The clinical status of HIV-1 infected patients initiated on modern anti-retroviral (ARV) therapy is consistently improved as indicated by reduced viral load (VL) and increased CD4+ T-cell count, however, the reported impact on patients' health related quality of life (HRQoL) has been variable. An appropriate assessment of HRQoL response over time in ARV-naïve HIV-1 infected subjects initiated on LPV/r combined with RAL or TDF/FTC could reveal the impact of these therapies on functional status and wellbeing.The PROGRESS study is an ongoing, randomized, open-label 96-week trial of LPV/r 400/100 mg BID combined with either RAL 400 mg BID (n=101) or TDF/FTC 300/200 mg QD (n=105) in ARV-naïve subjects. Subjects completed the MOS-HIV, a validated, disease-specific HRQoL instrument, at baseline and weeks 8, 24, and 48. The MOS-HIV comprises 35 items in eleven dimensions. Dimension specific scores and the two summary scores  each range from 0 to 100 points, with higher scores indicating better function or well being. Changes in score from baseline were analyzed using ANCOVA with the following covariates: baseline score, treatment arm, gender, race/ethnicity, age, time since HIV-1 diagnosis, baseline CD4+ T-cell count, and plasma HIV-1 RNA level (VL).Both LPV/r + RAL and LPV/r + TDF/FTC treatment arms achieved similar VL and CD4+ T-cell endpoints at 48 weeks; a similar proportion of subjects in each arm discontinued the study prematurely. For each assessment period, >79% of all subjects completed the MOS-HIV survey. There were no statistically significant differences between treatment arms in MOS-HIV scores on any dimension or summary score at any assessment period (p>0.100). When pooling data across all subjects, MCS improved significantly from baseline at week 8 , week 24 , and week 48 . PCS improved significantly from baseline only at week 24 . The General Health Perceptions dimension, an overall evaluation of health, was significantly improved at week 8 , week 24 , and week 48 .LPV/r in combination with either RAL or TDF/FTC improved HRQoL related to both mental and physical states as well as overall health in ARV naïve, HIV-1 infected subjects at weeks 8, 24, and 48. Additional HRQoL data through week 96 are being collected.


Mycobacterium genavense is a rare nontuberculous mycobacteria (NTM). Human infections are mostly disseminated in the setting of the AIDS epidemic or the use of aggressive immunosuppressive treatments. M. genavense culture is fastidious, requiring supplemented media. Pulmonary involvement rarely occurs as a primary localization.M. genavense infection: one kidney transplanted patient and one HIV-infected patient. Both patients were initially treated with anti-tuberculous drugs before the identification of M. genavense on sputum or broncho-alveolar lavage fluid culture. A four-drug regimen including clarithromycin and rifabutin was started. Gamma interferon has been helpful in addition to antimycobacterial treatment for one patient.We report here two patients with pneumonia as the predominant manifestation of M. genavense could be the etiologic agent of sub-acute pneumonia mimicking tuberculosis in patients with cellular immunodeficiency status.Clinicians should be aware that  M. genavense, is a fastidious growing mycobacteria found in water [M. genavense infection preferentially involves bowel and abdominal lymph nodes, a fact being ascribed to the presumed digestive contamination. Mortality of patients with disseminated M. genavense infection is high, ranging from 44% to 71% in 2 series [M. genavense infected patients with pulmonary involvement as the main clinical manifestation, one kidney transplant recipient and one HIV-infected patient.Host defenses against mycobacteria need an immune cellular response Th1/Th17) activated by cytokines and chemokines produced by innate immunity cells /Th17 act. M. genain water  with birin water . Human iin water . Thereafin water ,7. M. ge2 series ,9. Herei3, hemoglobin 11,1 g/dL, aspartate aminotransferase 88 U/L, alanine aminotransferase 40 U/L, gamma glutamyl transferase 310 U/L, alkaline phosphatases 210 U/L, lactico deshydrogenase 268 U/L. Her peripheral blood CD4+ cell count was 110/mm3 and HIV viral load was 6.39 log copies/mL. Serological tests for hepatitis B and C were positive. A chest radiograph demonstrated diffuse bilateral pulmonary infiltrates , clarithromycin (500 mg b.i.d), ethambutol (20 mg/kg/d) and amikacin  and HAART  were introduced. Clarithromycin was increased to 1000 mg b.i.d after 2 weeks of treatment because the treatment was well-tolerated and optimal treatment recommendations were sparse. HIV viral load decreased to 3 log copies/mL after 1 month without increase in CD4+ count (60 cells/mm3). Despite 3 months of this antimycobacterial treatment, the patient's clinical course worsened and infiltrates in the upper lobes on chest X-ray increased. A second bronchoscopy was performed and BAL fluid cytologic analysis showed the persistence of 20 to 30 AFB/field, which were again identified as M. genavense, without other pathogens. Then gamma interferon (IFN-γ) was started (50 μg/m2 sub-cutaneously 3-times a week). The HAART therapy was continued with undetectable viral load but a persistent CD4 lymphopenia (50/mm3). The patient's general condition gradually improved within 1 month of IFN-γ treatment with oxygen weaning, radiological improvement and negative respiratory specimens mycobacterial culture. The CD4 cell count reached 300/mm3 after twelve months of antimycobacterial treatment, so IFN-γ was withdrawn. The anti-mycobacterial treatment was withdrawn after 2 years. We decided to continue the treatment more than 6 months after immune reconstitution and 12 months after negative culture of respiratory specimen because chest radiograph demonstrated continuous regression of pulmonary infiltrate until 24 months, with minimal sequelae at the end of treatment  imaging demonstrated multiple abscesses. Cerebral toxoplasmosis was suspected and treatment with pyrimethamine and sulfadiazine was initiated. Biological results were: C-Reactive Protein (CRP) 19 mg/L, leukocyte count 3940/mms Figure . The thot Figure . At the 3; hemoglobin 10.9 g/dL. CD4+ lymphocyte count was 230 cells/mm3. Serological tests for HIV-1 and HIV-2 were negative. Chest radiograph  on Mycobacteria Growth Indicator Tube (MGIT) liquid medium. M. genavense was identified by INNO-LiPA probe identification on the product of the 16S-23S rDNA gene spacer region amplification. Anti-mycobacterial therapy was thus modified for clarithromycin (500 mg b.i.d), moxifloxacin (400 mg/d), rifabutin (5 mg/kg/d) and ethambutol (15 mg/kg/d). The immunosuppressive regimen was tapered with withdrawal of mycophenolate, tacrolimus was replaced by cyclosporin A, so the daily dose of prednisone was increased to 20 mg. Blood and bone marrow cultures were negative. Bone scan showed no additional localization. Intestinal malabsorption and anorexia required prolonged parenteral nutrition. After 3 months of treatment, the patient's condition gradually improved; fever, respiratory and digestive symptoms had resolved. The patient was no longer able to produce sputum for mycobacterial culture. Colonoscopic control after 2 months of treatment showed the regression of ulcerations. After 6 months, lung lesions and mesenteric lymphadenopathies had decreased on thoracic and abdominal CT scan. At the last medical visit, in October 2010, abdominal CT scan showed resolution of adenopathies. On thoracic CT scan, pulmonary consolidations had resolved. One right hilar lymph node (13 mm) remained unchanged. The antimycobacterial treatment was continued because of the sustained immunosuppressive regimen for chronic graft rejection.A 44 year-old renal transplant Guinean patient was admitted in September 2008 for fever, weight loss and non-productive cough worsening gradually for one month. He had undergone renal transplantation 3 years earlier because of focal segmentary hyalinosis. His immunosuppressive regimen included mycophenolate mofetil 1500 mg/d, prednisone 10 mg/d and tacrolimus 6 mg/d without any modification since. Blood test results were: CRP 28 mg/L; white blood cells 11300/mmh Figure  and thorM. genavense infection in two patients with cellular immune deficiency. Secondary dissemination occurred in one case. M. genavense infection has previously been reported in only two solid organ (kidney and heart) transplanted patients [M. genavense infection associates weight loss, fever, anemia, digestive manifestations and hepatosplenomegaly [M. genavense infection (of whom 20 were HIV positive) reported five patients with pulmonary symptoms in combination with extra-respiratory symptoms in all cases [M. genavense remains a rare cause of NTM infection. In the Microbiology Department of Foch Hospital, Suresnes, our two cases were the only two patients with M. genavense isolation among 176 patients with NTM isolated from clinical specimens between 2005 and 2008. The other mycobacteria identified were: Mycobacterium avium complex: 56; M. gordonae: 42; M. xenopi: 37; M. kansasii: 21; M. fortuitum: 8; M. abscessus complex or M. chelonae: 7; M. celatum: 1; non-identified: 2.We here emphasize the predominant thoracic involvement that revealed severe patients ,11. Bothpatients . In HIV-nomegaly ,12,13. Pll cases . HoweverM. genavense can be cultured from stools, sputum, urine and blood samples and tissue biopsy [M. genavense is fastidious, making microbiological diagnosis frequently difficult, requiring mycobactin J supplementation for optimal recovery on culture. Culture must be prolonged up to 4 months with a median positive culture time of 43 days (min. 10 days- max. 6 months) depending on the culture medium [e biopsy . Howevere medium ,13,15-18M. genavense can eventually be detected on clinical specimens with PCR [M. tuberculosis PCR on a positive AFB smear specimen is useful to exclude tuberculosis earlier. In case one, NTM treatment would have been considered if M. tuberculosis PCR had been performed on sputum. Furthermore, we think that M. genavense should be envisaged in immunocompromised patients with mycobacterial pulmonary disease and digestive involvement or negative mycobacterial culture lasting for more than one month.Molecular tools such as polymerase chain reaction (PCR) are of interest in two ways before cultures become positive. Firstly, with PCR -21. SecoM. genavense infection [M. genavense disease is correlated with CD4 cell count. Indeed M. genavense infection occurs in patients with profound CD4+ lymphopenia (50 cells/mm3 or less). However, CD4+ cell count is not a reliable marker of immunosuppression in non-HIV patients receiving immunosuppressive treatment. For example, Pneumocystis pneumonia is frequently observed in organ transplanted patients with more than 400 CD4+ cells/mm3 [3). Thus, M. genavense infection may occur in non-HIV patients under an immunosuppressive regimen with a much higher CD4+ cell count than seen in HIV-infected patients.Infection control requires decrease of the immunosuppressive regimen in patients who receive such treatment in combination with appropriate antimycobacterial treatment. Human and mouse models have pointed out the role of CD4+ and CD8+ T lymphocytes in nfection ,23. Explells/mm3 . In our M. genavense infection is unknown. The high effectiveness of clarithromycin and rifabutin has been demonstrated in a murine model [Mycobacterium avium intracellulare infection, treatment was continued 12 months after immune restoration was achieved for Case 1 [The optimal anti-mycobacterial therapy in ne model . Multidrne model . Even wine model . As recor Case 1 . ConcernM. genavense. Indeed, mice deficient for IFN-γ are unable to clear M. genavense infection [M. avium intracellulare complex infection [M. tuberculosis and NTM [In the case of our first patient, despite HAART introduction and appropriate anti-mycobacterial treatment, the infection was not controlled. The administration of IFN-γ was temporally associated with a significant clinical improvement and bacteriological clearance of infection. IFN-γ is essential for the control of nfection . In HIV-nfection . The cla and NTM .M. genavense infection may be a slowly progressive pneumonia mimicking tuberculosis in patients with cellular immunodeficiency; 2) Thus, this diagnosis should be evoked in an immunosuppressed patient who receives empirical anti-tuberculous therapy without improvement; 3) Extra-thoracic manifestations can be absent; 4) the microbiological diagnosis requires prolonged cultures and may be helped by PCR analysis of bronchial and lung specimens; 5) IFN-γ may be useful to control M. genavense pneumonia in addition to the anti-mycobacterial regimen; 6) CD4+ T cell count is not a reliable marker of the risk of M. genavense infection in non-HIV imunocompromised patients.Finally clinicians should be aware 1) That the main clinical manifestation of Th: T helper lymphocyte; HAART: Highly active anti-retroviral therapy; AFB: Acid fast bacilli; BAL: broncho-alveolar lavage; HIV: Human immunodeficiency virus; PCR: Polymerase chain reaction; NTM: Non tuberculous mycobacteria; IFN-γ: Interferon-gamma.The authors declare that they have no competing interests.Written informed consent was obtained from the patient for publication of this case report and accompanying images. A copy of the written consent is available for review by the Editor-in-Chief of this journalBR, EC, OL, LJC made substantial contributions to conception and design. ER, PH, DZ, MFM, PC, ML participated in acquisition of data. BR, EC drafted the manuscript. OL, LJC, ER, PH, DZ, MFM, PC, ML revised it critically. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2334/11/311/prepub


Once-daily compared with twice-daily antiretroviral therapy regimens increased adherence; however, the difference was modest and not associated with a difference in virological suppression. In addition, higher pill burden was associated with lower rates of virological suppression, whether once- or twice-daily regimens. Background. Contemporary antiretroviral treatment regimens are simpler than in the past, with lower pill burden and once-daily dosing frequency common. We performed a meta-analysis of randomized controlled trials (RCTs) to investigate the impact of pill burden and once-daily vs twice-daily dosing on ART adherence and virological outcomes.Methods. A literature search of 4 electronic databases through 31 March 2013 was used. RCTs comparing once-daily vs twice-daily ART regimens that also reported on adherence and virological suppression were included. Study design, study population characteristics, intervention, outcome measures, and study quality were extracted. Study quality was rated using the Cochrane risk-of-bias tool.Results. Nineteen studies met our inclusion criteria (N = 6312 adult patients). Higher pill burden was associated with both lower adherence rates (P = .004) and worse virological suppression (P < .0001) in both once-daily and twice-daily subgroups, although the association with adherence in the once-daily subgroup was not statistically significant. The average adherence was modestly higher in once-daily regimens than twice-daily regimens . Patients on once-daily regimens did not achieve virological suppression more frequently than patients on twice-daily regimens . Both adherence and viral load suppression decreased over time, but adherence decreased less with once-daily dosing than with twice-daily dosing.Conclusions. Lower pill burden was associated with both better adherence and virological suppression. Adherence, but not virological suppression, was slightly better with once- vs twice-daily regimens. Among human immunodeficiency virus (HIV)–infected patients, adherence to antiretroviral therapy (ART) is a primary determinant of virological suppression, disease progression, and death –3. ART rLittle is known about the impact of once- vs twice-daily ART and pill burden on adherence and virological outcomes. Indeed, in some patients with suboptimal adherence and/or virological failure, reducing the pill burden may be more important than switching from a twice-daily regimen to a once-daily regimen. Furthermore, governments, third-party payers, and HIV programs may prefer the use of non-coformulated ART generics because they are less expensive than brand name STRs. Therefore, as more generics become available, there is the potential for a paradoxical “desimplification,” with movement away from STR regimens , 11.A 2009 meta-analysis by Parienti and colleagues of 11 randomized trials reported that ART adherence rates were significantly better with once-daily than with twice-daily regimens , with a Thus, we conducted an updated meta-analysis to evaluate the impact of pill burden and once- vs twice-daily ART on adherence as well as virological outcomes in both ART-naive and -experienced HIV-infected adults.The study background, rationale, and methods were specified in advance and documented in a protocol that was published in the PROSPERO register (CRD42012002515).We included only randomized controlled trials (RCTs) that compared once-daily vs twice-daily regimens in either ART-naive or -experienced patients with objective measures of adherence and measures of virological outcomes.We systematically searched the following databases from their inception until 31 March 2013 : Cochrane CENTRAL, PubMed, Google scholar, and Web of Science. Our search terms included the following: “HIV,” “treatment simplification,” “co-formulation,” “fixed-dose combination,” “QD,” “twice-daily,” “once-daily,” “adherence,” “HAART,” “ART,” “cART,” and “patient preference.” We also searched abstracts from major HIV/AIDS and infectious diseases conferences (from 2008 onward) including Conference on Retrovirus and Opportunistic Infections, International AIDS Conference, International AIDS Society Conference on HIV Treatment, Pathogenesis and Prevention, International Conference on Antimicrobial Agents and Chemotherapy, and Infectious Diseases Society of America Conference. In addition, the bibliographies of relevant review articles, metaanalyses, and selected articles were examined for pertinent studies.We evaluated each identified study using the following predetermined selection criteria: open-label RCTs of HIV-infected subjects either ART naive or ART experienced that compared once-daily ART regimens with any twice-daily antiretroviral regimens and assessed both adherence  and viral suppression . Placebo-controlled, blinded trials were excluded because the regimen frequency was identical for the comparator arms (to maintain blinding) and, therefore, the impact of the placebo on adherence could not be measured. We chose to exclude trials that used self-reported adherence as the patients are more likely to overestimate adherence due to social desirability and typically these trials do not reflect true variability in adherence due to a ceiling effect –30.We used the Cochrane Collaboration's tool for assessing the risk of bias for quality assessment of the included studies . The stuThree reviewers  independently evaluated the eligibility and methodological quality of studies obtained from the literature search. These same reviewers also independently extracted and compared the data. For each identified study that met the selection criteria, details on study design, study population characteristics, intervention, outcome measures, and study quality were extracted. Discrepancies were resolved by consensus through discussion.P value; and imputation of the SD reported in similar studies.The primary measures of treatment effects were weighted mean difference (WMD) with 95% confidence interval (CI) for adherence to treatment and relative risk (RR) with 95% CI for virological suppression. We used the following methods to compute effect sizes, when incompletely reported: contact with the corresponding author; estimation of the standard deviation (SD) on the basis of the sample size, median, and range as suggested by Hozo and colleagues  or on thThe Spearman rank correlation coefficient (rho) was used to examine the associations between regimen pill burden (daily number of tablets), length of follow-up period, adherence rates, and virological response.2I statistic, which reports the percentage of total variation across studies due to heterogeneity rather than chance [We used DerSimonian and Laird  random en chance , 35. Basn chance , subgrouWe examined the reliability and conclusiveness of the available evidence using a trial sequential analysis (TSA) –39 and thttp://ims.cochrane.org/revman) were used for meta-analysis; Trial Sequential Analysis Software, version 0.9 beta (www.ctu.dk/tsa), was used for the trial sequential analyses.This review was performed according to the PRISMA recommendations for meta-analyses of RCTs . Stata 1Supplementary Table 1 shows the characteristics of studies that were excluded from the meta-analysis, and Supplementary Table 2 shows the assessment of bias risk among the included studies.The literature search yielded 428 articles Figure . After rA) between adherence and pill burden  for both once-daily and twice-daily regimens. However, when the analysis was stratified by the regimens, the association between adherence and pill burden was significant in the twice-daily regimens  but not in the once-daily regimens . There was also a statistically significant negative association  between pill burden and virological suppression , which was significant in both the once-daily  and twice-daily subgroups .There was a negative and statistically significant association Figure A betweenP = .0002; Figure Supplementary Figure 1). In prespecified subgroup analyses, the greater average adherence with once-daily vs twice-daily dosing was more pronounced in treatment-naive patients . There was no significant difference in virological suppression among patients following once-daily vs twice-daily regimens . Furthermore, there was no significant difference between once- and twice-daily regimens in virological suppression in the treatment-naive or -experienced subgroups . When the analysis was stratified by dosing regimens, twice-daily remained statistically significant , whereas the once-daily was not . Similarly, there was a significant negative association  between virological suppression and duration of follow-up , such that virological suppression declined with longer follow-up. The associations were similar to the overall for both twice-daily  and once-daily  regimens.Adherence declined significantly over time .This meta-analysis of 19 RCTs which included 6312 patients found that higher pill burden was associated with both lower adherence and worse virological suppression in both twice-daily and once-daily subgroups. In addition, adherence was higher with once-daily ART regimens than with twice-daily regimens when adherence was measured objectively using pill counts and/or MEMS caps. However, this difference was minimal and did not translate into better treatment outcomes. Furthermore, the greater adherence with once-daily dosing was only statistically significant in treatment-naive individuals and in those who switched from twice- to once-daily dosing with virological failure. Adherence did not increase among treatment-experienced patients who switched from twice- to once-daily dosing while virologically suppressed; adherence was likely high in these patients prior to the switch. Both adherence and virological suppression decreased with longer follow-up, but the adherence decrease was less pronounced with once-daily dosing than with twice-daily dosing.Interestingly, none of the included randomized trials directly evaluated the effect of an STR, which we consider an unanswered question for further research. However, in our study, there was a significant negative association between pill burden and virological suppression, suggesting that regimen simplification with STRs may be helpful in select situations. One small observational study conducted among marginally housed individuals and 2 large observational studies conducted found better adherence with STRs  , 56, 57,There are several possible explanations for the apparent lack of impact of once- vs twice-daily dosing on virological outcomes. First, the impact of once-daily dosing on adherence was relatively small (2.5% absolute increase in adherence); this was possibly too small to result in a clinically meaningful difference in virological suppression. Second, a substantial number of the trials included in this meta-analysis were of relatively short duration. Moreover, volunteers for clinical trials are likely to be more adherent than their counterparts managed in routine clinical practice, and there may be more resources available to support adherence in clinical trial settings . For theThese results have several important practical implications. Currently, as all recommended regimens are highly potent, ART combinations should be selected based on factors such as tolerability, potential drug interactions, patient preference for dosing frequency, and pill burden, as well as structural factors  . EffortsIn a mathematical simulation, Walensky and colleagues showed that the future use of a once-daily regimen that includes generic efavirenz plus generic lamivudine plus branded tenofovir in the United States could yield savings of almost $1 billion per year to HIV programs . Our res2 > 50%). Also, by focusing on once-daily vs twice-daily dosing, our analysis may have masked regimen-specific effects  that have little to do with the frequency of dosing. Finally, the impact of regimen frequency and pill burden on adherence and virological outcomes in RCTs may not necessarily generalize to desimplification, in which patients may perceive that their regimen has been reduced in quality. Such a change could adversely affect adherence and/or treatment outcome, and, as noted above, specific studies to investigate this question are needed.Our study has several strengths. We performed a comprehensive search of several databases and sources to identify eligible RCTs that provide the highest quality of evidence. Three authors independently evaluated each study for inclusion and data extraction. Furthermore, we performed a trial sequential analysis; this is an efficient decision-making tool that is used to establish whether firm evidence of effect has been obtained . RegardiIn this meta-analysis of 19 RCTs, we confirmed that once-daily ART regimens increased adherence when compared with twice-daily regimens, but the difference was modest and not associated with a difference in virological suppression. Importantly, we found that higher pill burden was associated with lower rates of virological suppression regardless of dosing frequency. The nonlinear correlation between pill burden and adherence or virological suppression suggests that, while ART desimplification from once-daily STRs to once-daily multitablet regimens may have adverse effects on virological outcomes, separating out STRs and/or fixed-dose combinations into their constituents is not likely to have a major detrimental impact on virological outcomes . Nevertheless, further research is needed to directly investigate the impact of such a switch, in particular among patients who are virologically suppressed at baseline. In the meantime, our results suggest that pill burden should be a consideration in the selection of an antiretroviral regimen, independent of dosing frequency.Supplementary materials are available at Clinical Infectious Diseases online . Supplementary materials consist of data provided by the author that are published to benefit the reader. The posted materials are not copyedited. The contents of all supplementary data are the sole responsibility of the authors. Questions or messages regarding errors should be addressed to the author.


Background. Eight decades ago, the ratio of monocytes to lymphocytes  was noted to affect outcomes of mycobacterial infection in rabbits. Recent transcriptomic studies support a role for relative proportions of myeloid and lymphoid transcripts in tuberculosis outcomes. The ML ratio in peripheral blood is known to be governed by hematopoietic stem cells with distinct biases.Methods. The predictive value of the baseline ML ratio was modeled in 2 prospective cohorts of HIV-infected adults starting cART in South Africa . Incident tuberculosis was diagnosed with clinical, radiographic, and microbiologic methods per contemporary guidelines. Kaplan-Meier survival analyses and Cox proportional hazards modeling were conducted.Results. The incidence rate of tuberculosis differed significantly by baseline ML ratio: 32.61 , 16.36 , and 51.80  per 1000 patient-years for ML ratios of less than the 5th percentile, between the 5th and 95th percentiles, and greater than the 95th percentile, respectively (P = .007). Neither monocyte counts nor lymphocyte counts alone were associated with tuberculosis. After adjustment for sex, World Health Organization human immunodeficiency virus disease stage, CD4+ T-cell counts, and previous history of tuberculosis, hazards of disease were significantly higher for patients with ML ratios of less than the 5th percentile or greater than the 95th percentile .Conclusions. The ML ratio may be a useful, readily available tool to stratify the risk of tuberculosis and suggests involvement of hematopoietic stem cell bias in tuberculosis pathogenesis. Mycobacterium tuberculosis infection [M. tuberculosis infection or a high risk for developing tuberculosis is a major challenge; tuberculin skin tests (TSTs) and the recently developed interferon γ–release assays are poor at predicting which individuals with latent infection will subsequently go on to develop tuberculosis, particularly in human immunodeficiency virus (HIV)–positive populations [Tuberculosis is the leading cause of death in patients commencing combination antiretroviral therapy (cART) in sub-Saharan Africa –3, yet pnfection . Howeverulations . The neeulations , 8 as opulations .Mycobacterium bovis [M. bovis, they demonstrated that reduced or elevated ratios of monocytes to lymphocytes  were associated with enhanced lethality of mycobacterial infection [Gene expression studies have identified biomarkers of active tuberculosis that provide insight into immune mechanisms of disease and may be useful for improving the diagnosis of tuberculosis , 11. Fleum bovis . They anum bovis  and humaum bovis , althougnfection . LymphoiData in this study were derived from several cohort studies or clinical trials –19 condu+ T-cell count <200 cells/µL were eligible for commencement of a cART regimen comprising 2 nucleoside reverse-transcriptase inhibitors and 1 nonnucleoside reverse-transcriptase inhibitor. During this period, the South African guidelines did not support the routine use of isoniazid preventive therapy for patients receiving cART. For this analysis, 1862 individuals who initiated cART and had at least 1 full blood count measurement available before cART were included. At enrollment, all individuals underwent clinical examination, including clinical screening for tuberculosis and other opportunistic infections. Individuals with active tuberculosis at enrollment were excluded. Participants were clinically evaluated at least twice before enrollment, and monthly after cART commencement. During follow-up after cART initiation, individuals with symptoms and/or signs of tuberculosis were assessed and received a diagnosis per South African guidelines, which include clinical examination, sputum microscopy, and/or culture and chest radiography as indicated [For the primary analysis participants were ambulatory, HIV-seropositive, cART-naive adults older than 18 years who enrolled in care and treatment between April 2004 and April 2011 at the CAT clinic in Vulindlela, rural KwaZulu Natal . Care fondicated .The replication cohort was a cohort of cART-naive adults who self-referred for care at the Prince Cyril Zulu Communicable Diseases Centre, a chest clinic that provides care for the eThekwini (Durban) municipality.HIV-uninfected women enrolled in the CAPRISA004 trial were included for analyses of the stability of the ML ratio . RoutineFor analyses of the impact of HIV acquisition and disease progression, data from the CAPRISA002 study , a prospFor analyses of the impact of antituberculous therapy on the ML ratio, data from the CAPRISA003/SAPiT trial were used . In thisIn all cohorts, data were collected in real time during prospective follow-up on standardized case report forms and captured electronically using the DataFax system  by at least 2 data encoders.Full blood counts of peripheral blood collected in ethylenediaminetetraacetic acid–containing tubes (Becton Dickinson) were performed by one of 2 clinical diagnostic laboratories, each using a 5-part differential hematology analyzer . Full blood counts were subject to strict quality assurance procedures, including twice-daily high and low internal quality control, fortnightly quality controls through the Thistle external quality assurance service , and annual quality assurance as part of the College of American Pathologists QC scheme. Both laboratories are accredited by the South African National Accreditation System in accordance with international standards ISO 17025:2005 and ISO 15189:2007.P value of < .05 was considered statistically significant. Poisson approximations were used to calculate confidence intervals (CIs) for estimations of the incidence rate. Bootstrapped estimates of the adjusted HR across the ML ratio continuum were generated with the boot package. Statistical analyses were performed in R , using the following packages: epiR, survival, date, and mfp.To calculate the ML ratio, the absolute monocyte count was divided by the absolute lymphocyte count. For the primary analysis of tuberculosis incidence, a survival analysis was performed. Study duration was calculated as the time from cART initiation to the first diagnosed episode of tuberculosis or death, withdrawal from the study, transfer to another facility, or last visit date (for patients remaining in follow-up), whichever occurred first. The log-rank test was used to compare survival curves stratified by ML ratio. To test the association between the ML ratio and tuberculosis incidence, a Cox proportional hazards regression model was performed. Covariates included in the model are given in the body text or tables. For the analysis of continuous variables, the Kruskal-Wallis rank sum test was used. All statistical tests were 2-sided, and a + T-cell count at baseline was 120 cells/µL. The median follow-up for this cohort was 17 months but extended up to 7 years. The incidence of tuberculosis, diagnosed per South African national guidelines, during follow up was 18.79 cases per 1000 patient-years of treatment , consistent with comparable cohorts [We assessed whether the ML ratio, when evaluated before commencement of cART, was predictive of the development of tuberculosis during cART in a prospective cohort of 1862 HIV-infected adults in South Africa. Baseline characteristics of this cohort are detailed in Table  cohorts , 25.TabM. bovis [+ T-cell counts, and those with greater WHO staging values were overrepresented in the group with the highest ML ratio (>95th percentile) relative to the other 2 groups. This observation is concordant with expectations of individuals with more severe lymphopenia having higher ML ratios . The overall incidence of tuberculosis among individuals with an ML ratio less than the 5th percentile was 32.61 cases per 1000 patient years , and the overall incidence among individuals with an ML ratio greater than the 95th percentile was 51.80 cases per 1000 patient years , compared with 16.36 per 1000 patient-years  for individuals with an ML ratio between the 5th and 95th percentiles (Table P = .003). Neither the monocyte count nor lymphocyte count alone were predictive of tuberculosis on formal testing .To validate these results, the association between the baseline pre-cART ML ratio and the risk of tuberculosis during cART was assessed in a replication cohort of 345 adults commencing cART at an urban treatment site implementing the same care and treatment protocol as in the primary cohort. The overall incidence of tuberculosis after cART commencement in this cohort was 21.62 cases per 1000 patient-years . Consistent with the finding in the primary cohort, pretreatment ML ratios that were less than the 5th percentile or greater than the 95th percentile were independently associated with a significantly greater incidence and hazard of tuberculosis . These data demonstrate a J-shaped association between ML ratio and incident tuberculosis, with both low and high ML ratios associated with subsequent tuberculosis.To refine estimates of the association between the ML ratio and incident tuberculosis, we generated bootstrapped estimates of effect across the ML ratio continuum Figure B. These + T-cell of <100 cells/µL, WHO stage 3 or 4 HIV infection or disease, male sex, and/or a past history of tuberculosis. To address these potential confounders, these covariates were included in the Cox proportional hazards model (Table P = .002). Moreover, the association was consistently observed in analyses stratified for each risk factor individually in the primary cohort  or replication cohort (data not shown).To exclude the possibility that the effect of ML ratios on the tuberculosis risk was mediated by modification of HIV treatment outcomes, we confirmed that baseline ML ratios were not associated with virological or immunological outcomes (per 2010 WHO criteria) 6 or 12 months after cART initiation in the primary cohort . Since the index takes into account lymphocyte and monocyte values, and since the latter may be independently regulated, variation of the ML ratio is of interest.We assessed how the ML ratio is affected by changes in clinical condition. Intuitively, the ML ratio is expected to vary following HIV acquisition, tuberculosis, cART initiation, or tuberculosis therapy because of changes in lymphocyte counts , and there was no difference in the distribution of the ML ratio between any 2 visits . Even when monocyte and/or lymphocyte counts were outside of their respective 5th–95th percentiles, the majority (65.7%) of individuals had an ML ratio between the 5th and 95th percentiles (both cell types increased or decreased). Conversely, at 4.3% of visits the monocyte and lymphocyte counts were between the 5th and 95th percentiles, but the ML ratio was not . These data demonstrate that although the ML ratio is stable among healthy individuals, about two thirds of individuals with altered numbers (<5th or >95th percentile) of either cell type retain apparently normal ML ratios, and conversely about 1 in 20 individuals with normal cell numbers may still have altered ML ratios. The ML ratio captures information not adequately captured in the monocyte or lymphocyte counts alone.The stability of the ML ratio was assessed among 790 healthy, HIV-uninfected South African women who underwent routine safety monitoring over 2 years while enrolled in a topical microbicide gel trial . The ML A). In contrast, ML ratios were significantly increased following HIV acquisition , consistent with postinfection lymphopenia. Moreover, the distribution of ML ratios broadened with HIV infection . The ML ratio increased significantly over the course of HIV disease progression . The ML ratio was not associated with the rate of HIV disease progression, defined as the time to achievement of a CD4+ T-cell count of <350 cells/mL, death, or the need for cART (data not shown).Next, ML ratios were compared before and after HIV acquisition in a cohort of 160 high-risk women prospectively followed for HIV acquisition in several studies . ML rati3The change in ML ratio with duration of cART was assessed in a cohort of 2023 individuals who underwent screening and commenced cART. The ML ratio was significantly altered by cART: over the first 24 months of therapy, the median ML ratio decreased to within the range observed for HIV-uninfected healthy volunteers and was maintained at this level for up to 7 years . Before tuberculosis therapy, 15% of individuals (96/642) had ML ratios in the normal range for an HIV-uninfected population, compared with 41.7%  following tuberculosis therapy (P < .0001).Finally, the impact of tuberculosis therapy on the ML ratio was evaluated. Among 642 adults with tuberculosis and HIV infection who commenced antituberculosis therapy before cART [The ability to stratify patients starting cART on the basis of their risk of tuberculosis during follow-up is necessary for targeting prevention interventions. Here, we demonstrate that extremes in the ratio of peripheral blood monocytes to lymphocytes, but neither monocyte or lymphocyte counts alone, were associated with an increased risk of incident tuberculosis over several years. Our observation that the ML ratio is associated with tuberculosis risk supports findings of recent microarray studies in HIV-negative infants  and extends findings made >80 years ago in rabbit models of mycobacterial infection . In moreM. tuberculosis infection may alter hematopoietic stem cells such that the peripheral ML ratio is altered. In support of this is the observation that in the microarray studies, alongside the ratio of myeloid transcripts to lymphoid transcripts being a signature of risk, transcripts suggesting activation/quiescence of hematopoietic stem cells were more prevalent among infants who developed tuberculosis. Alternatively, the ML ratio and underlying hematopoietic stem cell activity may be epiphenomena, and the ML ratio may be a marker of where along the spectrum [Whether the ML ratio is causatively involved in tuberculosis or simply reflects underlying traits that affect tuberculosis risk will require further study. It is likely that the ML ratio captures underlying differences in the immune microenvironment, as has been implied by its prognostic value in influenza  and malaspectrum  of tuberspectrum . FurtherThe clinical usefulness of the observation that the ML ratio is related to prospective risk of tuberculosis is promising but will benefit from extension and validation in non–tuberculosis-endemic regions, among children, and in HIV-negative adults. In particular, the diagnostic performance of the ratio and the development of standards or diagnostic thresholds require attention. Evaluation of whether the ML ratio is associated with other disorders, as is suggested by recent studies in influenza  and malaAlthough we observed that higher ML ratios were associated with tuberculosis, it is possible that tuberculosis was present but not diagnosed at the time of enrollment (reverse causation). But the observation that the association appears to hold out for several years, and is present even excluding early tuberculosis diagnoses, suggests that undiagnosed tuberculosis at enrolment does not account for the association.An important potential limitation of this study is that this was a secondary data analysis study, so it is plausible that there may have been misclassification of the tuberculosis diagnosis if tuberculosis was not specifically and prospectively assessed. But the data used here were obtained during the conduct of prospective studies in which assessment for any clinical disorder, particularly tuberculosis, was performed. The diagnostic algorithm for tuberculosis used in these studies followed the national South African guidelines, and so the diagnostic procedures in this study approximate those in clinical settings, which are known to suffer impaired sensitivity for incident tuberculosis. A further limitation of this study is that participants did not have TSTs or interferon γ–release assays performed. Finally, at the time of clinical assessment, although clinicians were not blinded to the full blood count, they were not aware of this hypothesis, thus diminishing diagnostic bias.M. tuberculosis infection to prevent tuberculosis ,or closer clinical monitoring to diagnose tuberculosis earlier. The ML ratio may be a useful, readily available tool to stratify risk of tuberculosis and provide a new insight into tuberculosis pathogenesis.Among patients starting cART, the ability to predict the risk of tuberculosis may facilitate targeted interventions such as vaccination, treatment for latent Supplementary materials are available at The Journal of Infectious Diseases online . Supplementary materials consist of data provided by the author that are published to benefit the reader. The posted materials are not copyedited. The contents of all supplementary data are the sole responsibility of the authors. Questions or messages regarding errors should be addressed to the author.


To estimate the population-level causal effect of source of payment for HIV medication on treatment adherence using Marginal Structural Models.+ T cell count and prior plasma HIV RNA. Traditional association models were also examined and the results compared.Data were obtained from an observational cohort of 76 HIV-infected individuals with at least 24 weeks of antiretroviral therapy treatment from 2002 to 2007 in Kampala, Uganda. Adherence was the primary outcome and it was measured using the 30-day visual analogue scale. Marginal structural models (MSM) were used to estimate the effect of source of payment for HIV medication on adherence, adjusting for confounding by income, duration on antiretroviral therapy (ART), timing of visit, prior adherence, prior CD4Free HIV treatment was associated with a 3.8% improvement in adherence in the marginal structural model, while the traditional statistical models showed a 3.1–3.3% improvement in adherence associated with free HIV treatment.Removing a financial barrier to treatment with ART by providing free HIV treatment appears to significantly improve adherence to antiretroviral therapy. With sufficient information on confounders, MSMs can be used to make robust inferences about causal effects in epidemiologic research. Access to antiretroviral therapy (ART) continues to expand at a rapid rate Many studies in resource limited settings have documented that the cost of medications is a major predictor of non-adherence to ART We used marginal structural models (MSMs) using a targeted maximum likelihood estimate (MLE) to estimate the causal effect of source of payment for ART on treatment adherence among HIV-infected individuals in Kampala, Uganda from 2002 to 2007. This was a time of rapid transition from exclusively self-pay to free HIV ART. Marginal structural models (MSMs), developed by Robins et al We utilized data from the Adherence Monitoring Uganda (AMU) study Marginal structural models (MSM) were used to estimate the difference in adherence means for a given month that would have been observed between the treatment group  and the control group  if source of payment for therapy had been assigned randomly. The marginal treatment effect is the parameter of interest. A targeted maximum likelihood estimator (TMLE) was used to estimate this parameter. A data set was created that consisted of a data point for each person-month during follow-up for which source of payment for medication and subsequent adherence were measured. Confounders considered included prior adherence, prior CD4 T cell count, prior plasma HIV RNA level, income, duration on ART and time of visit (period from study enrolment when study staff visited participant and assessed adherence).Counterfactual assumption: we assumed that counterfactuals exist and that the outcome observed for each patient was one of the potential outcomes. We thus assumed that this was a missing data problem. In conjunction with the counterfactual assumption, we assumed that the treatment or exposure was independent of the counterfactual outcomes given the covariates (Randomization assumption). That is, there were no unknown confounders (NUC). We are not very certain that all confounders were identified. However, using a directed acyclic graph (DAG) we tried to identify the important confounders and these were included in the analysis : We assumed that treatment was not assigned deterministically based on prior conditions. The probabilities of receiving the treatment were between 0.2 and 0.6 suggesting that the ETA assumption was met.clever covariate) was then calculated clever covariate was then used to target the parameter of interest. The targeting step was performed by regressing the outcome (adherence at a given visit) on the clever covariate using the initial estimate of the mean adherence level for each individual used as an offset. The coefficient on the clever covariate represents the degree of confounding in the parameter of interest. The targeting step is repeated until this coefficient is zero. For this analysis, convergence was attained in one step. Standard errors for calculating the 95% confidence intervals were estimated using clustered bootstrap (i.e. randomly sampling patients with replacement). All analyses were conducted using R software Version 2.7.2.The marginal effect of treatment  was estimated by calculating the difference between 1) the mean outcome when all patients was assumed to have been on free treatment and 2) the mean outcome when all patients was assumed to be on self-pay treatment. Targeted maximum likelihood estimation (TMLE) was used to reduce bias in the parameter of interest Parallel analyses to estimate the effect of treatment were conducted using generalized estimating equations (GEE) and ordinary least squares (OLS).10 copies RNA/ml of 5.53 (IQR 4.91–5.82). The majority of the cohort was female (63.9%). The mean age was 36 years (SD 7.5). Fifty-five percent of the participants earned less than 60 US dollars a month. One third of the study participants had completed up to a primary level of education. Details of participant characteristics at study entry have been published elsewhere Seventy-six participants were included in the analysis with a total of 251 observations. The 76 participants were observed for a total of 1669 person-months. Median follow up time was 22 person-months (IQR 18–27). Participants initiated therapy at advanced stages of HIV infection, with a mean CD4 cell count of 56 cells/ml [SD 130] and median log+ cell count and at later visits  over the course of follow up was 95.68%±16% with a median of 100% (IQR 100%–100%). Mean adherence in the self-pay person-months was 93.50%±19.16% while that in the free person-months was 98.56%±9.78% . In the Applying the targeted maximum likelihood estimator, receiving free HIV medication was estimated to increase adherence by 3.82% compared to self-pay treatment. This was slightly higher than the estimates from two traditional association models - generalized estimating equations (GEE) and ordinary least squares (OLS) .Using a marginal structural model, we estimated a 3.8% difference in mean adherence when HIV-infected patients receive free HIV treatment compared to when they pay for the treatment out-of-pocket. Our finding confirms other studies finding higher proportion of self-reported adherence Super Learner which is the best approach for selecting big models.The MSM is a semi-parametric model whose validity depends on meeting specific assumptions. However, some of the assumptions cannot be tested from the data. Non-testable assumptions are the time ordering and the counterfactual assumptions. Nevertheless, we are certain that the time ordering assumption was met because three of the authors participated in the design and implementation of the study. We are not very certain that the randomization assumption was met given the limited number of confounders that were collected. Statistical analysis showed that the ETA assumption was met. We believe that our model was correctly specified because we employed All participants in this analysis initiated ART with advanced disease. This was not intentional by the investigators neither was it a choice of the patients. At the time of enrolment of patients into this study, HIV drugs were not readily available and they were very costly. Those few who could access them were supported by family and friends through huge financial sacrifices. As such these patients initiated treatment very late as sustainability of treatment was not guaranteed. However later in the course of the study, the government of Uganda gradually introduced free ART for all HIV patients. Because of the large number of patients, poorer and sicker patients (as per their CD4 count) were often given priority over their fellow patients with slightly better immunity. Another cause for late initiation of ART was stigma. Patients did not want to be identified as HIV positive hence kept away from care until very late when there was no choice but to show up if they wanted to live. It is possible that the results of this study could have been affected by selection bias given that the majority of the study participants were of a low social economic class and hence could not afford medications which consequently led to switch to free ART with priority. However, a qualitative study conducted in the same population to understand how and why the patients had exceptional adherence revealed that the main reason for adhering to ARV medications was the desire to live and take care of other family members Our analysis using traditional models gave effects of 3.3% and 3.1% using a repeated measures model and the ordinary least squares model respectively. These effect sizes were less than the one from the MSM, underestimating the net effect of payment source on adherence, with the estimate from the ordinary least squares models failing to meet statistical significance (95% CI −0.57 to 7.24) Other analyses have shown greater attenuation of effects and even reversal of effects when using traditional statistical models There are several of limitations to our study. Our study was conducted among a small sample of ARV-naïve individuals who initiated treatment at an advanced stage of disease. This may limit generalizability of the findings. Although the analysis showed a significant association between source of payment for ART and adherence, there is a possibility that the results could have been biased. The following biases may have occurred although the extent of bias cannot be quantified. Selection bias is very likely to have influenced the results. Study participants were recruited from health facilities in Kampala. However, because of the extent of stigma that many HIV patients suffered at that time, many patients, especially those who could afford to purchase their medications, preferred to meet their care givers in private or to just send another person to the health facility to refill their prescriptions. Consequently, patients that were recruited in the study may not have been representative of all HIV-infected patients who were on ART. As such, the odds of selection for the exposed (those on free-pay) were not equal to the odds of selection for the non-exposed (those on self- pay) hence selection bias. There was an over representation of exposed non-cases (adherent) compared to the non-exposed cases (non-adherent). Information bias was another possible bias. This could have occurred as a result of loss to follow up as 50% of the patients who died (10% of those enrolled) died within 6 months of initiating ART and the majority of these were on self-pay treatment clever covariate which targets the parameter of interest. The MSM can correctly adjust for measured time-varying confounders that are affected by exposure. Given that this was a secondary data analysis that was not planned for when the study was designed, all confounding factors may not have been included in the analysis thus compromising the validity of our findings.Though useful in establishing causality, the marginal structural model has limitations. It makes the strong assumption of no unmeasured confounders. Causal effects can be estimated from the MSM parameters only if all relevant covariates are measured in the data and are adequately controlled in the analysis, including having appropriate models for determining the treatment mechanism and consequently the Our study also had several strengths. The study was conducted at a time when the healthcare system in Uganda was transitioning from self-pay to free treatment, which provided a “natural experiment” for study. It would be unethical at the present time to conduct a randomized trial to answer the question addressed in this analysis. Furthermore, state-of-the-art data analyses and the use of alternative methods to control for confounding improved the robustness of the findings.In summary, we found that receiving free HIV treatment was associated with better adherence among low income HIV-infected patients in a resource-limited setting. In a separate qualitative study conducted in the same population, lack of enough finances to purchase medications was reported as the main reason for missing doses


Since it was initiated in 2002, the China Free Antiretroviral Treatment (ART) Program has been progressing from an emergency response to a standardized treatment and care system. As of December 31, 2009, a total of 81,880 patients in 31 provinces, autonomous regions, and special municipalities received free ART. Gender differences, however, in mortality and immunological response to ART in this cohort have never been described.To understand whether women and men who enrolled in the China National Free ART Program responded equally well to the treatment.A retrospective analysis of the national free ART databases from June 2006–December 2008 was performed. HIV-infected subjects who were 18 years or older, ART naïve at baseline, and on a 3TC regimen enrolled in the program from June 1 to December 31, 2006, were included in this study, then followed up to 2 years.Among 3457 enrolled subjects who met the inclusion criteria, 59.2% were male and 40.8% female. The majority of the subjects were 19–44 years old (77%) and married (72%). Over the full 24 months of follow-up, the mortality rate was 19.0% in males and 11.4% in females (p = 0.0014). Males on therapy for 3–24 months were more likely to die than females  after adjusting for baseline characteristics. Compared to men, women had higher CD4+ counts over time after initiating ART (p<0.0001).Our study showed that women had an overall lower mortality and higher CD4+ counts than men in response to ART treatment, which may be attributed to adherence, biological factors, social, cultural and economic reasons. Further study is needed to explore these factors that might contribute to the gender differences in mortality and immunological response to ART. In the past two decades, advances in antiretroviral treatment (ART) have resulted in dramatic declines in death rates in countries where treatment is available, transforming a once-fatal disease into a manageable chronic illness In response to the growing HIV epidemic in China, the Chinese government responded in 2002 with a national ART program called the National Free Antiretroviral Therapy Program (NFATP), which provides antiretroviral (ART) drugs free to those most in need This analysis was approved by the institutional review board (IRB) of the National Center for AIDS/STD Control and Prevention (NCAIDS), China CDC. Per the IRB review, individual informed consent was waived because this analysis used currently existing data collected during the course of routine treatment and care and data were reported in aggregate without use of individual identifying information.As described previously A retrospective analysis was performed on subjects enrolled in the NFATP who had data in the Free ART Database from June 2006–December 2008. Eligible subjects had to be enrolled in the NFATP between June 1–December 31, 2006 to allow for two years of follow-up, at least 18 years of age, ART-naïve at time of enrollment, and received a HAART regimen that included 3TC (Lamivudine) as their initial therapy including D4T +3TC+NVP; D4T +3TC +EFV; AZT+3TC +NVP; or AZT+3TC +EFV. A 3TC regimen was required because it became readily available in China in 2005, has lower toxicity than other regimens, and has become the norm for initial therapy in China.After enrollment, subjects were followed up to 2 years to observe ART response including immunological function (CD4+ counts), side effects to the ART, adherence and mortality. Data from enrollment to visits through December 31, 2008 were included in the analysis to allow the last-enrolled subjects to have up to 2 years of follow-up after treatment initiation. For those who were enrolled earlier, follow-up visits after 24 months were eliminated from this analysis. Subjects with missing gender, missing treatment initiation date, first-line regimen with less than three single drugs or including a combination of AZT+D4T  were excluded from this study. A total of 4791 subjects enrolled into the NFATP from June 1 to December 31, 2006. Of those, 3457 subjects were eligible for this study. When subjects were lost to follow-up or terminated treatment during the two-year follow-up, they were censored from the study, and the length on study was defined as the date of enrollment to the date of the last available visit. Subjects who transferred from one site to another within the program were considered as adherent to study visits.Demographic and baseline characteristics were obtained, including gender, age, marital status, mode of HIV transmission, signs and symptoms at enrollment, and ART regimens. In this population, the HIV transmissions were mainly through commercial blood donations, noted as the former plasma donor (FPD); intravenous drug use (IDU); and sexually transmitted infection (STI). Laboratory test data included absolute CD4+ cell counts, hemoglobin and liver function, which were collected at baseline and follow-up visits. CD4+ T-cell counts were collected during at least one of the follow-up visits . When the CD4+ count was not available for a scheduled follow-up visit, an alternative CD4+ count within ±1.5 months of the scheduled visit time window was used for the analysis. HIV viral load was not available for analysis as a treatment outcome, since the lack of local resources during this time period prevented testing for viral load in most subjects. Based on Division of AIDS Table for Grading the Severity of Adult and Pediatric Adverse Events (GSAPAE) for HIV positive subjects only, hemoglobin was categorized as three groups , and liver function tests including AST and ALT were categorized as ≥100 U/L vs. <100 U/L (2.5×ULN). Self-reported current signs and symptoms of HIV infection such as fever, diarrhea, rash, fatigue, weight loss and lymphadenopathy, were collected at baseline. Self-reported medication adherence and fourteen self-reported adverse events  were also collected at follow-up visits. The self-reported medication adherence was collected as an ordinal variable with 0, 1–5, 6–10, 11–15, 16–20 and >20 times missing doses a month, but was redefined as a binary measure (100% vs. <100%) for this study. When a subject did not report having any missing dose, this subject was considered as perfectly adherent to ART medication (100%); otherwise, the subject was considered as less than 100% adherent (a subject reported missing at least one dose during follow-up).Chi-square was used to examine the differences in proportion of deaths between males and females. Kaplan-Meier survival curves and Log-Rank test were used to display and test gender differences in mortality over 2 years after initiating ART treatment. Univariate and multivariable Cox proportional hazard models were constructed to examine the gender effect on mortality. The demographic and baseline characteristics as well as medication adherence were included as the potential risk factors. The proportional hazard (PH) assumption over time was tested using a graphic approach and time-dependent variables where appropriate. When the PH assumption was not met because of a time/gender interaction, PH models were constructed separately for the subset of subjects who were in the program for up to three months (0–3 months) and for the larger subset who were in the program for up to 24 months (3–24 months). Another subset analysis was conducted among those who had CD4+ counts at baseline and at least three test results available after enrollment. The mixed linear model (MIXED) with repeated measurements was used to examine the effect of gender and other potential covariates on CD4+ count change over time. The unstructured covariance was assumed for the covariance structure on CD4+ counts. Model fit statistics  were used to choose the best covariance structure.Among 3457 enrolled subjects who met the inclusion criteria, 59.2% were male and 40.8% females . MajoritOverall, 443 (12.8%) of 3457 subjects were died in the first two years after initiating ART treatment. Among them, 151 (10.7%) of 1410 females and 292 (14.3%) of 2047 males were died. A significantly greater death incidence from Kaplan-Meier (KM) estimates in males 24 months after enrollment was detected when compared with female subjects . The estimated two-year overall survival rate, unadjusted for covariates, was 84.3%; two-year survival rate for females was significantly higher than for males . Of the 443 deaths observed in the first two years, about 248 (56%) occurred in the first three months after ART initiation. In addition, for multivariate analysis adjusting for covariates, the PH assumption of constant hazard was not satisfied over the entire 24-month period. There was a time/gender interaction (p = 0.0411), with the hazard rate for females changing over time. Therefore, the separated analyses were further performed: those in the program for 0–3 months and those in the program for greater than 3 months. The PH assumption was satisfied for these two separate groups. There was no difference in survival between males and females among those early deaths , Panel ACD4+ T-cell count was examined to assess gender differences in immunological response to ART. Due to limited resources, CD4+ counts were only done at a few visits for some subjects. Thus, we were unable to adjust for CD4+ cell count change over time in the multivariate proportional hazard models described earlier. We were, however, able to make inferences on CD4+ change over time for subjects who had baseline and at least 3 follow-up CD4+ count test results. The results from the multivariable mixed model with repeated measurements indicated that the average CD4+ cell count change over time in males was significantly lower than in females (p<0.0001) after adjusting for other factors including being older, infected through FPD, having past and current signs and symptoms, lower baseline CD4+ counts, without abnormal liver function, and less than 100% ART adherence .We attempted to examine the self-reported adverse events at the selected follow-up visits . When the events were missing at a specific month, the imputation was made by using an available data point within ±1.5 months around that specific follow-up visit. The results indicated that females tended to report more adverse events at most follow-up visits. For example, the overall proportion of self-reporting at least one adverse event among females was significantly higher than males at 3 months , 6 months , and 12 months .Our current study on this homogenously-treated population confirmed that females have better survival at 2 years on ART than men do. Compared to men, females had overall better immunological response overtime while on ART. Our findings are consistent with others. Hall et al Our finding that mortality did not differ by gender in the first 3 months on treatment may suggest that a certain proportion of patients entered therapy at a late enough stage of disease progression that there is little opportunity for therapy to work. This hypothesis is supported by our findings that mortality was much higher in the first 3 months and that it was strongly associated with greater baseline signs/symptoms and lower baseline CD4+ counts. After 3 months the hazard rate of females diverged from that of males.There are a number of possible explanations for our findings: (1) biologically better response to the drugs; (2) better adherence to treatment; and (3) other factors such as co-morbid conditions, general health status, and psychosocial differences. Our analysis, together with the recent findings of Rotger, et al. Our study has limitations because of the nature of the information that was retrospectively obtained from the Free ART database from 31 provinces of China, certain levels of recall bias and measurement variations may be introduced in data collection. Second, HAART initiation relies on both WHO staging and CD4+ count; however, WHO stage data was not collected until 2010. Although signs/symptom data were analyzed, this may not truly reflect WHO stage. Third, we were not able to analyze viral load response because it was infrequently collected in the field. Fourth, the CD4+ analysis was restricted to the subjects who had CD4 tests 3 times or more after the treatment initiation. Although the demographic characteristics  from this subset were comparable to the rest of the population, this subset may not truly represent the entire population of subjects who were included in the analysis. Fifth, self-reported treatment adherence data may not be completely accurate, and it is possible that some treatment response differences might still be attributable to difference in adherence. Finally, we had limited information collected on other possible explanatory factors, such as co-morbidities, general health, and psychosocial status.Our findings of lower mortality rates among females after 3 months on ART and better immunological response among females in a subset of our cohort demonstrate a difference in outcomes between men and women in the first 2 years of treatment with ART. Given the limitation of data it is unclear whether this is due to environmental, psychological, or biological factors. Further controlled prospective data collection should be undertaken to determine whether biological factors related to gender play an important role in determining outcomes to treatment. Further prospective studies within our own treatment cohort are necessary to explore other possible explanatory factors such as adherence, co-morbidities, general health status and psychosocial factors.Recommendations: Viral load is a gold standard to evaluate whether patients are virologically suppressed after ART. It is important to collect viral load data at certain time points after ART. Intensive adherence counseling for patients on failing regimens would increase the optimization of first line regimens and save money in the long term.


Improving the HBe seroconversion rate of patients without HBeAg loss after long-term lamivudine therapy has become an urgent clinical problem that we have to face. Unfortunately, there is no consensus on the mananement of these patients. The aim of this study was to evaluate the efficacy of pegylated interferon (PEG-IFN) α2a in patients without HBeAg loss after the withdrawal of long-term lamivudine therapy.Fifty patients with chronic hepatitis B without the loss of HBeAg after ≥96 weeks of lamivudine treatment were enrolled to withdraw from treatment to induce a biochemical breakthrough. Patients who achieved a biochemical breakthrough within 24 weeks received 48-weeks of PEG-IFN α2a therapy, and were then assessed during a subsequent 24-week follow-up period.5 copies/mL (76.2%) compared to patients with HBV DNA ≥105 copies/mL (27.3%) after 24 weeks of therapy (P=0.004).Forty-three (86.0%) patients achieved a biochemical breakthrough within 24 weeks of lamivudine withdrawal. The rates of combined response (both undetectable HBV DNA and HBeAg loss) and HBsAg loss were alone 51.2% and 20.9%, respectively after 48 weeks of PEG-IFN α2a therapy, and 44.2% and 18.6%, respectively, at 24 weeks after treatment cessation. The end-of-treatment combined response rate was 65.4% among patients with a baseline HBsAg <20,000 IU/mL, which was significantly higher than 29.4% of patients with HBsAg ≥20,000 IU/mL (P=0.031). For patients with HBsAg levels <1,500 IU/mL at 12 and 24 weeks therapy, the end-of-treatment combined response rate was 68.2% and 69.0%, which were both significantly higher than patients with HBsAg ≥1,500 IU/mL . The end-of-treatment combined response rate was significantly higher among patients with HBV DNA<10Retreatment with PEG-IFN α2a was effective and safe for patients without HBeAg loss after the withdrawal of long-term lamivudine therapy. HBsAg levels at the baseline, 12 and 24 weeks of therapy, and HBV DNA levels at 24 weeks of therapy, can predict the effect of PEG-IFN α2a after 48 weeks of therapy. Chronic infection with hepatitis B virus (HBV) is a major global health problem, which affects more than 400 million people worldwide ,2. ApproThe current treatment options for chronic HBV infection consist of interferon-α (IFN-α) and nucleos(t)ide analogues (NUCs), such as lamivudine, adefovir, entecavir and telbivudine. Since lamivudine was initially approved for the treatment of chronic HBV infection in China, several million affected patients have received long-term antiviral treatment with NUCs to prevent disease progression. Most patients hope that the period of antiviral treatment is temporary. In HBeAg- positive patients, durable HBe seroconversion is a satisfactory end-point as it is associated with an improved prognosis . HoweverIFN-α exerts an antiviral effect by degrading viral mRNA and proteins, and upregulates the immunological response to HBV by enhancing human leukocyte antigen class I expression on hepatocytes, its use has been confirmed as effective in the treatment of chronic HBV infection ,5-8. The3 copies/mL;iv) positive for HBsAg and HBeAg, but negative for anti-HBs and anti-HBe;v) normal liver function tests;vi) no history of liver failure or cirrhosis,and the lack of cirrhosis confirmed by a computed tomography scan; vii) neutrophil count ≥ 1.5×109/L and platelet count ≥100×109/L. The exclusion criteria included: i) superinfection with hepatitis A, C, D or E, cytomegalovirus, HIV, or Epstein–Barr virus as confirmed by enzyme-linked immunosorbent assay; ii) other liver diseases such as alcoholic liver disease, drug-induced hepatitis, Wilson disease and autoimmune hepatitis; iii) severe medical or psychiatric illness; iv) a history of diabetes, cardiac disease, hypertension, or renal, pulmonary disease or thyroid disease; v) pregnant or breastfeeding women; vi) an unwillingness or inability to provide informed consent or fulfill the the requirements of the study.Inclusion criteria included: i) age 18–60 years; ii) continuous lamivudine treatment >96 weeks; iii) serum HBV DNA levels <10Fifty outpatients with chronic hepatitis B  were recruited at the Southwest Hospital, Third Military Medical University, China, between January 2006 and June 2010. They were asked to withdraw from lamivudine and any other medications. Liver function tests were performed every 4 weeks. When a biochemical breakthrough was observed, patients underwent a serum HBV DNA test and retreatment with either PEG-IFN α2a  or lamivudine, in accordance with the study design shown in Figure 3 to 109 copies/mL). We performed HBV genotyping by direct PCR sequencing in a residual serum sample taken from each patient at the baseline timepoint before they started the PEG-IFN α2a treatment.Serum HBsAg and anti-HBs, HBeAg and anti-HBe were tested using the ARCHITECT (Abbott Laboratories). The range of HBsAg detection was 0.05 to 250 IU/mL. Serum HBV DNA was tested using real-time PCR  were tested 4-weekly during lamivudine withdrawal and the first 12-weeks of PEG-IFN α2a therapy. Patients undergoing PEG-IFN α2a therapy were then tested every 12 weeks during a further 60-week period. Peripheral leukocytes and platelets were tested at weeks 4, 8, 12, 24, 48 and 72 of therapy. Serum free thyroxin 3 and 4 (FT3 and FT4), thyroid stimulating hormone (TSH) were tested at the baseline, then 12 weekly until the end of treatment. The abnormal laboratory tests results and symptomatic adverse effects were recorded and closely monitored at each follow-up visit. We assessed all adverse events for the likelihood of causality by the study drug.P-value less than 0.05 was considered statistically significant. All statistical tests were two-sided.We performed all statistical tests with SPSS, version 10.0 . We expressed continuous variables as the median±standard deviation. The levels of serum HBV DNA and HBsAg were logarithmically transformed for the analyses. A 5 copies/mL). They received a treatment with PEG-IFN α2a  patients were observed to have a biochemical breakthrough (ALT levels ≥2×ULN) within 24 weeks of lamivudine withdrawal, and to have a virological breakthrough (HBV DNA ≥103 copies/ml and HBeAg loss) and HBsAg loss were 51.2% and 20.9%, respectively, at the end of 48 weeks therapy, and 44.2% and 18.6%, respectively, 24 weeks after the end of treatment. The rates of undetectable HBV DNA, HBeAg seroconversion and loss, and HBsAg seroconversion were shown in Figure 3 copies/mL. Among the 22 patients with a combined response at the end of 48 weeks therapy, 19 (86.4%) patients had a sustained combined response 24 weeks after the end of treatment. A normalization of the ALT levels was observed in 25 patients (58.1%) at the end of 48 weeks treatment, and among 32 patients (74.4%) by 24 weeks after the end of treatment.Among 43 patients treated with PEG-IFN α2a therapy, the rates of combined response . For patients with HBV DNA <105 at 24 weeks of therapy, the rate of combined response at the end of 48 weeks of therapy reached 76.2%, which was significantly higher than in patients (27.3%) with HBV DNA ≥105copies/mL, P=0.004, shown in Figure 5copies/mL at 12 weeks and 24 weeks of therapy, which were lower than among patients with HBV DNA ≥105copies/mL (7.53 ± 0.76 and 7.56±0.79 LC/mL) at 12 weeks and 24 weeks of therapy; these were not significant differences at either time point . For patients with HBsAg <1,500 IU/mL at 12 weeks and 24 weeks of therapy, the rates of combined response at the end of 48 weeks were 68.2% and 69.0%, which were significantly higher than in patients with HBsAg ≥1,500 IU/mL .The rate of combined response at the end of 48 weeks treatment was 61.5% among 26 patients with HBV DNA <10The rates of HBsAg loss at the end of 48 weeks treatment were 31.0% (9/29), 38.9% (7/18) and 63.6% (7/11) among patients with HBsAg < 1500, 100,10 IU/mL, but only reached 0 (0/14), 8.0% (2/25) and 6.3% (2/32) among patients with HBsAg ≥1500, 100,10 IU/mL at 24 weeks of therapy, having statistically significant difference .3 copies/mL  or HBsAg <1500 IU/mL  compared to that of patients with HBV DNA ≥103 copies/mL  or HBsAg ≥1500 IU/mL  at the end of 48 weeks treatment, P<0.01.The rate of sustained combined response was significantly higher among patients with HBV DNA <10From the withdrawal of lamivudine to the initiation of PEG-IFN α2a treatment, none of 50 patients had any evidence of hepatic decompensation or a severe acute exacerbation of chronic HBV infection characterized by ALT levels being >10×ULN or total bilirubin levels being >51 μmol/L. During 48 weeks treatment with PEG-IFN α2a, four (9.3%) patients developed a severe ALT flare-up (ALT levels >10×ULN), of which two patients developed it at 4 weeks of treatment , one patient developed it at 8 weeks of treatment (552 IU/L), and another patient at 12 weeks of treatment (616 IU/L). At 24 weeks after the end of treatment, two (4.7%) patients developed a severe ALT flare-up (487 and 2042 IU/L). However, during 48 weeks treatment with PEG-IFN α2a and 24-week follow up period, T-Bil levels did not exceed 51 μmol/L among 43 patients. Other adverse symptoms and events related to the use of PEG-IFN α2a included upper respiratory tract symptoms , fever , alopecia , abdominal discomfort , malaise , headache , myalgia , arthralgia , reduced appetite , local erythematous reactions , weight loss , a decreased neutrophil count  and a decreased platelet count . No patient required a dosage adjustment for PEG-IFN α2a, and no patient developed thyrotoxicosis or hypothyroidism.Improving the HBe seroconversion rate of patients without HBeAg loss after long-term lamivudine therapy has become an urgent clinical problem that we have to face. Unfortunately, there is no consensus on the management of these patients. In this study, we administered a new retreatment strategy by switching patients without HBeAg loss after long-term lamivudine therapy with a withdrawal-induced ALT flare to PEG-IFN α2a therapy. The rationale of this new retreatment strategy was that lamivudine withdrawal usually induces a biochemical breakthrough and a host antiviral immune response, and the biochemical breakthrough characterized by high serum ALT levels is associated with a better response to IFN-α ,9-13. OuAlthough patients who relapse after lamivudine withdrawal can be treated with any available effective therapeutic agent, but treatment with NUCs has an indefinite duration and causes low rates of HBeAg seroconversion or loss, and furthermore, HBeAg seroconversion induced by lamivudine retreatment is not permanent ,10,11. T7 copies/mL and baseline HBV DNA <107 copies/mL. No significant difference was also found between patients with baseline ALT ≥3×ULN and those with baseline ALT <3×ULN in this study, which indicated that baseline ALT level alone or HBV DNA level alone was not a good indicators for predicting the effect of PEG-IFN α2a therapy. Possible reasons for these results included the fact that the sample size in this study was too small to achieve a statistically significant difference, and that the interaction of both ALT and HBV DNA affected the response to treatment. Our results showed that the rate of combined response was significantly higher among patients with baseline ALT ≥3×ULN or HBV DNA <107copies/mL than patients with both baseline ALT <3×ULN and HBV DNA ≥107 copies/mL, which indicated that combined baseline ALT and HBV DNA levels may predict the effect of PEG-IFN α2a therapy, and was better than ALT or HBV DNA levels alone. Our results also showed that the rate of combined response at the end of 48 weeks of therapy was significantly higher among patients with HBV DNA <105 at 24 weeks of therapy than those with levels ≥105copies/mL, indicated that the HBV DNA level at 24 weeks of treatment might be able to predict the response rate at the end of 48 weeks of PEG-IFN α2a therapy, this had been described in other studies [Recent studies indicated that HBsAg levels at the baseline and during treatment can predict the effect of PEG-IFN α in the treatment of chronic hepatitis B ,17. Our  studies . Our resIn this study, we also found that the rate of a sustained combined response at 24 weeks after the end of treatment was closely associated with HBeAg status, HBV DNA levels and HBsAg levels at the end of 48 weeks therapy. Therefore, we suggested that extended treatment with PEG-IFN α2a may be helpful for patients who only achieved virological response or combined response but having high HBsAg levels at the end of 48 weeks therapy, in order to achieve a better sustained combined response. Long-term maintained therapy with NUCs should be considered for patients without virological response or HBeAg loss at the end of 48 weeks therapy.Hepatic decompensation and severe acute exacerbation of chronic hepatitis B induced by lamivudine withdrawal are major clinical concerns. In this study, none of 50 patients had a severe acute exacerbation of chronic hepatitis B during the 48 weeks follow-up after lamivudine withdrawal, which indicated that the withdrawal of lamivudine was safe for these patients without HBeAg loss and underlying liver cirrhosis, if liver function tests are monitored monthly for 24 weeks after lamivudine withdrawal. Our results also indicated that lamivudine withdrawal-induced hepatic decompensation and severe acute exacerbation were uncommon events and could be controlled for patients without underlying liver cirrhosis, which was in accordance with other previous reports . This stIn conclusion, a retreatment strategy of switching to PEG-IFN α2a therapy in patients with lamivudine withdrawal-induced ALT flare was shown to be safe and efficacious for patients without HBeAg loss and underlying liver cirrhosis after long-term lamivudine treatment. A sustained combined off-treatment response and HBsAg loss were achieved. HBsAg levels at the baseline, 12 and 24 weeks of therapy, and HBV DNA levels at 24 weeks of therapy were critical factors for predicting the effect of 48 weeks therapy with PEG-IFN α2a. However, our conclusions remain conservative because of the small sample size and nonrandomized controlled study.DNA: Deoxyribonucleic acid; HBV: Hepatitis B Virus; HBsAg: Hepatitis B surface antigen; HBeAg: Hepatitis B e antigen; NUCs: Nucleo(s)tide analogues; PEG-IFN: Pegylated interferon; ALT: Alanine aminotransferase; ULN: Upper limit of normal; T-Bil: Total bilirubin; PCR: Polymerase chain reaction; FT3 and FT4: Free thyroxin 3 and 4; TSH: Thyroid stimulating hormone.There are no conflicts of interest to disclose for all authors. There are not any commercial relationships with other departments except Southwest Hospital of Third Military Medical University.Financial competing interests1. In the past five years, we did not received any reimbursements, fees, funding, or salary from any organization that may in any way gain or lose financially from the publication of this manuscript. There is not any organization financing this manuscript (including the article-processing charge).2. We do not hold any stocks or shares in an organization that may in any way gain or lose financially from the publication of this manuscript, either now or in the future.3. We are currently applying for any patents relating to the content of the manuscript. We did not receive reimbursements, fees, funding, or salary from any organization that holds or has applied for patents relating to the content of the manuscript.4. We have not any other financial competing interests.Non-financial competing interestsThere are not any non-financial competing interests  to declare in relation to this manuscript.XQZ conceived of the study, participated in its design, the selection and treatment of patients, performed the statistical analysis and drafted the manuscript. HYZ collected the clinical data of all patients, JPY worked for follow up of patients. QM conceived of the study, participated in its design and revised this manuscript. All authors read and approved the final manuscript.