD4+ was 389 [285] cells/µL. 91% had HIV RNA < 50 copies/ml. Median adherence was 92% [range 0-100%] and 28% had suboptimal adherence. In univariate analyses, recent illicit drug use, on methadone, higher CES-D score, taking a higher number of ARV medications, greater pill burden, missed clinic appointments and lower CD4+ were associated with suboptimal adherence. In multivariate analysis, missed clinic appointments  a higher CES-D score  and being on a higher number of antiretroviral medications  were all independent predictors of suboptimal adherence.In a cohort where many patients are on once daily ARV, although attending clinic visits and psychological status remain important, the number of antiretroviral medications is the strongest independent predictor of adherence. Medication complexity (number of ARV) rather than the pill burden is more predictive of poor adherence in this cohort of patients from varied demographic backgrounds. Single pill, fixed dose combinations (FDC) may improve adherence and these data support further development of FDC especially for those with drug addiction and psychological issues in which current FDC medications may not be suitable.


Ritonavir-boosted lopinavir (LPV/r) or nevirapine (NVP) combined with two nucleosides are recommended for first-line regimens in antiretroviral-naïve HIV-1 patients. There are few comparative studies between these different class-based regimens. Efficacy and safety may vary from randomized studies to actual clinical practice.We analyzed retrospective data from 167 HIV-1 infected antiretroviral-naïve individuals initiating LPV/r or NVP plus two nucleoside transcriptase reverse inhibitors (NRTI) (between 1999 and 2006), according to current guidelines.LPV/r was given to 46.7%, whereas 53.3% received NVP. Average patient age was 42 years , 23.4% were women and 72.5% Caucasians. Co-infection with hepatitis viruses was present in 34.1% of all patients. The first most frequently used NRTI backbone was zidovudine-lamivudine . An alteration on NRTI backbone without study-drug discontinuation was permitted. There were no statistically significant differences between groups in the former baseline variables. Patients receiving a LPV/r-based regimen had, in average, lower baseline T CD4 cell counts  and a higher viral load  compared with those receiving NVP. Early response to treatment was evaluated by the number of patients with a viral load decline >1.0 log10 after one month of treatment: 91.7% for LPV/r (n=33/36) and 77.1% for NVP (n=27/35). Undetectable viral load after one year of treatment was 79.3% for LPV/r (n=46/58) and 82.8% for NVP (n=48/58); with an increase in T CD4 cell count by 8.9 and 1.9-fold for LPV/r (n=46) and NVP (n=54), respectively (P=0.003). The overall number of patients that discontinued therapy before completing one year of treatment were, respectively for LPV/r and NVP, 17.9% (n=14/78) and 23.6% (n=21/89). Toxicity was the most referred reason for study-drug discontinuation in both groups. After one year of treatment, toxicity grade III/IV blood biochemistry analyzed values  were 10.2% (n=27/264) for LPV/r and 8.33% (24/288) for NVP, compared to 6.44% (n=17/264) and 7.99 % (n=23/288) on baseline.LPV/r seems to have a better early response and immunological improvement. However, excluding the early discontinuation of therapy due to toxicity, NVP seems to have a lesser toxicity impact in the long-term.


As antiretroviral treatment (ART) continues to expand in resource-limited countries, the emergence of HIV drug resistance mutations (DRMs) is challenging in these settings. In Gabon , no study has yet reported the virological effectiveness of initial ART given through routine HIV care.Following the World Health Organization (WHO) recommendations, a cross-sectional study with a one-time HIV-1 RNA viral load (VL) measurement was conducted in Gabon to assess virological failure (VF) defined by a VL result ≥1000 copies/ml and DRMs among adult patients living with non-B HIV-1 strains and receiving first-line non-nucleoside reverse transcriptase inhibitor (NNRTI)-based antiretroviral therapy for at least 12 months. Risk factors associated with VF and DRMs were assessed.P=0.033), having experienced unstructured treatment interruptions (P=0.0044), and having low CD4+ counts at enrolment (P<0.0001). A longer time on ART (P=0.0008) and being followed up at the rural centre (P=0.021) were risk factors for DRMs.Between March 2010 and March 2011, a total of 375 patients were consecutively enrolled from two decentralized  HIV care centres. Median time on ART was 33.6 months . Overall, the rate of VF was 41.3% (36.4–46.4). Among viremic patients, 56.7% (80/141) had at least one DRM and 37.6% had dual-class resistance to nucleoside reverse transcriptase inhibitors (NRTIs) and NNRTIs. The most frequent DRMs were K103N/S (46.1%) and M184V/I (37.6%). Thymidine analogue mutations were found in 10.6%. Independent risk factors associated with VF were being followed up at the semirural centre (This is the first study conducted in Gabon providing VF rates and DRM patterns in adult patients receiving first-line ART. In sub-Saharan Africa, where NNRTI-based regimens are recommended as the standard for first-line ART, strengthening virological monitoring together with preventing unplanned treatment interruptions are a global public health priority. Over the past decade, the roll-out of antiretroviral therapy (ART) has dramatically increased in low- and middle-income countries. More than six million people were receiving ART in these countries at the end of 2010 ,4.In Gabon  where HIV-1 seroprevalence is high (approximately 6%) , ART becAccording to the WHO recommendations , we condPatients were consecutively enrolled from two HIV care centres. One ARV centre, located at Franceville , was a semirural site managing a total of approximately 1000 patients on ART. The second centre, located at Koulamoutou  was a rural setting with approximately 400 treated subjects. A standardized form was filled out by medical doctors to record the following variables: sex, age, ARV use , and previous CD4 counts obtained at ART initiation. Previous history of unstructured treatment interruptions (TIs) was also extracted from clinical records. Treatment interrupters were defined as subjects who defaulted and returned to care during their follow-up. When available, the duration of unplanned TIs was recorded. Patients who were transferred to other care centres were not considered as interrupters. If patients had a treatment interruption at the time of blood sampling, they were excluded from this survey.Whole blood specimens were collected in EDTA tubes at each centre. For Franceville, they were processed within six hours at the Centre International de Recherches Médicales de Franceville (CIRMF) Retrovirology Laboratory. After centrifugation, plasma was frozen at −80°C. For Koulamoutou, plasma specimens were processed similarly in the centre, stored at −20°C, and transported by road in a cool box once per week to the CIRMF Retrovirology Laboratory.http://www.hivfrenchresistance.org). Mutations were also studied using the Stanford HIVdb genotypic resistance algorithm and then coded as major resistant mutations using the International AIDs Society (IAS) list from January 2012 (http://hivdb.stanford.edu). HIV-1 subtyping was performed by phylogenetic analysis of pol region sequences. Our laboratory participated in the 2011 quality control assessment of HIV-1 drug resistance sequencing implemented by the ANRS [HIV seropositivity was confirmed using an in-house serotyping V3 immunoassay . CD4 couthe ANRS .2 for categorical variables and the Mann-Whitney-U test for continuous variables. Second, VF (≥1000 copies/mL) and DRM prevalence rates with 95% confidence intervals (95% CIs) were determined. Third, logistic regression models were used to examine the following explanatory variables as potential factors associated with VF: gender, age, centre, time on ART, unstructured TIs, and CD4 count at ART initiation (month 0) and at the time of VL assessment. The same seven variables plus HIV-1 RNA levels were further examined as predictors of at least one significant DRM among patients with VF. Finally, all variables with P-values <0.2 in univariate analyses were entered in multivariate analyses.All statistical analyses were performed using Stata 10™ software . First, a comparison of patients’ characteristics between the two care centres was made using the χ+ count was 159 cells/µl , with no significant difference between women (158 cells/µl) and men (164 cells/µl). At the time of enrolment, the median time on treatment was 33.6 months  and was slightly higher for males (35.4 months) compared with females (32.3 months). The median CD4+ count reached 337 cells/µl , with very similar values for women (334 cells/µl) and men (343 cells/µl). Previous unstructured TIs were recorded in more than 40% of patients (in women and men), with an overall median duration of seven months. There was no significant difference in the patients’ characteristics between the two care centres except the median time on ARV therapy, which was slightly longer in the semirural site than in the rural centre (P=0.03). Also unstructured TIs were more frequently recorded in patients from the semirural centre (approximately 46%) versus the rural site (approximately 33%) (P=0.05).From March 2010 to March 2011, a total of 375 adult patients on first-line ART were consecutively included . In bothOverall, 171  had a VL ≥300 copies/ml and were therefore defined as viremic on ART, with a median of 49,900 copies/ml . When using a threshold value of 400 copies/ml, 165  subjects were viremic. For drug resistance assessment, genotypic tests were only done among the 155 (41.3%) viremic patients with a VL result ≥1000 copies/ml. Overall, 141/155 (91.0%) virus were successfully sequenced (ten could not be amplified and four had insufficient volume) and showed a very high HIV-1 genetic diversity. Although the predominant HIV-1 variants were CRF02_AG  and subtype A , 15 additional subtypes/CRFs  and 19 mosaic unique recombinant forms (URFs) were also identified.By using the ANRS algorithm, among the 141 sequenced specimens, 80 (56.7%) harboured ≥1 DRM . OverallOf the 56 NRTI-associated DRMs, M184V/I selected by 3TC/FTC was the most frequent , followed by any thymidine analogue resistance TAM mutation . Ten patients had viruses with the TAM-1 mutation T215Y/F associated with AZT/D4T resistance. Other TAM-1 were M41L (n=2) and L210W (n=1). TAM-2 mutations included K219Q/E/R (n=4), D67N/G (n=4), and K70R (n=3). Three (3.7%) patients had viruses with at least three of these TAMs associated with AZT/D4T resistance. No patient harboured viruses having accumulated TAMs that confer also cross-resistance to ddI or ABC or TDF. No patient displayed viruses harbouring the K65R or K70E mutation selected by TDF. Two patients had HIV-1 strains harbouring the multi-NRTI resistance Q151M mutation associated with M184V/I and Y188L.Among the 77 (54.6%) patients harbouring viruses resistant to at least one molecule of the NNRTIs class, the most commonly detected DRM was the K103N/S . Major NNRTIs DRMs were also obtained at positions P225H (n=12), K101E (n=11), Y181C (n=10), G190A (n=7), Y188L (n=6), V90I (n=5), E138A/G (n=5), M230L (n=4), A98G (n=3), H221Y (n=3), L100I (n=3), V179D (n=2), and V106I (n=1). In two patients treated with NVP, viruses were found resistant to etravirine (ETV), the second generation NNRTI drug, due to the simultaneous presence of the Y181C and H221Y mutations. Five additional viruses were predicted to be possibly resistant to ETV, including three viruses harbouring the E138A/G/Q/R DRM, one with the three DRMs K101E/H/I/P/R, Y181C, and G190A/S, and one with the three DRMs V90I, K101E/H/I/P/R, and G190A/S.P=0.033), having experienced unstructured treatment interruptions (P=0.0044), and having low CD4+ counts at enrolment (P<0.0001) (P=0.24), age, or CD4 at M0. A longer time on ART (P=0.0008) and being followed up at the rural centre (P=0.021) were risk factors for DRMs. We found no significant association between the presence of DRMs and sex (P=0.95), age, or CD4 at M0. VL levels were not associated with DRM acquisition  . We founuisition .+ counts at inclusion were strong predictors of VF. Also, a higher VF rate was found for the semirural care centre. Second, among participants who virologically failed ART, 56.7% carried ≥1 DRM and almost 40% had dual-class resistance to NRTIs and NNRTIs. DRMs were consistent with expected patterns given first-line NNRTI-based therapy predominantly used in Gabon. The proportion of patients with DRMs significantly increased with time on ART and was higher for the rural site.Here, for the first time in Gabon, we provided VF rates and DRM patterns in adult patients living with non-B HIV-1 strains and receiving mostly (>95%) first-line thymidine analogue, 3TC, and NNRTI-based ART for a median period of 34 months. Our survey, conducted in semirural and rural settings, first revealed a high rate of VF, with 41% of patients showing VL results ≥1000 copies/ml. In our survey, a history of unplanned treatment interruptions and CD4Comparison of VF (or success) and DRM profiles rates obtained in different African populations must be made cautiously because study design, VL assays and threshold used, and duration of ART at time of failure may vary. In a review, Barth and colleagues reported virological success (defined by a VL <400 cp/ml) rates in sub-Saharan Africa of 78%, 76%, and 67% after 6, 12, and 24 months of ART, respectively [et al. obtained 24% of drug-resistant viruses in a two-months-off, four-months-on ART arm, compared with 9% in a continuous strategy arm, with a highly significant difference for resistance to mostly NNRTIs. By contrast, very few studies “in real world settings” investigated the impact of unplanned treatment interruptions on the development of DRMs [In addition, the burden of DRMs found in our survey was consistent with previous studies in which prevalence of DRMs showed highly contrasted results, with NNRTI mutations ranging from 47 to 100%, the M184V/I mutation from 24 to 81%, and any TAM from 0 to 63% –22. Howe of DRMs ,27. In oOur study is limited by its cohort cross-sectional design and does not provide any information on the overall efficiency of the national ART program. Even if it is known that rates of lost to follow-up are higher in the first year and tend to decrease thereafter, no information was available on the dropout/death rates of the clinics and on the presence (or not) of DRM before ART initiation. In addition, subjects with a VL result ranged between 300 and 1000 copies/ml may harbour mutant HIV strains with DRMs. Therefore, the drug resistance rate that we obtained in our study is most likely a minimal estimate. Another limitation of our study is represented by the fact that no data on baseline prevalence of HIV drug resistance was available for our studied population. However, all studies recently conducted among untreated Gabonese subjects revealed low levels (≤1%) of transmitted HIV DR ,28,29. FIn summary, this is the first study conducted in Gabon providing VF rates and DRM patterns in adult patients receiving first-line ART. In sub-Saharan Africa, where NNRTI-based regimens are recommended as the standard of care for first-line ART, strengthening virological monitoring aimed at early failure detection, together with preventing unintended treatment interruptions, are a global public health priority.Access to virological monitoring should be also increased to limit the level and complexity of drug resistance observed in non-B HIV-1 strains circulating in resource-limited settings. Increase in VL monitoring will prevent inappropriate switches and preserve susceptibility to second-line regimens in Africa –32. As rFinally, the WHO/HIVResNet Global HIV drug resistance (HIVDR) prevention, surveillance, and monitoring strategy must be strengthened worldwide at sentinel sites to inform national health authorities on the efficiency of first-line ART and to allow recommendations on future ART strategies. Our study showed that, in the vast majority of our patients, the second-line regimen recommended by WHO would be active in our settings. Nevertheless, a minority showed complex drug resistance profiles predictive of resistance to the usual second-line regimens. Emerging multi-NRTI resistance in sub-Saharan Africa would not only compromise second-line treatment options and the success of antiretroviral roll-out but could also contribute to the spread of drug-resistant variants worldwide . Thus fr


Background. Combination antiretroviral therapy (cART) has progressively decreased mortality of HIV-associated tuberculosis .To date, however, limited data on tuberculosis treatment outcomes among coinfected patients who are not ART-naive at the time of tuberculosis diagnosis are available. Methods. A multicenter, observational study enrolled 246 HIV-infected patients diagnosed with tuberculosis, in 96 Italian infectious diseases hospital units, who started tuberculosis treatment. A polytomous logistic regression model was used to identify baseline factors associated with the outcome. A Poisson regression model was used to explain the effect of ART during tuberculosis treatment on mortality, as a time-varying covariate, adjusting for baseline characteristics. Results. Outcomes of tuberculosis treatment were as follows: 130 (52.8%) were successfully treated, 36 (14.6%) patients died in a median time of 2 months (range: 0–16), and 80 (32.6%) had an unsuccessful outcome. Being foreign born or injecting drug users was associated with unsuccessful outcomes. In multivariable Poisson regression, cART during tuberculosis treatment decreased the risk of death, while this risk increased for those who were not ART-naive at tuberculosis diagnosis. Conclusions. ART during tuberculosis treatment is associated with a substantial reduction of death rate among HIV-infected patients. However, patients who are not ART-naive when they develop tuberculosis remain at elevated risk of death. The wider access to combined antiretroviral therapy (cART) had a profound impact on HIV-associated tuberculosis.Prospective studies conducted in high-burden and low-burden countries have clearly shown that incidence of tuberculosis is strikingly reduced in persons receiving cART –5, and mA number of observational studies have also demonstrated that starting cART during treatment for tuberculosis reduces mortality in ART-naive patients in spite of the increased risk of immune reconstitution syndrome . The effHowever, little information is available on the outcome of tuberculosis occurring in patients who are not ART-naive at the time of tuberculosis diagnosis.In a multicenter study conducted in Italy in the context of wide availability of cART, we found that more than one-third of cases of HIV-associated tuberculosis occurred in patients who were already on antiretroviral treatment . In thisDesign of this multicenter, prospective, observational study has been previously described . BrieflyIndividuals (in and out-patients) 18 years of age or older, with confirmed HIV infection, diagnosed with tuberculosis in the participating units during a 15-month period, were included in the study. All patients were given an identification code to guarantee confidentiality and each of the participating centers sought ethical clearance according to local regulations.At baseline, the following data were collected for each enrolled subject: age, sex, country of birth, place of residence, education, employment, date of first positive HIV test, mode of HIV infection, history of active tuberculosis, results of chest radiographs and clinical evaluation and microbiological examinations for mycobacterial infection and disease. Data on concomitant AIDS defining diseases, CD4+ lymphocyte count and current and/or previous antiretroviral therapy were also recorded. Followup data were obtained from hospital wards and outpatient clinic records, and included information on antiretroviral therapy, tuberculosis treatment outcome, and vital status at the end of the study period.All data were collected onto coded standardised forms. All forms were checked by scientific staff at the coordinating center for logical errors.Cultures for mycobateria were performed on radiometric method  and/or Löwenstein-Jensen medium.Mycobacterium tuberculosis from a clinical specimen and/or demonstration of M. tuberculosis from a clinical specimen by nucleic acid amplification) and/or signs and symptoms compatible with tuberculosis , completed diagnostic evaluation and decision by physician to treat with a full course of antituberculosis chemotherapy. On the basis of microbiological, clinical, radiological or histological findings, tuberculosis was classified as pulmonary, both pulmonary and extra pulmonary, extra pulmonary only.A case of tuberculosis was defined as a physician diagnosis of tuberculosis in a person who has bacteriological evidence of active disease  tuberculosis included cases resistant to at least both. Patients were defined aware of HIV serostatus if patients they had their first HIV positive test performed al least 3 months before tuberculosis diagnosis.A new case of tuberculosis was defined as a patient who has never had treatment for tuberculosis, or who has taken antituberculosis drugs for less than one month. Previously treated cases were defined as patient who had received at least 1 month of antituberculous therapy in the past. Drug-resistant (DR) tuberculosis was defined as caused by a For the purpose of the present analysis, patients were defined ART-naive if they had never received antiretrovirals or they received antiretrovirals for less than one month before diagnosis of tuberculosis. Patients were defined on ART at the time of tuberculosis diagnosis if they received antiretrovirals for at least one month in the three months preceding tuberculosis diagnosis.Tuberculosis treatment outcomes were defined according WHO definitions . For theχ2 or Fisher's Exact Test, as appropriate, were used to compare proportions. Odds ratios (ORs) with the associated 95% confidence intervals (CI) were calculated to measure the association between variables and treatment outcome. By fitting a polytomous logistic regression, we analyzed association of baseline characteristics associated with death and unsuccessful outcome of tuberculosis treatment, compared to successful outcome.Descriptive statistical methods were used to provide a general profile of the study population. The To investigate the impact of cART on mortality rate, a Poisson regression model was used. Results of this analysis are presented as mortality rate ratios (MRRs) with the associated 95% CI. Patients were included from the initiation of antituberculosis treatment until completion of treatment, death or loss to followup, whichever comes first; cART was included in the analysis as a time-dependent variable together with potential confounders. Analyses were performed with STATA software .We considered for inclusion in the analysis 271 HIV-infected patients with who were diagnosed with tuberculosis during the study period. Among these patients, 25 (9.22%) did not start tuberculosis treatment, 5 because they were transferred-out and 20 who were lost to follow up immediately after diagnosis. The remaining 246 patients entered the present analysis.The median time from first date of documented HIV seropositivity was 36.9 months (range: 0–201.3), and 96 (39%) were not ART-naive at the time of tuberculosis diagnosis. Of these patients, 34 received antiretroviral therapy for a median of 13.5 months (range 1–86), but not in the three months preceding diagnosis of tuberculosis, and their last ART regimen included a protease inhibitor (PI) in 20 patients and a nonnucleoside reverse transcriptase inhibitor (NNRTI) in 11 patients. At baseline the median value of CD4 lymphocytes was 120.5/mmc (range 0–1111), and viral load median value, calculated in 241 patients, was 4.94 log copies/mL. At least one concomitant AIDS defining illness disease was recorded in 60 (24.4%) patients. We recorded tuberculosis treatment outcomes for the 246 patients included in the analysis. A successful outcome was recorded for 130 patients (52.8%), among them 75 (30.5%) were cured and 55 (22.4%) completed treatment. Eighty patients (32.5%) had unsuccessful outcomes: 44 (17.9%) were lost to follow up in a median time of 1 month, and 25 (10.2%) were defaulters, 9 (3.7%) were transferred-out, and 2 (0.8%) were failures. Thirty-six patients (14.6%) died a median time of 2 months after tuberculosis treatment initiation. P = 0.008), which was also more likely for injecting drug users. Risk of death was also associated with being injecting drug users as well as to a lower CD4 cells count at the time of tuberculosis diagnosis and MDR tuberculosis. In multivariable polytomous logistic regression analysis , not beiAmong the patients enrolled, 151 (61.4%) received cART and tuberculosis treatment concurrently. Of these patients 62 were already on cART when tuberculosis was diagnosed and 89 started cART during tuberculosis treatment, 56 (62.9%) in the initial phase and 33 (37.1%) in the continuation phase and included a PI in. Patients who were already on cART at initiation of TB have been receiving antiretrovirals for a median of 24 months (range 3–108) before diagnosis of tuberculosis and their last cART regimen included a PI in 35 cases and an NNRTI in 23. An additional 21 patients were not ART-naive but not on ART at tuberculosis diagnosis. ART administered during tuberculosis treatment included a PI in 75 cases (49.7%). We performed a further analysis in order to estimate the impact of use of cART during tuberculosis treatment on death rate of HIV-infected patients with tuberculosis. During 161.2 person-years (PY) of observation, 36 deaths occurred with an overall mortality rate of 22.3 per 100 PY (95% CI: 16.1–31.0). Among the patients who died, 17 were not ART-naive, 7 were ART-naive and started cART during tuberculosis treatment and 12 patients never started cART. P < 0.001), whereas being not ART-naive at tuberculosis diagnosis caused a more than four-fold increase in the same risk . Risk of death was also associated with a lower CD4 cell count, age ≥ 40 at diagnosis, and MDR tuberculosis. In multivariable analysis , the useIn this multicentre study conducted in a low tuberculosis incidence country, a successful outcome of tuberculosis treatment was documented in slightly more than 50% of HIV-infected patients; the death of the patient during treatment was recorded in almost 15% patients. When we analyzed the impact of cART before and during tuberculosis treatment on the risk of death, we found the cART use during tuberculosis treatment reduced the probability of dying, while this risk was increased in those who were not ART-naive at tuberculosis diagnosis. The overall success rate of tuberculosis treatment observed in the present study is lower than that reported in other European studies in general population. A survey conducted in 10 European countries  found anHowever, the main difference with surveys of tuberculosis treatment outcome in general population is the increased proportion of death, and HIV infection per se may most likely account for the observed discrepancies in death rates. Indeed the above-referenced Spanish multicentre survey  found a We further explored the association between the risk of death and the use of cART. In our study population, almost 50% of patients continued or started cART during the initiation phase of tuberculosis treatment and an additional 14% initiated cART during the continuation phase. Overall, the use of cART was associated with a greater than six-fold reduction of the risk of death. This effect is of the same order of magnitude of that observed both in high and low tuberculosis incidence countries . Thus ouA relevant finding was the increased risk of death for individuals who were not ART-naive when diagnosed with tuberculosis, which remained significant when we adjusted in the analysis for current cART use as well as for other factors associated with the risk of death such as a low CD4 cells count, older age at tuberculosis diagnosis, and multidrug resistance. The reasons for this association remains to be elucidated, nevertheless some hypothesis can be put forward. First of all, some patients had interrupted cART treatment before tuberculosis was diagnosed and this may be a marker of poor adherence to cART or previous cART failure or cART toxicity. Thus, even if cART is resumed during tuberculosis treatment, a reduced effect could be expected in these patients. Moreover, treatment interruptions per se have been associated with an increased risk of death . OccurreTuberculosis occurring in cART-treated patients may in some instances be due to the so-called unmasking, which is defined as clinical manifestation of preexisting tuberculosis infection that is due to ART-induced immune restoration and which may sometimes result in severe or even fatal disease . We do nIn this study we do not have details of antiretroviral treatment history of patients and in particular we cannot determine if those who were not ART-naive had virological treatment failures and/or antiretroviral resistance at the time of tuberculosis diagnosis, and thus we could not estimate the impact of these factors on patients' outcome. The high proportion of patients who abandoned treatment may also have affected the analysis of factors associated with death. A further limitation is that the study was conducted on patients treated relatively early in the cART era, and thus the conclusions on the effect on cART may not necessarily be applicable to the newer cART regimens. In conclusion these study shows an alarmingly high proportion of unsuccessful outcome of tuberculosis in HIV-infected persons who inject drugs or are migrants and stresses the need of intervention aimed at keeping these patients into care. Tuberculosis occurred frequently in patients who were not ART-naive, and these patients had an increased risk of death compared to those who were ART-naive, also after taking into account the use of ART during tuberculosis treatment. Tuberculosis occurring in patients who already received ART may represent in the future an important issue to be addressed in high tuberculosis incidence countries in which scaling up of ART is currently underway.