Cotrimoxazole (CTX) prophylaxis is recommended by the World Health Organization (WHO) for HIV-1-infected individuals in settings with high infectious disease prevalence. The WHO 2006 guidelines were developed prior to the scale-up of antiretroviral therapy (ART). The threshold for CTX discontinuation following ART is undefined in resource-limited settings.3 in a malaria-endemic region in Kenya. Participants were randomized and followed up at 3-mo intervals for 12 mo. The primary endpoint was a composite of morbidity  and mortality. Incidence rate ratios (IRRs) were estimated using Poisson regression.Between 1 February 2012 and 30 September 2013, we conducted an unblinded non-inferiority randomized controlled trial of CTX prophylaxis cessation versus continuation among HIV-1-infected adults on ART for ≥18 mo with CD4 count > 350 cells/mmp < 0.001), driven by malaria morbidity. There were 34 cases of malaria, with 33 in the CTX discontinuation arm . Diarrhea and pneumonia rates did not differ significantly between arms . Study limitations include a lack of placebo and a lower incidence of morbidity events than expected.Among 538 ART-treated adults screened, 500 were enrolled and randomized, 250 per arm. Median age was 40 y, 361 (72%) were women, and 442 (88%) reported insecticide-treated bednet use. Combined morbidity/mortality was significantly higher in the CTX discontinuation arm  are currently infected with HIV, the virus that causes AIDS. HIV, which is usually transmitted through unprotected sex with an infected individual, gradually destroys CD4 lymphocytes and other immune system cells. Because destruction of the immune system leaves HIV-infected individuals susceptible to “opportunistic” infections, early in the AIDS epidemic, most HIV-positive individuals died within ten years of infection. In 1996, effective antiretroviral therapy (ART)—drug regimens that stop HIV replicating and allow the immune system to recover its ability to fight infections—became available. For people living in affluent countries, HIV/AIDS became a chronic condition, but because ART was expensive, HIV/AIDS remained fatal in resource-limited countries. In 2003, the international community began to work towards achieving universal access to ART. Now, at least a third of people living with HIV have access to ART, and the global mortality (death) rate from HIV/AIDS is falling.3 blood should take CTX and that, in settings with a high prevalence of HIV infection and limited health infrastructure, all HIV-infected adults should take CTX. However, these guidelines were developed before access to ART was scaled-up, and CTX prophylaxis has risks  as well as benefits. In this unblinded non-inferiority randomized controlled trial, the researchers investigate the effects of CTX discontinuation among ART-treated HIV-infected adults in Kenya. A randomized controlled trial compares the outcomes of individuals randomly assigned to different treatments; in an unblinded trial, everyone involved in the trial knows who is taking which treatment; a non-inferiority trial investigates whether one treatment is not worse than another treatment.To prevent opportunistic infections, the World Health Organization (WHO) recommends cotrimoxazole (CTX) prophylaxis for HIV-infected individuals. CTX is a combination of two antimicrobial drugs that is active against a range of bacterial, fungal, and parasitic infections; prophylactic antimicrobials are taken to prevent infection. CTX decreases morbidity (illness) and mortality among HIV-infected individuals primarily by reducing the rates of malaria, pneumonia, diarrhea, and severe bacterial infections. The 2006 WHO guidelines for CTX prophylaxis recommend that, in resource-limited countries, HIV-infected patients whose CD4 count is ≤350 cells/mm3, and who were taking CTX. Half the participants continued to take CTX; the remainder stopped taking CTX. After 12 months of follow-up, the combined rate of morbidity  and mortality was significantly higher in the CTX discontinuation arm than in the CTX continuation arm (a significant difference is one that is unlikely to have happened by chance). The difference in this primary outcome between the trial arms was driven by malaria morbidity—there were 33 cases of malaria in the CTX discontinuation arm but only one case in the CTX continuation arm. By contrast, the rates of pneumonia and diarrhea did not differ significantly between the trial arms.The researchers enrolled 500 HIV-infected adults living in a malaria-endemic region of Kenya  who had been treated with ART for ≥18 months, who had a CD4 count of >350 cells/mmBecause the difference between the two trial arms in terms of the primary endpoint was greater than the researchers’ predefined non-inferiority limit, these findings suggest that CTX discontinuation is inferior to CTX continuation among ART-treated, immune-reconstituted HIV-infected adults living in a malaria-endemic region. The accuracy of these findings may be affected by the lack of blinding—the clinicians’ care decisions and the patients’ threshold for seeking care may have been influenced by their knowledge of their trial arm assignment. Overall, however, these findings suggest that malaria endemicity should be considered when making decisions about whether to stop CTX after ART-driven immune reconstitution. Indeed, in December 2014, following the release of preliminary data from this trial and the completion of an independent trial on CTX discontinuation, WHO issued supplemental guidelines on CTX prophylaxis. These state that CTX prophylaxis can be discontinued for HIV-infected adults who are clinically stable on ART and have evidence of immune recovery and viral suppression but recommend that CTX prophylaxis be continued regardless of CD4 cell count or HIV/AIDS clinical stage in settings where malaria is endemic and/or severe bacterial infections are common.http://dx.doi.org/10.1371/journal.pmed.1001934.This list of resources contains links that can be accessed when viewing the PDF on a device or via the online version of the article at HIV infection and AIDSInformation is available from the US National Institute of Allergy and Infectious Diseases on NAM/aidsmap provides basic information about HIV/AIDS, summaries of recent research findings on HIV care and treatment, information on opportunistic infections and on CTX prophylaxis, and personal stories about living with HIV/AIDSAvert, an international AIDS charity, on many aspects of HIV/AIDS, including information on HIV/AIDS in Kenya, on ART, and on opportunistic infections and CTX prophylaxis: Avert also provides personal stories about living with HIV/AIDSInformation is available from HIV/AIDS, including its 2013 guidelines on using ART to treat HIV infection and its December 2014 supplement to these guidelines, which contains updated recommendations on CTX prophylaxis; a factsheet about CTX prophylaxis is also availableWHO provides information on all aspects of 2015 World AIDS Day Report provides up-to-date information about the AIDS epidemic, including progress towards universal access to ARTThe this trial is availableMore information about  Cotrimoxazole (CTX), fixed-dose trimethoprim-sulfamethoxazole, is a low-cost and widely utilized broad spectrum antibiotic used to prevent opportunistic infections in patients with human immunodeficiency virus type 1 (HIV-1). Prophylaxis with CTX has been shown to decrease mortality, morbidity, and hospitalizations among HIV-infected adults  and chil3) to prevent Pneumocystis jiroveci pneumonia and toxoplasmosis [3) [Recommendations for CTX use in adults with HIV-1 vary by setting. In the United States and Europe, CTX is recommended for HIV-1-infected adults with severe immunosuppression (CD4 count < 200 cells/mmlasmosis –14. Follosis [3) ,15–17.3 [In resource-limited countries, the threshold for CTX discontinuation following ART remains undefined. The 2006 World Health Organization (WHO) guidelines recommend CTX prophylaxis for HIV-1-infected patients with a CD4 cell count of ≤350 cells/mm3 . For set3 . HoweverAt the time of our protocol submission, there were no randomized trial data on the effects of discontinuing CTX following ART in resource-limited settings. However, prior to study initiation, a randomized controlled trial (RCT) conducted in Uganda demonstrated an increased incidence of malaria and diarrhea among ART-treated adults who discontinued CTX compared to those who continued . In this3) to determine whether discontinuation of CTX was non-inferior to continued CTX prophylaxis in decreasing morbidity. Following completion of our RCT, the WHO published supplemental guidelines on the use of CTX [We conducted an RCT among adults on ART with evidence of immune recovery , and individuals in the CTX discontinuation arm discontinued CTX at the time of enrollment. The primary aim of the trial was to determine whether CTX prophylaxis can be discontinued in ART-treated, immune-reconstituted adults without significant harm.3). Additionally, participants had to be currently taking CTX for HIV and willing to return to the clinic every 3 mo for the 12-mo study follow-up period. We excluded individuals who were pregnant or breastfeeding, who were taking second-line ARVs, or who had a documented allergy to CTX.We recruited HIV-infected individuals between February 1, 2012, and August 27, 2012, at the HIV Treatment and Care Clinic, known as the Patient Support Center (PSC), of Homa Bay District Hospital in western Kenya. Individuals were eligible for the study if they were ≥18 y old, HIV seropositive, and taking first-line antiretrovirals (ARVs) with evidence of immune recovery  to either the CTX continuation or CTX discontinuation arm. Each participant’s arm allocation was concealed from study investigators, staff, and participants in a randomization envelope until the end of the enrollment visit, when the envelope was opened; participants and study staff were not masked to treatment assignment thereafter.A history and physical examination was conducted at randomization and at 3-mo intervals during follow-up. At randomization, 6 mo, and 12 mo, specimens were obtained for a complete blood count with CD4 count per Kenyan guidelines . If clinParticipants were encouraged to come to the clinic to see study providers for any illness as a sick visit. Sick visits included more intensive workup, with malaria rapid diagnostic test, thick/thin blood smears, and measurement of alanine aminotransferase level and HIV-1 RNA viral load, as clinically indicated and available. Moreover, participants were contacted every 2 wk by study counselors using text messaging or phone calls and asked about recent illness and compliance with study arm protocol. Any participant reporting any symptoms was asked to return to the clinic for evaluation by study staff. To decrease the risk of malaria and diarrhea in this cohort, we provided insecticide-treated bednets and water filters to all participants.At the completion of the study, participants were transferred back to PSC care providers. Those who had stopped CTX were counseled on restarting CTX again as per Kenyan guidelines .The primary morbidity/mortality endpoint was a composite of malaria, pneumonia, and diarrhea events and non-trauma mortality events. Malaria was defined as a fever, measured or self-reported, and either a positive rapid diagnostic test or thick smear showing the presence of parasites. Pneumonia was defined as fever, cough, or tachypnea and either a clinically abnormal chest examination or evidence of infiltrate on chest radiograph significant enough to require hospitalization. Diarrhea was defined as a self-report of three or more loose stools in a 24-h period in the past week. An independent clinician  who was blind to arm assignment at the time of adjudication evaluated all serious adverse events (SAEs) and designated their likelihood of being research-related. Each event was deemed potentially related to the research if it could have been due to either CTX use or CTX discontinuation.Prespecified secondary endpoints were 12-mo CD4 change and ART treatment failure. Treatment failure was defined by Kenya National AIDS & STI Control Programme guidelines and was categorized as clinical, immunologic, or virologic . VirologWe estimated the needed sample size for the comparison of interest: the incidence rate ratio (IRR) comparing the rate of morbidity/mortality observed in the CTX discontinuation arm to the rate observed in the CTX continuation arm. We chose a non-inferiority limit of 1.25, thus planning to conclude non-inferiority of CTX discontinuation if the upper bound of the two-sided 90% confidence interval (CI) for the IRR was less than 1.25 . We usedFor analysis of the primary endpoint, we calculated a combined morbidity/mortality incidence rate for each treatment arm. We used Poisson regression with robust error variance to estimate the IRR, using the natural logarithm of follow-up time as an offset variable. We conducted both intention-to-treat (ITT) and per-protocol (PP) analyses. ITT analyses assigned all follow-up time from a participant to that participant’s original randomization arm. In contrast, PP analyses reflected changes to CTX regimen. Specifically, for participants randomized to discontinue CTX who restarted CTX due to pregnancy, PP analyses excluded person-time following the restart of CTX.3, >600 cells/mm3) and duration of ART at enrollment . We also calculated IRRs for the rate of SAEs and the rate of all grade 2 or higher AEs. To estimate IRRs, we used Poisson regression as described above or exact Poisson regression where appropriate. We compared 12-mo change in CD4 count in the two groups using a linear mixed effects model that included main effects for arm, time, and a time–treatment arm interaction. This model used CD4 data collected at the scheduled enrollment, month 6, and month 12 study visits. We used Cox proportional hazards regression to compare time to ART treatment failure between arms.In secondary analyses, we calculated IRRs comparing the CTX discontinuation arm to the continuation arm for each individual component of the morbidity/mortality outcome, as well as for subgroups defined by enrollment CD4 count (≤600 cells/mmAn independent data safety and monitoring board (DSMB) was convened four times during the course of the study to review study procedures, participant retention, and safety data by arm and included review of any relevant new data. The DSMB adopted a safety stopping rule based on SAEs deemed potentially research-related (due to either discontinuing CTX or continuing CTX) by the DSMB safety monitor, who would be blind to arm assignment at the time of adjudication. Non-severe  malaria, pneumonia, and diarrhea events were not included as criteria for early stopping, based on the rationale that rates of non-severe AEs in the CTX discontinuation arm may reflect rates in HIV-uninfected community members and could be offset by non-severe AEs related to CTX use. The interim safety analysis was conducted in accordance with the Lan–DeMets implementation of the O’Brien–Fleming boundary for harm; the alpha spending function was based on an information fraction equal to the interim number of events in all arms divided by the total number of expected events by the conclusion of the study. The interim analysis was conducted at an information fraction of 0.64, and the study stopping criterion was not met.We used Stata version 12.1  for all 3, and median ART duration was 4.5 y. Randomization resulted in baseline characteristics that were generally well-balanced between the two arms  participants randomized to the CTX discontinuation arm had restarted CTX due to pregnancy.p < 0.001), we could not conclude non-inferiority of CTX discontinuation. The difference by arm was largely driven by the malaria component: the rate of malaria was 0.4 per 100 person-years in the CTX continuation arm and 13.0 per 100 person-years in the CTX discontinuation arm . PP results were similar to ITT results, with an IRR of 2.27 (95% CI 1.52–3.39) for the composite endpoint of morbidity and mortality and an IRR of 33.56 (95% CI 4.60–245.01) for malaria. Among individuals in the CTX discontinuation arm, malaria cases occurred throughout the study follow-up period, without evidence of an early “rebound”  in the subgroup with CD4 count > 600 cells/mm3 .In subgroup analyses Tables  and 4, mReported bednet use at enrollment was high, at 88%. Reported bednet use during follow-up was high (96.4%) and was similar between both arms.n = 108), non-malaria febrile or flu-like illness (n = 53), upper respiratory tract infection (n = 49), diarrhea (n = 38), and malaria (n = 34). Anemia was more frequent in the CTX discontinuation arm: there were 42 anemia events in the CTX continuation arm and 66 anemia events in the CTX discontinuation arm. The rates were 16.6 per 100 person-years in the CTX continuation arm and 26.1 per 100 person-years in the CTX discontinuation arm, a statistically significant difference . There were no hypersensitivity reactions.There were 34 SAEs, of which 27 were grade 3 or higher AEs (including one death), and 18 were deemed potentially related to the research (five in the CTX continuation arm and 13 in the CTX discontinuation arm) . The ratp < 0.001).The estimated rate of grade 2 or higher AEs was 63.5 per 100 person-years in the CTX continuation arm and 99.5 per 100 person-years in the CTX discontinuation arm, a statistically significant difference  in the CTX continuation arm and of 31.6 cells/mm3/y (95% CI 10.7–52.6) in the CTX discontinuation arm (p = 0.85).Analyses of 12-mo CD4 change included CD4 counts at enrollment, at month 6, and at month 12 for all participants contributing data at these time points. There was an increase in CD4 count of 28.8 cells/mmtion arm . There wp = 0.73).There were a total of nine ART treatment failure events: six immunologic treatment failures, two virologic treatment failures, and one clinical treatment failure. Five ART treatment failure events were in the CTX continuation arm, and four were in the CTX discontinuation arm. There was no statistically significant difference between arms in time to treatment failure .Only one other randomized control trial to date has examined CTX discontinuation after ART initiation in adults in a resource-limited setting . A recenData from this study were presented at the Conference on Retroviruses and Opportunistic Infections in 2014  and subsThe prior CTX discontinuation study in Uganda was stopped early after the researchers observed increased malaria, and therefore it had a limited follow-up time of up to 4 mo . It is pIn contrast to other CTX discontinuation studies, we did not observe an increase in diarrhea in participants in the CTX discontinuation arm ,10. OverThis study complements findings by Walker et al. , who notOur study demonstrated low rates of treatment-limiting AEs such as hematological AEs, rash, or hypersensitivity among participants who continued CTX. All participants were on CTX at the study start, therefore likely limiting the group of participants who would have reactions to the antimicrobial and suffer from adverse effects. AEs were significantly more common among those who stopped CTX, consistent with prior studies ,6,10,11.3 in both arms and had been on ART for a median of over 4 y with few prior hospitalizations in the past 3 mo. This, coupled with high retention, suggests that these patients may represent an elite subgroup of patients who took advantage of a strong PSC.Our study had several strengths and limitations. Because our study was an unblinded clinical trial without a placebo, clinician’s care decisions, as well as the patients’ threshold to come in for care, may have been influenced by trial arm. A second limitation, for statistical analyses, was an observed incidence of morbidity events that was lower than expected. Additionally, our study was limited to 12 mo of follow-up. A strength of the study was high retention (98% in each arm). The PSC at the Homa Bay District Hospital had a long history of collaboration with Médecins Sans Frontières and a reputation for excellent care, which contributed to retention in the trial. Participants had median CD4 counts approaching 600 cells/mmIn conclusion, CTX discontinuation among ART-treated adults in a region with endemic malaria results in increased incidence of clinical malaria but not pneumonia or diarrhea. The implications are broad, and our results suggest that CTX prophylaxis should continue in regions with endemic malaria. However, in regions without malaria or with less endemicity, CTX cessation may be possible after immune recovery.S1 Table(DOCX)Click here for additional data file.S1 Text(PDF)Click here for additional data file.S2 Text(DOC)Click here for additional data file.S3 Text(DOCX)Click here for additional data file.


Increased HIV testing frequency among high-risk populations such as men who have sex with men (MSM) and male-to-female transgender women (TW) can lead to earlier treatment and potentially reduce HIV transmission.We analyzed baseline survey data from 718 high-risk, young (median age 29 [interquartile range 23–35]) MSM/TW enrolled in a community-based HIV prevention trial between 2008–2009. Participants were recruited from 24 neighborhoods in and around Lima, Peru. We assessed HIV testing frequency, testing behaviour, and motivations and barriers to testing. Multivariate analysis identified correlates to prior HIV testing.Overall, 79.6% reported HIV testing within their lifetimes, however, only 6.2% reported an average of two tests per year, as per Peruvian Ministry of Health guidelines. The most commonly reported motivators for testing were to check one’s health (23.3%), lack of condom use (19.7%), and availability of free testing (14.0%), while low self-perceived risk for HIV (46.9%), fear of a positive result (42.0%), and lack of access to testing services (35.7%) were the most frequently reported barriers. In multivariate analysis, factors independently associated with HIV testing included age , transgender-identification vs. gay-identification , history of transactional sex , and prior sexually transmitted infection diagnosis .An overwhelming majority of participants did not meet the standard-of-care for testing frequency. The reported motivations and barriers to testing highlight issues of risk perception and accessibility. Our findings suggest utilizing non-traditional outreach methods and promoting HIV testing as a routine part of healthcare in Peru to encourage testing and knowledge of HIV serostatus. The complex issue of HIV prevention requires a multidimensional approach that includes regular voluntary HIV testing and counseling (HTC), especially among key populations such as men who have sex with men (MSM) and male-to-female transgender women (TW) ,2. KnowlPsychosocial and operational barriers -8 prevenThe current standard of care recommended by the Peruvian Ministry of Health (MoH) for MSM/TW is HTC every six months , howeverComunidades Positivas trial [We conducted a cross-sectional analysis of baseline survey data from the as trial ,18. ThisTraditional ethnographic methods were used to identify 24 low-income neighborhoods that had a strong network of MSM/TW. Potential participants were recruited using a snowball technique. Inclusion criteria included biologically born males aged 18 to 45 years who reported at least one sexual encounter with a man or transgender woman in the past 12 months, lived or worked near one of the 24 identified neighborhoods, planned to stay in the area for the duration of the study and willing to participate in the study. Baseline survey data was collected between 2008–2009. Eligible participants provided written informed consent prior to the initiation of any study procedures. The study was approved by the Institutional Review Boards of University of California, Los Angeles and Universidad Peruana Cayetano Heredia.The baseline survey interview was conducted face-to-face by a native Spanish speaker for all sections except for the HIV testing history, which was administered by audio computer-assisted self-interviewing (ACASI) due to fears of reporting bias among HIV positive participants given stigmatization of people living with HIV. Variables assessed included socio-demographic characteristics, HIV testing history , a sexual risk assessment , and a problem drinking assessment using the CAGE instrument .All participants underwent pre-test counseling for HIV and syphilis after which rectal and pharyngeal swab samples and 10 ml of blood were collected for testing. Test results and post-test counseling were provided within two weeks of sample collection. Participants with syphilis were given appropriate antibiotic therapy. Participants with newly diagnosed HIV infections were referred to designated MoH facilities for ongoing care and treatment. HIV antibody status was determined using Genetic Systems HIV-1/HIV-2 Plus O EIA  and positive results were confirmed with GenScreen HIV-1/HIV-2 Western Blot . Syphilis infection was determined by Rapid Plasma Reagin , followed by TPPA confirmation  and serial dilution of RPR titers. For the purposes of this analysis, diagnosis of syphilis infection was based on an RPR titer of 1:8 or higher. For quality control, 10% of HIV samples were sent to the San Francisco Department of Public Health laboratory , and 10% of syphilis samples were sent to the US Navy Medical Research Unit-6 Bacteriology laboratory in Lima, Peru for confirmatory testing.The main outcome of “HIV testing” was based on participant self-report and defined as a dichotomous variable according to lifetime testing history (ever-testers and never-testers). “HIV testing” was also assessed as a continuous variable (according to the number of reported lifetime HIV tests) in order to determine testing frequency. In the descriptive analysis, median values and interquartile ranges (IQR) were estimated for continuous variables and proportions calculated for categorical variables. “Ever-testers” were defined as participants who reported undergoing HIV testing prior to the baseline survey and “never-testers” were defined as those who reported no prior history of HIV testing. Participants were defined as newly HIV-positive if they reported HIV-negative status but had a positive Western blot test at baseline. Participants who accurately reported their HIV-positive status at baseline were considered to have a known HIV-infection, and these individuals were excluded from further analysis.Survey questions on testing motivations and barriers permitted multiple answers. The possible answer choices were provided from a previously defined list of possible responses with a write-in answer space available. The write-in answers that fell underneath the scope of the pre-defined list of answers were included in the analysis. Write-in answers that fell outside of the scope of pre-defined answers were minimal and not included in the analysis. The mean number of tests per sexually active year was calculated by dividing the number of prior HIV tests by the number of sexually active years since age 18. Self-reported sexual behaviours with the previous three partners were pooled to identify whether individuals had engaged in unprotected anal intercourse (UAI) within the past six months. Problem drinking was defined as answering “yes” to two or more of the four CAGE questions .We used chi-square and Wilcoxon rank sum tests to evaluate associations between the outcome and categorical and continuous variables respectively. To measure the strength of association between the main outcome and independent variables, prevalence ratios (PR) with 95% confidence intervals (CI) were calculated for bivariate and multivariate analyses. We used Poisson-family generalized linear models with a logarithmic link function and robust variance to estimate PR for correlates of ever HIV testing ,21. VariComplete clinical and demographic characteristics of the study sample are shown in Table The sample consisted of 718 participants with a median age of 29 (IQR 23–35) years and a median number of sexually-active years of 14 (IQR 9–21). Participants identified most commonly as gay (64.8%) or transgender (29.0%), and most had not obtained post-secondary education (71.6%). Many participants reported high-risk behaviours such as transactional sex, , UAI within the past six months (61.7%), or problem drinking, as denoted by a positive CAGE screen (57.8%).Of the 718 participants, the majority (79.6%) reported at least one HIV test within their lifetime  were newly diagnosed with HIV of whom 30 (36.6%) tested within the past year, 31 (37.8%) tested over a year ago, and 18 (22.0%) had never been tested. Three participants with newly diagnosed HIV infection (3.66%) declined to answer whether they had ever been tested.Although the HIV prevalence between ever-testers and never-testers was similar , ever-testers were significantly more likely to engage in transactional sex, report problem drinking, have a positive RPR, and report greater number of sexual partners in the past six months than never-testers , as the questions did not differentiate between primary and secondary factors. Nonetheless, the majority of write-in categories were reiterations of the potential responses provided, which is consistent with prior studies conducted within and outside of Peru ,6,8.A final note is warranted with respect to our study population. In Peru, the social networks of MSM and TW tend to overlap, and it is a frequent challenge of studies to characterize the distinct attributes of each group. Moreover, at the time of study design, these group differences were not as well recognized as they are today, and the development of improved study methodologies continues to be necessary for an improved understanding of sexual dynamics among Peruvian MSM/TW. Nonetheless, our sample population involved high-risk, low-income MSM/TW, and while not representative of all MSM/TW in Lima, Peru, it highly corresponded with those targeted by MoH HIV testing programs.Our study calls for alternative outreach methods and public education to improve adherence to the MoH recommended standard of care for HIV testing among MSM/TW through a norm of client-initiated HTC. The use of mobile testing and availability of HIV testing in MSM/TW frequented venues would capture MSM/TW not reached by traditional healthcare approaches, which places responsibility on patient-initiated visits to an STI clinic ,25. In a


CD36 has been associated with obesity and diabetes in human liver diseases, however, its role in age-associated nonalcoholic fatty liver disease (NAFLD) is unknown. Therefore, liver biopsies were collected from individuals with histologically normal livers (n=30), and from patients diagnosed with simple steatosis . Patients were divided into two groups according to age and liver biopsy samples were immunostained for CD36. NAFLD parameters were examined in young (12-week) and middle-aged (52-week) C57BL/6J mice, some fed with chow-diet and some fed with low-fat  or high-fat diet  for 12-weeks. CD36 expression was positively associated with age in individuals with normal livers but not in NAS patients. However, CD36 was predominantly located at the plasma membrane of hepatocytes in aged NAS patients as compared to young. In chow-fed mice, aging, despite an increase in hepatic CD36 expression, was not associated with the development of NAFLD. However, middle-aged mice did exhibit the development of HFD-induced NAFLD, mediated by an increase of CD36 on the membrane. Enhanced CD36-mediated hepatic fat uptake may contribute to an accelerated progression of NAFLD in mice and humans. Therapies to prevent the increase in CD36 expression and/or CD36 from anchoring at the membrane may prevent the development of NAFLD. The world is experiencing a dramatic increase in numbers of elderly people, rising to an estimated 22% of the world population aged 60 years and older by 2050 . AlthougNonalcoholic fatty liver disease (NAFLD) is, worldwide, the most common liver disease, affecting one-third of the overall population . NAFLD iCD36 facilitates the uptake of long-chain fatty acids (LCFA) across the cell membrane and high levels of expression can contribute to lipid accumulation in heart and skeletal muscle . EnhanceWe have, therefore, sought to investigate whether increased CD36 expression underlies the increased susceptibility to the development of NAFLD with age. Our data show that aging is associated with a dramatic increase in CD36 expression in human NAFLD as well as in aged mice on a normal diet. Our data also show that aging, in combination with HF-feeding, triggers the presence of CD36 at the cell surface of hepatocytes, which may contribute to enhanced fat uptake in NAFLD and drive the progression of simple steatosis towards NASH. Therefore, our data suggest that therapies to prevent the increase in CD36 expression and CD36 from anchoring at the membrane may prevent the development of NAFLD.To determine whether CD36 expression in the human liver is increased by the physiological process of aging, immunohistochemical staining for CD36 was performed in liver biopsy samples from young (20-38 years) and aged individuals (50-83 years) with histologically normal livers, and liver biopsy samples from young (20-42 years) and aged individuals (52-74 years) diagnosed with non-alcoholic steatosis (NAS) Fig. . We obseTo further investigate the correlation between aging and hepatic CD36 expression, we assessed CD36 expression in middle-aged mice fed a chow-diet for one year. Body weight was significantly increased Fig.  in middlIl1β, Tnf, IkBα, Cd68, Mcp1, Cxcl1) in middle-aged mice as compared to young mice  was not affected by aging in these mice , suggesting that fat oxidation is impaired in middle-aged mice irrespective of HF-feeding. Although we did not perform direct measurements of fatty acid oxidation, this is in line with a previous study showing that mitochondria of middle-aged mice display many electron transport chain defects and decreased complex I and IV activities . Our resFurthermore, our results indicate that minor lipid droplet formation is indeed present in middle-aged mice fed a regular chow diet for one year Fig. . Howeveret al. [Although our findings seem to contradict the current understanding that aging is associated with hepatic steatosis in humans and rodents, they are in line with the recent study of Fontana , who shoet al. . This coper sedoes not contribute to the development of NAFLD; however, it does increase susceptibility to the development of diet-induced NAFLD. Furthermore, we report for the first time that aging increases CD36 membrane expression in the livers of both mice and humans. Therefore we propose that increased CD36 localization/stabilization at the plasma membrane may be a key to enhanced hepatic fat uptake, and may play an important role in advancing NAFLD with age. Our results suggest that drugs that prevent the increase in CD36 expression and/or CD36 from anchoring at the plasma membrane may alleviate NAFLD and prevent the transition from benign steatosis toward more advanced NASH.In summary, we have shown that aging et al [This study comprised 56 non-diabetic patients who underwent a needle liver biopsy during programmed laparoscopic cholecystectomy. Liver biopsy sections were collected as previously described  and histet al  was usedet al , NASH paImmunostaining and computational image analysis using analySIS® software  were performed on formalin-fixed paraffin-embedded liver biopsy sections (4 m thickness) from all patients of each group, as previously described . All immAfter a 12 h overnight fast, clinical and anthropometric data as well as venous blood samples were obtained from each patient at the time of liver biopsy. All serum chemistry analyses were performed at the Central Laboratory Service of the University Hospital Santa Cristina  as described . InsulinThis study was performed with the approval of the University of Alberta Animal Policy and Welfare Committee and the University of Groningen Ethics Committee for Animal Experiments, which adheres to the principles and guidelines established by the European Convention for the Protection of Laboratory Animals. Experiments were carried out on young (12-14 week old) and middle-aged (52-58 week old) male C57BL6/J mice, maintained on a 12-hour light-dark cycle and given free access to water and chow diet unless otherwise stated. At 12-14 or 32-34 weeks of age, a subset of mice was randomly divided into a low-fat diet (LFD) group  and a high-fat diet (HFD) group  for a period of 12 weeks. Insulin was measured in plasma from fasted mice as described .Paraffin-embedded sections of the liver (4μm) were stained with hematoxylin-eosin (H&E). Microscopy was performed with a Leica DM 3000 microscope with a DFC420 camera .Ppia). Primers are presented in Total RNA was isolated from the liver using QIAazol reagent . First-strand cDNA synthesis was performed using the QuantiTect Reverse Transcription Kit . qRT-PCR was performed using a 7900HT System  and values were corrected using the housekeeping gene Cyclophillin A  and immunoblotted with antibodies against, Erk1/2, p-Erk1/2, Actin, α-tubulin, Parkin , Pan Cadherin , CD36  and Oxphos . Immuno-positive bands were visualized by chemiluminescence  and quantified using the Molecular Imager ChemiDoc xrs+system from Bio-Rad .2 test or Fisher exact test for categorical variables and the unpaired t test or Mann-Whitney U test for continuous variables. Spearman's rho test was used to evaluate correlations. Experimental data were compared using the unpaired t test or Mann-Whitney U test. The level of significance was set at P<0.05. All statistical analyses were performed in GraphPad Prism .Data are presented as mean ± SEM unless stated otherwise. The characteristics of the patients studied were compared by the Pearson


Autism spectrum disorder (ASD) is more prevalent in males, and the mechanisms behind this sex-differential risk are not fully understood. Two competing, but not mutually exclusive, hypotheses are that ASD risk genes are sex-differentially regulated, or alternatively, that they interact with characteristic sexually dimorphic pathways. Here we characterized sexually dimorphic gene expression in multiple data sets from neurotypical adult and prenatal human neocortical tissue, and evaluated ASD risk genes for evidence of sex-biased expression. We find no evidence for systematic sex-differential expression of ASD risk genes. Instead, we observe that genes expressed at higher levels in males are significantly enriched for genes upregulated in post-mortem autistic brain, including astrocyte and microglia markers. This suggests that it is not sex-differential regulation of ASD risk genes, but rather naturally occurring sexually dimorphic processes, potentially including neuron–glial interactions, that modulate the impact of risk variants and contribute to the sex-skewed prevalence of ASD. Autism spectrum disorder is approximately 4.5 times more likely to occur in boys than girls. Here, Werling, Geschwind and Parikshak characterized sexually dimorphic gene expression in the non-diseased, post-mortem, adult and prenatal human brain, and show genes expressed at higher levels in males are significantly enriched for genes upregulated in autistic brain. FMR1) or Smith–Lemli–Opitz syndrome (DHCR7)88Autism spectrum disorder (ASD) is a developmental condition characterized by deficits in social communication and restricted, repetitive behaviours or interests34rome DHCR789. The One robust risk factor for ASD is sex: for every female with ASD in the United States, there are 4.5 affected males121379Several theories have been proposed regarding sex-differential risk and protective mechanisms, though the evidence supporting each is varied167918We reasoned that since sex-differential gene expression patterns contribute to the development and function of a sexually dimorphic brain, evaluating genome-wide sex-differential gene expression in neural tissue could elucidate points of overlap with ASD risk genes and related pathways. Here we test two basic hypotheses about the relationship between sexually dimorphic expression and ASD risk genes: (1) ASD risk genes are expressed at different levels in males and females. With differing baseline expression levels, the magnitude of the impact of a disruptive mutation in a risk gene is likely to differ by sex. Accordingly, we expect to observe enrichment of sex-differentially expressed (sex-DE) genes among known ASD risk genes. (2) ASD risk genes are expressed at the same level in males and females, but genes in interacting molecular pathways and/or cellular processes are differentially expressed by sex. In this case, the downstream impact of sex-neutrally expressed ASD risk genes is likely modulated by their interactions with sexually dimorphic processes. If this were the case, we would expect to observe enrichment of sex-DE genes among gene sets representing processes associated with ASD pathophysiology, but not among ASD risk genes themselves.To evaluate these hypotheses, we identify sexually dimorphic gene expression patterns in adult and prenatal human cerebral cortex tissue and test ASD-associated gene sets for enrichment of sex-differential expression. We observe significantly male-biased expression patterns for microglial-function-enriched, ASD-upregulated, co-expression modules from post-mortem ASD brain232526To identify sex-differential gene expression patterns in the human brain, we analysed RNA-sequencing (RNA-seq) gene expression data from human cerebral cortex tissue from the BrainSpan project3023823242526273233We first analysed RNA-seq data from 58 post-mortem cortex samples from the BrainSpan projectXIST, an X-chromosome transcript that initiates X-chromosome inactivation and is only expressed in females , or higher expression in females . We found no evidence from either test for significant sex-differential expression of any of these diverse sets of ASD risk genes in the adult cerebral cortex, providing no evidence supporting hypothesis 1  candidate genes from a manually curated databasethesis 1 .V, downregulated in ASD and enriched for genes with neuronal and synaptic functions; and asdM16V, upregulated in ASD and enriched for genes involved in immune and inflammatory responses)et al.G, downregulated in ASD, and asdM6G, upregulated in ASD, both modules are enriched for neuronal markers and synaptic genes and overlap with the asdM12V moduleG, which is upregulated in ASD, enriched for M2-state microglial genes, and overlaps with the asdM16V moduleNext, we tested gene sets with evidence of ASD-associated expression patterns in adult human cortex232526P<0.05) in males than in females , we find a sevenfold enrichment of ASD-upregulated genes =1.5e-04; P (PBinom_adj)=6.9e-03, V also show enrichment for significantly male-DE genes  that were also tested in the ASD-DE analysis, we observe a highly significant positive correlation between the sex-differential and ASD-differential FDs . We note that among the 47 genes expressed at significantly higher levels in both males and in ASD, 22 are asdM16V membersV membersV and asdM12V, and that this pattern may be more broadly indicative of sexually dimorphic transcriptomes originating from specific neural cell types.To assess the extent of these parallel expression patterns across the transcriptome, we compared the FDs from sex-DE genes with FDs observed in the differential expression analysis of ASD cortexP<0.005, 98 genes at P<0.01 and 519 genes at P<0.05 s, r=0.57, P=3.6e-65; P<0.05) from these data  and 70 are astrocyte markers , further supporting the characterization of astrocytic genes and ASD-upregulated, microglial-function-enriched asdM16V genes, as displaying male-biased expression in the adult human cortex  in the more susceptible male population, and higher expression of a cohort of synaptic genes (exemplified by asdM12V), in the more protected, female population.Taken together, these findings fail to support the hypothesis that ASD risk genes are differentially expressed by sex in the human cerebral cortex. Rather, we observe that sex-DE genes are enriched among specific classes of changes observed in post-mortem cortex from ASD patients, which is indicative of shared biology between ASD and typical sexual dimorphisms. These patterns of sex-DE enrichment follow what is observed in ASD cortex, with higher expression of microglial and astrocyte genes  a greater proportion of astrocytes and microglia to neurons in male cortex results in higher measured gene expression in males; or (2) males' cortical astrocytes and microglia are more transcriptionally active than females'. Aside from early, tenuous observations of greater cortical neuron density in males45464850525354795758We also acknowledge several limitations of our study. First, our analyses utilize gene expression data from the human cerebral cortex, a region that other studies have indicated is a convergent area where many ASD risk genes likely exert their action30The mechanism(s) of the sex-biased expression observed here are not known, but present significant opportunities for understanding ASD's sex-biased pathogenesis. For example, transcription of sex-DE genes identified here may be regulated by sex-differential factors, such as transcription factors from the sex chromosomes, or by sex steroid receptor binding to hormone response elements upstream from transcription start sites. Chromatin immunoprecipitation experiments in human neural tissue coupled with functional assays in model systems will be needed to thoroughly characterize binding sites for these regulatory elements throughout the genome.Alternatively, the sex differences we have observed may not result from differential transcription regulation, but instead from sex differences in the proportions of different neural cell types within male and female tissue. The overlap between male-DE genes and astrocyte and microglial marker genes is consistent with this possibility, although the processes that lead to sex differences in cell type proliferation or differentiation are unclear. Quantitative histological studies of male and female human brain will be critical to characterize the extent to which cell type composition differs between the sexes, and neurodevelopmental follow-up will be required to demonstrate how these differences arise.V, enrichment of genes carrying ASD risk variants provides evidence for a causal, primary role in ASD aetiology. However, ASD-associated co-expression modules that lack enrichment for genetic risk variants, such as asdM16V, may be linked to ASD in one of several ways: (1) They reflect direct downstream consequences of variants in, or dysregulation of, primary, ‘causal' genes  that actively participate in the pathophysiological processes that shape and maintain an autistic-functioning brain; (2) they represent risk pathways that are independent of primary risk genes, for example, consequences of an environmental exposure, that also actively participate in ASD's aetiology; or (3) they exemplify consequences of living with ASD to the brain, which may modify the presentation of ASD or its comorbidities, but that do not have a role in ASD's aetiology. For example, the gene expression signature captured by non-variant-enriched, co-expression module asdM16V, which is upregulated in post-mortem adult autistic cortex, may reflect the impact of chronic social stress experienced across the lifespan. If male and female brains respond differently to stress23The extent to which the convergent transcriptional changes previously observed in ASD cortex23In any case, the sex-DE patterns that we observe in multiple, independent samples demonstrate that currently identified ASD risk genes are not innately sex-differentially expressed in the developing or adult human cerebral cortex. Instead, it is the molecular, cellular or circuit-level context in which ASD risk genes operate that is likely responsible for modulating ASD liability in a sex-differential manner. This notion fits with the remarkable consistency of the male bias in ASD prevalence, despite ongoing evidence of considerable genetic heterogeneity in ASD. Study of astrocyte–neuronal and microglia–neuronal synapse interactions, and study of the causes and functional consequences of higher expression of astrocyte and microglial genes in male brain and synaptic genes in female brain, are needed to determine how sex-differential functioning of these pathways impact sexually dimorphic risk for a variety of neurodevelopmental disorders, including ASD.2-transformed (log2(RPKM+1)). Samples from regions of the frontal, temporal and parietal cortex from subjects aged 13–40 years, with RNA integrity number (RIN) of at least 8.0, were retained for this analysis. Non-expressed genes defined as having a log2-transformed RPKM expression level of <1 in more than 50% of all male or female samples were removed. Outlier samples were identified by evaluating inter-sample correlations and hierarchical clustering, first within each sex, and then on the full data set (samples of both sexes considered together); samples with inter-sample correlations >2.5 s.d.'s from the mean were removed. The data were then quantile normalized to mitigate the effects of systematic differences in the distribution of expression levels across samples on the differential expression analysis results. After gene filtering and outlier removal, 72 samples  and 16,843 expressed genes remained. To balance the male and female sample sets, we then matched a subset of the female samples to the male samples by age and brain region, and again filtered for expressed genes. The final data set consisted of 58 samples from 10 subjects  and brain region. These samples were a subset of a larger set acquired from the Harvard Brain and Tissue Bank and the Eunice Kennedy Shriver National Institute for Child Health and Human Development Brain and Tissue Bank for Developmental Disorders following the tissue acquisition policies of the respective brain banks.Approximately 100 mg of tissue from each sample was dissected on dry ice; RNA isolations were performed using the miRNeasy kit (Qiagen), RNA quality was quantified using the RIN2 values, and ensured that there were no sample outliers with a summed sample correlation Z-score>2  and removed. Outlier samples with inter-sample correlations >2.5 s.d.'s from the mean, either within each sex or within the complete sample set, were removed. The expression data for the remaining samples were then quantile normalized. The final filtered data set consisted of 13 samples from 10 subjects (Reads were mapped to the human reference genome (hg19) using Gencode v18 annotations with TopHat2, allowing for up to 10 multiple mappings per read41e>2 ref. . Genes nsubjects , and 19,subjects .et al.Exon array data analysed by Kang 2-transformed median probe set intensity of <6 in >80% of male or female samples within the selected sample set, and were removed. Outlier samples were detected by evaluating inter-sample correlations and hierarchical clustering within each sex and within the complete data set, and samples with inter-sample correlation >2.5 s.d.'s from the mean were removed. Expression data for the remaining samples were quantile normalized. After these processing steps, 142 samples  and 9,985 expressed genes remained. To balance the number of male and female samples for analysis, we then matched a subset of the female samples to the male samples on age and brain region, and again filtered for expressed genes. The final data set consisted of 86 samples from 8 subjects  in sporadic ASD cases from exome sequencing of families from the Simons Simplex CollectionASD candidate genes were selected from the SFARI Gene Autism Databaseet al.q-value<0.1, 88 genes) or (2) lower levels in ASD than in control cortex (125 genes); (3) ASD-associated, unsigned co-expression modules from weighted gene co-expression network analysis, asdM12V ; and (4) asdM16V . From a 2014 study by Gupta and colleagueset al.G , (6) asdM6G  and (7) asdM5G   .Since gene expression patterns in the brain often reflect the transcriptional activity of distinct neural cell types39Recent evidence has also suggested complementary roles for microglia and immune cells in mediating behaviour in male versus female animalset al.et al.27, we used the genome-wide set of one-to-one human–mouse orthologues as background  at each of the three significance levels for the sex-DE input genes.To evaluate the enrichment of gene sets of interest among sex-DE genes, we applied a two-sided Fisher's exact test separately to sex-DE gene sets expressed more highly in males  and in females  at three 2(fold difference), at any P value); or higher expression in females (negative log2(fold difference), at any P value). We then applied a two-sided binomial test against the distribution of male-higher and female-higher genes in the corresponding background gene set . Computer code used in this study is available on request.How to cite this article: Werling, D. M. et al. Gene expression in human brain implicates sexually dimorphic pathways in autism spectrum disorders. Nat. Commun. 7:10717 doi: 10.1038/ncomms10717 (2016).Supplementary Figures 1-2 and Supplementary Table 1Sex-differential expression results from all three data sets: adult BrainSpan RNA-seq, adult replication set RNA-seq, prenatal exon array.Tables for all ASD-risk and ASD-associated gene sets tested for enrichment of sex-differential expression.Over-representation analysis results for adult BrainSpan RNA-seq data set.Tables for all neural cell type marker gene sets tested for enrichment of sex-differential expression.Tables for all adaptive immune system cell type marker gene sets tested for enrichment of sex-differential expression.Over-representation analysis results for adult replication RNA-seq data set.Over-representation analysis results for prenatal exon array data set.Tables for all gene sets used as background in over-representation analyses.Membership in gene sets of interest, background gene sets, and sex-differentially expressed gene sets for all expressed genes from each of the three data sets analyzed.


Background. Human papillomavirus (HPV) vaccines are recommended for girls prior to sexual debut because they are most effective if administered before girls acquire HPV. Little research has been done on HPV prevalence in girls who report not having passed sexual debut in high HPV-prevalence countries.Methods. Using attendance registers of randomly selected primary schools in the Mwanza region of Tanzania, we enrolled girls aged 15–16 years who reported not having passed sexual debut. A face-to-face interview on sexual behavior and intravaginal practices, and a nurse-assisted self-administered vaginal swab were performed. Swabs were tested for 13 high-risk and 24 low-risk HPV genotypes.Results. HPV was detected in 40/474  girls. Ten different high-risk and 21 different low-risk genotypes were detected. High-risk genotypes were detected in 5.3% . In multivariable analysis, only intravaginal cleansing (practiced by 20.9%) was associated with HPV detection .Conclusion. This cohort of adolescent Tanzanian girls had a high HPV prevalence prior to self-reported sexual debut, and this was associated with intravaginal cleansing. This most likely reflects underreporting of sexual activity, and it is possible that intravaginal cleansing is a marker for unreported sexual debut or nonpenetrative sexual behaviors. Cervical cancer is the most common form of cancer in women in sub–Saharan Africa and the highest age-standardized incidence of cervical cancer in the world is found in East Africa at over 30.0 per 100 000 person-years . AlthougThe 2 available HPV vaccines, Gardasil (MSD) and Cervarix , cover the 2 most common cancer-causing HPV genotypes HPV-16 and -18. Additionally, Gardasil covers HPV-6 and -11, which, though low-risk for cervical cancer, are common causes of genital warts. HPV vaccination is most effective if administered prior to acquiring infection with these vaccine-related genotypes , 9. HoweTo address the gaps in knowledge, we present baseline cross-sectional results from a cohort study in Tanzania that enrolled adolescent girls who reported that they had not passed sexual debut, and who were tested for vaginal HPV DNA. This study is the first to describe HPV prevalence in girls in sub–Saharan Africa who report no previous sexual intercourse, and the associations between reported sexual behaviors, vaginal practices, and HPV infection.The cohort was enrolled between January and August 2012 from previously prepared attendance lists of randomly selected government primary schools in 3 districts of the Mwanza Region in Tanzania. These lists, covering all primary schools in the districts, had been drawn up in 2010 to prepare for a trial of delivery methods for HPV vaccine . The lisEligibility criteria included being aged 15 or 16 years at enrolment, enrolled in class 6 in 2010 in 1 of the selected schools, not pregnant (self-reported), planning to stay in the study area or able to travel to appointments, self-reporting never having had vaginal sex, and being willing to self-administer vaginal swabs. A subsample of 26 of the 82 schools were randomly selected and, in order to prevent stigmatization of girls who reported sex, at each of these 26 schools the first girl who reported ever having had sex was enrolled. An additional eligibility requirement for these girls was having passed sexual debut within the past 12 months.The London School of Hygiene and Tropical Medicine Ethics Committee and the Medical Research Coordinating Committee, Tanzania, approved the study protocol. Because all potential participants were considered minors in Tanzania (under 18 years old), written informed consent was required from a parent/guardian with subsequent participant informed assent. Consent for enrolment was taken before any assessment of previous sexual debut. The consent procedure involved separate face-to-face explanations of the study to parents and daughters by a study nurse, provision of written information, and time to ask questions. Individuals unable to read were consented in the presence of an independent witness who provided an additional signature. Individuals unable to write provided a thumbprint. Parents/guardians were compensated for their time with Tsh5,000 (approximately $3), and participants were provided with a toothbrush and toothpaste.After enrolment, girls had a face-to-face interview with a female study nurse. Interviews were carried out in private at the participant's home, school, or local health center, depending on participant preference. Interviews were carried out in Swahili using a structured paper questionnaire, which had been translated and back-translated from English. Questions covered demographic and socioeconomic details, menstrual and vaginal hygiene practices, nonpenetrative sexual behaviors, and details of previous penetrative-sex frequency and partners. Interviews included colloquial terms for sexual behaviors that had been collected during focus group discussions with similar-aged girls. One self-administered vaginal Dacron swab was obtained after instructions from a study nurse, who remained in the room and provided verbal and positional hand guidance if necessary. Girls are being followed every 3 months for 18 months. We report results from the enrolment visit.Immediately after collection, swabs were stored dry in cryotubes and placed into cold boxes with ice packs. They were submitted daily to the reference laboratory in Mwanza and stored at −20°C until they were shipped to the Catalan Institute of Oncology, Barcelona, Spain. HPV detection and genotyping were performed using the Linear Array HPV genotyping assay  which detects 37 HPV genotypes . For this study, 13 HPV genotypes, HPV-16, -18, -31, -33, -35, -39, -45, -51, -52, -56, -58, -59, and -68, were classified high-risk [Briefly, DNA was extracted from the specimens by silica-gel-based methods . Extracted material was amplified with the PGMY PCR system, and the generated amplicons were detected and typed by reverse-line blot reaction . Polymerase chain reaction in this assay is based on a multiplex system, including human β-globin amplification primers. This provides an internal quality control for sample material. Specimens consistently negative for β-globin amplification were excluded because it was assumed that vaginal sampling was unsuccessful. All protocols were performed according to manufacturer's instructions: each step was performed in separate rooms and negative controls were used.Questionnaire data were double-entered into OpenClinica LLC , and analyzed using STATA V12.0 . Baseline cohort characteristics were examined, and the prevalence of any HPV infection and prevalence of individual HPV genotypes were calculated. Household wealth was estimated using a score based on the number of possessions owned by the head of the household in which the participant currently resided. Median age of menarche in the cohort was calculated using Kaplan–Meier survival methods to account for those who had not reached menarche.P < .1 after adjustment for age were included in a multivariable model. Vaginal cleansing was investigated as a binary term for ever having cleansed, and as frequency of cleansing categorized as never, ≤2, and ≥3 times per day. All P values are from likelihood ratio tests.Factors associated with prevalent HPV infection (treated as a binary outcome for any HPV genotype vs none) were identified using logistic regression models. Age was considered an a priori potential confounder, and factors reaching Of 1555 potentially eligible girls on the original school attendance lists, 1177 (76.5%) were located. From these, 801 (68.1%) were confirmed to be the appropriate age, and 628 (78.4%) consented to be screened for enrollment  and the remainder (52.5%) were 16 years old . In total, 24  girls had a high-risk HPV genotype. Multiple genotype infections were detected in 20  girls, comprising 50.0% of those with HPV DNA. The 2 most commonly detected genotypes were HPV-42 and HPV-58, with a prevalence of 1.9% (N = 9) and 1.1% (N = 5), respectively , and high-risk HPV was detected in 22.7% . Though the numbers involved were small, their demographic profile was similar to that of girls who reported no previous sex (data not shown).A total of 22 eligible girls who reported having passed sexual debut within the last year were enrolled from 26 randomly selected schools. HPV DNA was detected in 7  of these 22 girls, significantly higher than in those who reported never having had sex (P = .10). Only the reporting of ever having practiced intravaginal cleansing was associated with HPV DNA detection in univariable analysis, and this persisted after adjustment for age  (Table P = .002). However, among girls who reported cleansing, there was no evidence of an association of HPV detection with method (fingers vs cloth), or substance used   based on the testing of self-administered vaginal swabs. This is higher than in 2 previous studies among women who reported no previous sex: 1 study of 130 Swedish women aged 10–25 years reported a prevalence of 1.5%, and 1 longitudinal study of American women aged 18–20 years reported that 1.7% of specimens were found to have HPV DNA . A furthPrevious studies have confirmed vaginal sex as the predominant method for acquisition of cervical HPV infection in women, as demonstrated by rapid HPV acquisition after reported sexual debut, and a significant increase in the risk of HPV infection with increased number of partners and with high-risk partners , 15. LacAlternative explanations for the presence of vaginal HPV DNA in girls and women who report no previous sex include mother-to-child transmission (MTCT) of HPV, nonpenetrative sex practices, or transmission via fomites . MTCT isIn the age-adjusted analysis in our study, disclosure of ever having practiced intravaginal cleansing was associated with more than doubled odds of prevalent HPV. Furthermore, a positive dose-response relationship was seen with increasing frequency of vaginal cleansing. Commonly, women use intravaginal practices (IVP) to manage menstruation, as part of their sexual practice , and to improve genital hygiene. IVP includes both cleansing inside the vagina with water, soap, or other products, and the insertion of products into the vagina  . IVP is Trichomonas vaginalis in Zambian girls aged 13–16 who reported no previous sexual contact [HPV has been detected in fingernail and fingertip specimens of young sexually active women in the United States, as well as on toilet seats in several European airports, and on surfaces in a sexual health clinic in the United Kingdom , 33. HPV contact . Intrava contact .An important and likely explanation for the observed association between intravaginal cleansing and HPV is that intravaginal cleansing may be a marker of unreported sexual debut or nonpenetrative sex behaviors. Girls may be more likely to start vaginal cleansing after sexual debut because they believe that this may prevent pregnancy or STIs, relieve STI-related symptoms, or because they have been taught that this practice is appropriate for sexually active women . IncreasMultiple HPV-genotype infections were as common as single genotype infections in this study, consistent with observations in adolescent girls in previous studies in the United States , 39. MorMajor strengths of this study include the interview method, in which the questionnaire included local age-specific colloquial terms for sexual practices, and was administered by trained nurses experienced in adolescent sexual behavior research. In addition, vaginal swabs were self-administered but collection was directly observed by nurses. Self-administered vaginal swabs in a previous local study (median age 17) demonstrated collection of epithelial cells in 233/244 (95%) of swabs , and 99%One of the limitations of this study is that the enrollment list came from primary schools. Although primary school attendance is a legal requirement in Tanzania, and has been reported as 97% , it may The proposed HPV vaccination program in Tanzania will target girls in primary school class 4, where the median age is 10 years, and a catch-up campaign in older girls has not thus far been proposed. This study provides useful evidence for policymakers in relation to the likely effectiveness of such a campaign. Overall, 80.3% of 15–16 year old girls reported no previous sex, 99.2% of whom did not have infection with either of the 2 high-risk HPV genotypes included in current HPV vaccines (HPV-16/18). The WHO recommend including older girls in catch-up campaigns if a significant proportion of girls are naive to HPV vaccine types [The data we present strongly link prevalent HPV infection with reported intravaginal cleansing. Intravaginal cleansing may be a marker for undisclosed sexual activity in our population, but could alternatively be a novel mechanism for the nonsexual transmission of HPV. If this is the case, the identification of this potentially modifiable risk factor would be highly relevant for a country with one of the highest rates of cervical cancer in the world. Further research to confirm and understand the link between intravaginal cleansing and HPV is required and is currently underway in Mwanza.


Hospitalisation for adversity-related injury  has been described as a “teachable moment”, when intervention may reduce risks of further harm. Which adolescents are likely to benefit most from intervention strongly depends on their long-term risks of harm. We compared 10-y risks of mortality and re-admission after adversity-related injury with risks after accident-related injury.n = 333,009) or for accident-related injury . We used Kaplan–Meier estimates and Cox regression to estimate and compare 10-y post-discharge risks of death and emergency re-admission. Among adolescents discharged after adversity-related injury, one in 137 girls and one in 64 boys died within 10 y, and 54.2% of girls and 40.5% of boys had an emergency re-admission, with rates being highest for 18–19 y olds. Risks of death were higher than in adolescents discharged after accident-related injury , as were risks of re-admission . Risks of death and re-admission were increased after all combinations of violent, drug/alcohol-related, and self-inflicted injury, but particularly after any drug/alcohol-related or self-inflicted injury , for which age-adjusted hazard ratios for death in boys ranged from 1.67 to 5.35, compared with 1.25 following violent injury alone . The main limitation of the study was under-recording of adversity-related injuries and misclassification of these cases as accident-related injuries. This misclassification would attenuate the relative risks of death and re-admission for adversity-related compared with accident-related injury.We analysed National Health Service admissions data for England (1 April 1997–31 March 2012) for 10–19 y olds with emergency admissions for adversity-related injury  as a way to deal with the emotional upheaval of adolescence. Consequently, adolescents often sustain adversity-related injuries . In England, for example, a third of all adolescents admitted to hospital for any injury have an adversity-related injury; the remaining injuries are mainly accident-related. Because adolescents who present with an adversity-related injury often re-present later with other adversity-related injuries, hospitalization for such injuries is a “teachable moment,” a time when relevant interventions  can potentially reduce the risk of further harm. But which adolescents are likely to benefit from such interventions depends on their long-term risks of harm. Here, the researchers use hospitalization data for England to compare the ten-year risks of mortality (death) and re-admission among adolescents after adversity-related injury with the risks after accident-related injury.The researchers used National Health Service hospital admissions data collected between 1997 and 2012 for 10–19 year olds with emergency admissions for adversity-related or accident-related injury  to estimate the ten-year risks of death and emergency re-admission among injured adolescents after discharge. Among adolescents discharged after an adversity-related injury, one in 137 girls and one in 64 boys died within ten years. Also, 54.2% of girls and 40.5% of boys had a subsequent emergency re-admission for an adversity-related injury, and emergency re-admission rates were highest in 18–19 year olds. The risks of both death and emergency re-admission were higher among adolescents discharged after an adversity-related injury than among adolescents discharged after an accident-related injury. For example, boys discharged after an adversity-related injury were about twice as likely to die within the next ten years as boys discharged after an accident-related injury. Risks of death were increased after all combinations of adversity-related injury but particularly after combinations that included drug- or alcohol-related or self-inflicted injury. Finally, the risks of emergency re-admission were highest after injuries that included self-inflicted injury in both girls and boys.These findings indicate that adolescents discharged from the hospital after an admission for violence-related, drug- or alcohol-related, or self-inflicted injuries have increased risks of subsequent harm up to a decade later. Misclassification of some adversity-related injuries as accident-related injuries may affect the accuracy of these findings. Another important limitation of this observational study is residual confounding. That is, although the researchers adjusted for known factors likely to affect the risk of re-admission or death in their analysis, the adolescents who were re-admitted or died subsequent to discharge after an adversity-related injury may have shared other unknown characteristics that were responsible for their increased risk of harm. Nevertheless, these findings identify several risk factors that clinicians and service providers can use to identify those adolescents admitted to hospital with an injury who are at high or low risk of subsequent harm. Specifically, these findings suggest that the introduction of strategies for reducing subsequent harm after discharge should be considered for all types of adversity-related injury, particularly when it occurs in older adolescents.http://dx.doi.org/10.1371/journal.pmed.1001931.This list of resources contains links that can be accessed when viewing the PDF on a device or via the online version of the article at adolescence  and links to WHO documents concerned with adolescent healthThe World Health Organization (WHO) provides brief information on surviving adolescence for parents, teachers, and young peopleThe UK Royal College of Psychiatrists provides a fact sheet on stages of adolescence (in English and Spanish)The American Academy of Pediatrics also provides information on the Young Minds has real stories about self-harm and about other aspects of emotional well-being and mental health among young peopleThe UK not-for-profit organization adolescent development, self-harm, teen violence, and injuriesMedlinePlus provides basic information and links to further resources about violence, self-harm, and drug or alcohol misuse among adolescents admitted to the hospital for injuries is availableA previous open-access paper by the researchers about  Adolescents (10–19 y olds) are a vulnerable population –4. TheseA presentation to hospital for injury related to violence, drug/alcohol misuse, or self-harm provides an opportunity to assess adolescents’ psychosocial needs or initiate interventions, at a time when these individuals may be willing to consider behaviour change –10. CurrWe have previously reported the prevalence of emergency admissions for violent, drug/alcohol-related, and self-inflicted injury for adolescents in England. Approximately 4% of girls and boys have such an admission at least once between the ages of 10 and 19 y old, accounting for one-third of adolescents admitted with any injury (the majority of the remaining two-thirds are related to accidents) . In EnglWe used national hospitalisation data for England to determine the cumulative risks of death and emergency re-admission in adolescents over the 10 y after discharge following an admission for violent, drug/alcohol-related, or self-inflicted injury. We determined whether risks after adversity-related injury were increased compared with after accident-related injury (our hypothesis was that they would be). We also examined whether risks of death or emergency re-admission differed by sex and age or were associated with underlying chronic conditions, ethnicity, or deprivation.We used anonymised Hospital Episode Statistics (HES) data comprising all hospital admissions to the National Health Service (NHS) in England from 1 April 1997 to 31 March 2012 . We compWe identified adolescents (aged 10–19 y inclusive) who had one or more emergency  admissions for injury. We considered any multiple admissions within 1 d of each other, or relating to a hospital transfer, to be the same admission. We defined “emergency” admissions by the HES “method of admission” variable , and “inWe received a standard, de-identified data extract from the Health and Social Care Information Centre (HSCIC), which does not require research ethics approval or patient consent . The oriWe used ICD-10 codes recorded in the index admission record to define two cohorts of adolescents who were alive at discharge . We defiThe outcomes were death and emergency re-admission at least 1 d after discharge from the index admission and up to 10 y later. Death was captured by linking hospital admissions to UK death registration data from the Office for National Statistics . Re-admiWe included age, chronic condition status, deprivation (socioeconomic status), and ethnicity, as recorded at the index admission, as possible confounding variables, or independent risk factors, for death and re-admission, based on previous studies that have shown associations between these factors and harm in adolescents ,24. OtheWe grouped adolescents into three age groups  to reflect stages of development . We grouWe addressed data quality by replacing missing or inconsistent variables with the corresponding modal value for all admission records on the same individual. We replaced 0.8% of records with modal values for sex, 30.2% for ethnicity, and 1.8% for deprivation value. We analysed any residual missing variables for sex, ethnicity, or deprivation as “missing”. No data were missing for age or chronic condition status.All analyses were carried out separately for girls and boys, given well-established differences in the frequency of adversity-related injury between the sexes . We usedTo determine the absolute risks of death and emergency re-admission in each cohort, we calculated Kaplan–Meier cumulative probabilities and 95% confidence intervals (CIs) for each age group from 1 d to 10 y after discharge from the index admission. We also calculated 1-, 5-, and 10-y risks of death and re-admission following any violent, any drug/alcohol-related, and any self-inflicted injury. To allow comparison of the risks of death in the adversity-related and accident-related injury cohorts with those in the general population, we derived general population estimates of risks of death in 10–19 y olds in 1997–1999 for the next 1–10 y in 1-y increments using aggregate statistics published by the Office for National Statistics  with 95% CIs. Models were adjusted for confounders in stages. We first estimated crude HRs between the two cohorts, and then estimated HRs adjusting for age; age and chronic condition status; and age, chronic condition status, ethnicity, and deprivation. Estimated HRs were attenuated after adjusting for age, . However, further adjustments for chronic condition status, ethnicity, and deprivation did not substantially alter age-adjusted HRs. We therefore present the main comparison of the two cohorts adjusted by age only. However, because chronic condition status and deprivation were independently associated with death and re-admission, we report the absolute 10-y risks of these outcomes after adversity-related and accident-related injury in Table A6 in As different types of adversity-related injury tend to co-occur , we estiWe tested the goodness of fit of the Cox regression models by plotting the Nelson–Aalen estimate of the cumulative hazard function against Cox–Snell residuals . Analysen = 333,009) formed the adversity-related injury cohort, and 60%  formed the accident-related injury cohort . Compared with the accident-related injury cohort, adolescents in the adversity-related injury cohort were on average older at their index admission, more likely to have a chronic condition, and more likely to be from the most deprived areas according to IMD score . The mosIn the adversity-related injury cohort, girls were admitted predominantly for drug/alcohol-related (90.1%) or self-inflicted (72.4%) injury, while boys were most often admitted for violent (46.7%) or drug/alcohol-related (56.5%) injury. The distribution of ethnicity did not differ substantially between the two cohorts. The median follow-up time from the index admission ranged from 6.8 to 7.7 y in both cohorts   and 15.6/1,000 for boys , compared with 3.7/1,000 girls  and 6.0/1,000 boys  in the accident-related injury cohort  higher in girls and 113% (95% CI 98%–129%) higher in boys, after adjusting for age  and 40.5% for boys (95% CI 40.2%–40.9%) , adjusted for age, chronic condition status, ethnicity, and deprivation are presented in Table A4 in We found no association between ethnicity and long-term risks of death or emergency re-admission after adjusting for deprivation and underlying chronic condition  and living in the most deprived areas  increased the risks of death and re-admission.One strength of our study is the use of administrative data to study the entire population of adolescents hospitalised with injury within the English publicly provided NHS. Second, linkage of all hospital admissions and death records made it possible to study outcomes 10 y after discharge from the index admission. Third, this is the first study to our knowledge that has quantified risks of harm in adolescents after all three of violent, drug/alcohol-related, and self-inflicted injury within the same cohort. Long-term follow-up has previously been reported only after self-inflicted injury .One limitation of our study is potential misclassification of violent, drug/alcohol-related, or self-inflicted injury as accident-related injury ,33, whicAnother limitation is that linkage error may lead to a failure to capture death or re-admission and is more common in certain ethnic minorities. For example, in a study where the HES linkage algorithm was applied in another routine paediatric dataset, children and adolescents with black or “other”  ethnicity were approximately 2.4 to 4.1 times more likely to have a false or missed match than their white counterparts . FailureFinally, HES data do not include information on interventions received by the adolescents during or after admissions, such as referral to mental health services. Effective interventions could have confounded the association between adversity-related injury and later harm. This confounding could have caused underestimation of the association between adversity-related injury and later harm if adolescents in the adversity group were more likely to receive effective interventions than those in the accident-related injury group. In practice, however, psychosocial assessments are routinely recommended only for adolescents with self-inflicted injury , and it Our estimates of the relative increase in risks of death and emergency re-admission are likely to be generalisable to other hospitalised adolescent populations within the UK. However, generalisability to other countries and healthcare systems depends on the similarities of their populations of adolescents admitted with adversity-related injury to that of England. Generalisability also depends on the availability and intensity of psychosocial support for vulnerable adolescents after hospital discharge. Patterns of self-harm and drug/alcohol use during adolescence in other parts of Europe ,38, but Mortality rates in this study were either similar to or slightly higher than those reported for 14–24 y olds presenting with violent injury to the emergency department in the US . The risAdolescents discharged from hospital after an adversity-related injury have substantially increased risks of death and emergency re-admission during the next 10 y compared with adolescents discharged after an accident-related injury; 10 y after discharge from hospital in England for an adversity-related injury, one in 52 boys and one in 90 girls aged 18–19 y will have died. These risks may be underestimated because of under-recording of adversity in hospital discharge records.The risk of future harm was increased after all types of adversity-related injury. These findings justify extending national policy for psychosocial assessment after self-inflicted injury to all types of adversity-related injury. Consideration of psychological and social circumstances is good clinical practice, particularly for vulnerable adolescents. However, extending mandated psychosocial assessment from self-inflicted injury to all three types of adversity-related injury may have implications for services, as injured young people aged 16 y or older are often managed on adult surgical wards in the NHS, where expertise in psychosocial assessment and support for young people may be limited. Those who are male, older, have an underlying chronic condition, or are from deprived areas, and those exposed to multiple types adversity  have the highest risks of future harm Click here for additional data file.S1 Fig(TIF)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOC)Click here for additional data file.S1 Text(DOC)Click here for additional data file.


Pain and function improve dramatically in the first three months after hip and knee arthroplasty but the trajectory after three months is less well described. It is also unclear how pre-operative pain and function influence short- and long-term recovery. We explored the trajectory of change in function and pain until and beyond 3-months post-operatively and the influence of pre-operative self-reported symptoms.The study was a prospective cohort study of 164 patients undergoing primary hip (n = 80) or knee (n = 84) arthroplasty in the United Kingdom. Self-reported measures of pain and function using the Western Ontario and McMaster Universities Osteoarthritis index were collected pre-operatively and at 3 and 12 months post-operatively. Hip and knee arthroplasties were analysed separately, and patients were split into two groups: those with high or low symptoms pre-operatively. Multilevel regression models were used for each outcome (pain and function), and the trajectories of change were charted (0–3 months and 3–12 months).Hip: Most improvement occurred within the first 3 months following hip surgery and patients with worse pre-operative scores had greater changes. The mean changes observed between 3 and twelve months were statistically insignificant. One year after surgery, patients with worse pre-operative scores had post-operative outcomes similar to those observed among patients with less severe pre-operative symptoms. Knee: Most improvement occurred in the first 3 months following knee surgery with no significant change thereafter. Despite greater mean change during the first three months, patients with worse pre-operative scores had not ‘caught-up’ with those with less severe pre-operative symptoms 12 months after their surgery.Most symptomatic improvement occurred within the first 3 months after surgery with no significant change between 3–12 months. Further investigations are now required to determine if patients with severe symptoms at the time of their knee arthroplasty have a different pre-surgical history than those with less severe symptoms and if they could benefit from earlier surgical intervention and tailored rehabilitation to achieve better post-operative patient-reported outcomes. Joint replacement is a common elective surgical procedure, with more than 158,000 primary hip and knee arthroplasties performed during 2013 in England, Wales and Northern Ireland .Many studies have reported important improvement in patient-reported outcomes by comparing scores completed at baseline and at one single post-operative assessment usually 6 or twelve months after the arthroplasty. FindingKnown determinants of post-operative outcome following hip and knee arthroplasties include patients’ expectations, mental health status, co-morbidities and the severity of pre-operative pain and function –25. PatiFrom the data available it is difficult for a surgeon to make an accurate prediction of how an individual patient is likely to recover post-operatively  and it iThe aims of our study were to use information obtained from a cohort study to describe the trajectory of change after primary hip and knee arthroplasty, and to determine the effects of pre-operative pain and function on these trajectories and on outcomes one year after surgery.th December 2009 by the Southwest 4 Research Ethics Committee (09/H0102/72) and all patients provided informed, written consent.ADAPT is a single-centre UK prospective cohort study including people undergoing hip and knee arthroplasties. The study is registered on the NIHR Clinical Research Network Portfolio (UKCRN ID 8311). Its main purpose is to assess ways in which function can be assessed. The study was approved on 24Detailed information on study design, patient recruitment, inclusion-exclusion criteria, and assessment methods are provided in the published study protocol . BrieflyAssessments were conducted prior to surgery  and then at 3 and 12 months after surgery. At each post-operative assessment time, participants completed a postal questionnaire. Pain and functional ability were assessed by self-report using the Western Ontario and McMaster Universities Osteoarthritis index (WOMAC) function and pain sub-scales . The WOMTo ensure we had a sufficient number of patients to perform meaningful data analysis, we aimed to recruit approximately 250 patients for the main ADAPT study. However, the trajectory of recovery was expected to differ between patients listed for primary surgery and those listed for revision surgery. The latter participants were not considered in this research and analyses reported here focus on the group of ADAPT participants who underwent a primary arthroplasty. Analyses were conducted separately for patients undergoing hip and knee surgery. To investigate the influence of pre-operative function on the post-operative recovery pattern, patients were split into two groups: those with high or low pre-operative function using the median pre-operative WOMAC-function score as a cut-point. The median was chosen as a cut-point to prevent major imbalance by creating groups of equal size. A similar strategy was used for pain. Two participants undergoing hip surgery and one undergoing knee surgery who participated in the post-operative follow-up had missing pre-operative pain and function WOMAC scores. Their pre-operative high/low profiles were determined with a single imputation technique using regression models and the longitudinal measures of WOMAC-pain and function as independent factors.We used univariate linear multi-level regression analysis, one for each outcome (WOMAC-pain and WOMAC-function) to model longitudinal trajectories (group changes) and conduct between-group comparisons of the change pattern. This approach accounts for repeated and unequal number of measurements per participant while producing estimations valid under the missing at random assumption . FunctioChange was modelled as two linear splines (a line between two points): one spline for the “short-term change” occurring between the pre-operative assessment and the second assessment (3 months post-operative) and another spline for the “long-term change” occurring between the two post-operative assessments (3 and 12 months). These changes were normally distributed  allowing us to use the models presented above. The regression models were stratified by high/low pre-operative status. They were adjusted for the above two time splines, and random effects on the intercepts and slopes were modelled. Differences in the short-term or long-term changes between groups were tested using appropriate linear contrasts. Contrary to the changes between measurement points, outcome scores at each post-operative measurement occasion (3 months or 12 months) were non-normally distributed. Group comparisons of time-specific post-operative observed scores were therefore conducted with Mann-Whitney tests. A p-value of <0.05 was considered statistically significant. All models were fitted using Stata SE 13.1 and MLwiN v2.31 using Stata runmlwin command .Overall, 1451 eligible patients listed for hip or knee arthroplasty were approached about the ADAPT study and 264 agreed to take part (recruitment rates of 20% for patients waiting for hip surgery and 17% for those waiting for knee surgery). Of those who gave informed consent to participate, we excluded three people who did not subsequently undergo surgery and 12 people in whom all pre-operative data were unavailable. Patients who underwent a revision surgery (n = 85) were not considered for this study. Of those included in the final analysis, 80 had a primary total hip arthroplasty and 84 had knee arthroplasty .At 12 months post-operative, 89% of the participants who underwent primary hip surgery and 86% of those who underwent primary knee surgery had complete WOMAC-pain and function scores. These rates were comparable by pre-operative high/low pain or function status using Fisher’s exact test: hip: pain p = 0.73, function p = 0.15; knee: pain p = 0.54, function p = 0.76. For hip surgery, three patients withdrew after their surgery, one after the three months assessment and five were lost to follow-up. For knee surgery, six patients withdrew after their surgery, two after the three months assessment and four were lost to follow-up.2  for hip participants and 31 kg/m2  for knee participants.The demographics and clinical characteristics of the cohort are shown in The distributions of participants by high or low pre-operative WOMAC-function and pain scores are presented in Self-report functional ability and pain both improved after surgery, as shown in Improvements in function and pain mainly occurred within the first 3 months. Patients with low pre-operative function experienced short-term functional improvements that were larger than those reported by patients with high pre-operative function, between-group difference in monthly mean change in WOMAC-function standardised score 0.39 (95% CI 0.30, 0.48; p<0.001, Short-term pain improvements were also larger for those with high pre-operative pain, between-group difference in monthly mean change in WOMAC-pain standardised score 0.42 (95%CI 0.34, 0.51, Patients experienced an improvement in self-reported function and pain after their knee surgery, with this improvement occurring primarily in the first three months, as shown in Patients with low pre-operative function reported short-term functional improvements that were larger than those reported by patients with high pre-operative function, between-group difference in monthly mean change in WOMAC-function standardised score 0.14 (95%CI 0.04, 0.25; p <0.01, Short-term pain improvements in pain were also larger for participants with high pre-operative pain compared to those with low pre-operative pain, between-group difference in monthly mean change in WOMAC-pain standardised score 0.17 (95%CI 0.03, 0.30; p = 0.014, The majority of the symptomatic gain made after both hip and knee arthroplasty was achieved within the first three months after surgery. No significant mean change beyond three months was observed in our sample. Patients with worse pain and functional limitations prior to surgery had the greatest capacity to benefit from arthroplasty. They had larger improvements during the first three months of their surgery than those with better pre-operative scores; beyond this period, the mean changes in pain and function were not related to pre-operative scores. One year after surgery, the levels of pain and function achieved by those who underwent a primary total hip arthroplasty were not dependent on pre-operative pain and function. In contrast, patients who had the most severe pain and functional impairment prior to knee arthroplasty never achieved as good an outcome as those with less severe pre-operative pain and functional limitations.Our results on improvements in function and pain after surgery are in agreement with previous studies which show that most of the recovery as measured by self-reported outcomes occurs in the first few months after arthroplasty , 16, 39.We have found that the associations between pre-operative pain and function scores and the course of recovery differ between patients undergoing hip and knee arthroplasty. Our findings showing that the level of pre-operative pain/function influences the course of recovery and level of score reached at 12 months after knee surgery is consistent with previous research , 28, 40.The absence of an association between the pre-operative score and post-operative pain and function outcomes following hip arthroplasty is in disagreement with several studies , 28, 40 The absence of significant improvements observed after three months may be a consequence, at least in part, from the ceiling effect inherent in patient reported outcome measures used in arthroplasty research –46. PatiThe strengths of the study include the use of robust and validated outcome measures, good follow-up rates, more than one follow-up point, and a reasonable sample size for this type of investigation. The use of linear mixed regression models allowed the modelling of the repeated measures of pain and function and facilitated the use of all available observations including those of patients who did not participate in all follow-ups while providing estimations valid under the missing at random assumption.The apparent limitation of a relatively low participation rate is not, in our view, a major problem because this was not a trial, the overall demographics of those taking part are similar to those found in other studies, and we achieved the wide variation in disease severity at baseline which we were aiming for. Participation rates are explained by the high burden of attending additional research appointments in the ADAPT study. It is possible that those who agreed to take part differed from those who declined with regard to important determinants of the outcome of arthroplasty, such as mental health. Furthermore, this cohort is a single centre study limiting its external validity as context and culture may influence outcomes .The modest sample size limited our ability to adjust for factors known to be associated with post-operative outcomes such as age, gender, mental health and co-morbidities , 50, 51.We analysed results of patients who underwent unicompartmental and total knee arthroplasty together to keep a large enough study group. Evidence suggests that following unicompartmental knee surgery there is a faster recovery and lower rates of adverse events , 53. OthMeasuring outcomes only at 3 and 12 months is another limitation of the study, as inclusion of additional assessment points would have allowed more detailed investigation of recovery trajectories. We considered that additional assessment points would be too much of a burden for participants with a probability of increased levels of attrition.Losina and Katz  discusseMost of the improvement following hip and knee arthroplasty occurs within the first three post-operative months with no subsequent statistically significant improvement. Patients with worse pre-operative function or pain report poorer outcome at 12 months after knee arthroplasty, but not hip arthroplasty. Further investigations are now required to determine if patients with severe symptoms at the time of their knee arthroplasty have a different pre-surgical history than those with less severe symptoms and if they could beneficiate from earlier surgical intervention, when symptoms are less severe, and/or tailored pre- and post-operative rehabilitation to achieve better post-operative patient-reported outcomes.