BACKGROUND: Clinical trials showed a single oral dose of doxycycline taken after sex protects against STIs among men who have sex with men (MSM) but not women. Pharmacokinetic data at vaginal, rectal and penile sites of STI exposure are lacking. We examined vaginal, rectal and urethral doxycycline concentrations in men and women to better inform STI prevention. METHODS: Doxycycline pharmacokinetics in male and female participants 18-59 years of age were evaluated in blood and urine and on rectal and vaginal swabs collected at 1, 2, 4, 8, 24, 48, 72, 96 and 168 h after receiving a 200 mg oral doxycycline dose in a non-randomised single dose open label single centre study in Atlanta, Georgia. Rectal, vaginal, and cervical biopsies and male urethral swabs were collected 24 h after dosing (Trial registration: NCT04860505). Doxycycline was measured by liquid chromatography-mass spectrometry. FINDINGS: Eleven male and nine female participants participated in the study. Doxycycline concentrations on rectal and vaginal swabs collected up to 96 h after dosing were approximately twice those of plasma and remained above minimum inhibitory concentrations (MICs) for at least four, three, and two days for Chlamydia trachomatis, Treponema pallidum, and tetracycline-sensitive Neisseria gonorrhoeae, respectively. Geometric mean doxycycline concentrations in male urethral secretions (1.166 μg/mL; 95% CI 0.568-2.394 μg/mL), male rectal (0.596 μg/g; 0.442-0.803 μg/g), vaginal (0.261 μg/g; 0.098-0.696 μg/g) and cervical tissue (0.410 μg/g; 0.193-0.870 μg/g) in biopsies collected 24 h after dosing exceeded MICs. Plasma and urine doxycycline levels defined adherence markers up to four and seven days postdosing, respectively. No adverse events were reported in this study. INTERPRETATION: Doxycycline efficiently distributes to the rectum, vagina and urethra. Findings can help explain efficacy of STI prevention by doxycycline. FUNDING: Funded by CDC intramural funds, CDC contract HCVJCG-2020-45044 (to CFK).

INTRODUCTION: Globally, many young women face the overlapping burden of HIV infection and unintended pregnancy. Protection against both may benefit from safe and effective multipurpose prevention technologies. METHODS: Healthy women ages 18-34 years, not pregnant, seronegative for HIV and hepatitis B surface antigen, not using hormonal contraception, and at low risk for HIV were randomized 2:2:1 to continuous use of a tenofovir/levonorgestrel (TFV/LNG), TFV, or placebo intravaginal ring (IVR). In addition to assessing genital and systemic safety, we determined TFV concentrations in plasma and cervicovaginal fluid (CVF) and LNG levels in serum using tandem liquid chromatography-mass spectrometry. We further evaluated TFV pharmacodynamics (PD) through ex vivo CVF activity against both human immunodeficiency virus (HIV)-1 and herpes simplex virus (HSV)-2, and LNG PD using cervical mucus quality markers and serum progesterone for ovulation inhibition. RESULTS: Among 312 women screened, 27 were randomized to use one of the following IVRs: TFV/LNG (n = 11); TFV-only (n = 11); or placebo (n = 5). Most screening failures were due to vaginal infections. The median days of IVR use was 68 [interquartile range (IQR), 36-90]. Adverse events (AEs) were distributed similarly among the three arms. There were two non-product related AEs graded >2. No visible genital lesions were observed. Steady state geometric mean amount (ssGMA) of vaginal TFV was comparable in the TFV/LNG and TFV IVR groups, 43,988 ng/swab (95% CI, 31,232, 61,954) and 30337 ng/swab (95% CI, 18,152, 50,702), respectively. Plasma TFV steady state geometric mean concentration (ssGMC) was <10 ng/ml for both TFV IVRs. In vitro, CVF anti-HIV-1 activity showed increased HIV inhibition over baseline following TFV-eluting IVR use, from a median of 7.1% to 84.4% in TFV/LNG, 15.0% to 89.5% in TFV-only, and -27.1% to -20.1% in placebo participants. Similarly, anti-HSV-2 activity in CVF increased >50 fold after use of TFV-containing IVRs. LNG serum ssGMC was 241 pg/ml (95% CI 185, 314) with rapid rise after TFV/LNG IVR insertion and decline 24-hours post-removal (586 pg/ml [95% CI 473, 726] and 87 pg/ml [95% CI 64, 119], respectively). CONCLUSION: TFV/LNG and TFV-only IVRs were safe and well tolerated among Kenyan women. Pharmacokinetics and markers of protection against HIV-1, HSV-2, and unintended pregnancy suggest the potential for clinical efficacy of the multipurpose TFV/LNG IVR. CLINICAL TRIAL REGISTRATION: NCT03762382 [https://clinicaltrials.gov/ct2/show/NCT03762382].

BACKGROUND: Given future potential use of vaginal rings to prevent HIV infection, we examined the association of contraceptive vaginal ring (CVR) non-adherence with user dissatisfaction, tolerability, demographic, and behavioral factors. METHODS: In an open-label single-group study, sexually active women aged 18-34 years using oral or injectable hormonal contraception, conveniently sampled from general population, were assigned to 6-month use of a commercial CVR currently not licensed for use in Kenya. Non-adherence in any CVR cycle completed was assessed from: (1) self-report (not used for at least 1 day), and (2) pharmacy record (failure to timely receive a new CVR or return a used one). Additionally, non-adherence was assessed in a subset of participants by residual progestin and estrogen levels measured in returned CVRs. RESULTS: Of 202 participants who underwent CVR insertion by a study clinician, 142 completed all 6 visits, 172 responded to questions about ring use, and 43 provided used CVRs from months 1, 3, and 6 for residual hormone analysis. Non-adherence was 14.0% (24/172) by self-report and 54.5% (110/202) by pharmacy record. Non-adherence by pharmacy record was significantly reduced among women with a salary-based income (prevalence ratio (PR) 0.71, 95% confidence interval (CI) (0.55-0.91)] compared to women with income not salary-based or no income. Participants dissatisfied with CVR on ≥4 aspects (ambiguity of instructions, inconvenience of use, sensation, sexual discomfort, etc.) were more likely to report non-adherence (PR 2.69, 95% CI=(1.31-5.52)] compared to those dissatisfied with ≤3 aspects. Non-adherence by residual hormone levels was identified in 46.5% (20/43) participants. Over time, this subset of participants showed increasing non-adherence (P=0.004). We found lack of agreement among the various measures of non-adherence. CONCLUSIONS: Economic empowerment interventions, especially those emphasizing partner-independent income options, and expanded education on CVR features may alleviate non-adherence. Addressing CVR dissatisfaction preemptively may also help mitigate non-adherence.

BACKGROUND: Event-driven HIV prevention strategies are a priority for users who do not require daily pre-exposure prophylaxis (PrEP). Regimens containing integrase strand transfer inhibitors (INSTIs) are under evaluation as alternatives to daily PrEP. To better understand INSTI distribution and inform dosing selection we compared the pharmacology of two-dose boosted elvitegravir and unboosted bictegravir regimens in MSM. MATERIALS AND METHODS: Blood, rectal and penile secretions and rectal biopsies were collected from 63 HIV-negative MSM aged 18-49 years. Specimens were collected up to 96 h after two oral doses of tenofovir alafenamide and emtricitabine with elvitegravir boosted by cobicistat or unboosted bictegravir given 24 h apart. Antiretroviral drugs were measured by LC-MS. RESULTS: Mean bictegravir plasma concentrations remained above the 95% protein-adjusted effective concentration 96 h after dosing [273 (95% CI: 164-456) ng/mL] whereas elvitegravir plasma concentrations became undetectable 48 h after the second dose. Bictegravir and elvitegravir reached rectal tissues within 2 h after the first dose, and elvitegravir tissue concentrations [1.07 (0.38-13.51) ng/mg] were greater than bictegravir concentrations [0.27 (0.15-0.70) ng/mg]. Both INSTIs became undetectable in tissues within 96 h. Elvitegravir and bictegravir were not consistently detected in penile secretions. CONCLUSIONS: Whereas bictegravir plasma concentrations persist at least 4 days after a two-oral-dose HIV prophylaxis regimen, elvitegravir accumulates in mucosal tissues. Differing elvitegravir and bictegravir distribution may result in variable mucosal and systemic antiviral activity and can inform dosing strategies for event-driven HIV prevention.

BACKGROUND: HIV pre-exposure prophylaxis (PrEP) persistence and adherence are critical to ending the HIV epidemic in the United States. SETTING: In 2017 National HIV Behavioral Surveillance, HIV-negative men who have sex with men (MSM) in 4 U.S. cities completed a survey, HIV testing, and dried blood spots (DBS) at recruitment. METHODS: We assessed three PrEP outcomes: persistence (self-reported PrEP use at any time in the past 12 months and had tenofovir, emtricitabine, or tenofovir diphosphate (TFV-DP) detected in DBS), adherence at ≥4 doses/week (self-reported past-month PrEP use and TFV-DP concentration ≥700 fmol/punch), and adherence at 7 doses/week (self-reported past-month PrEP use and TFV-DP concentration ≥1250 fmol/punch). Associations with key characteristics were examined using log-linked Poisson regression models with generalized estimating equations. RESULTS: Among 391 MSM who took PrEP in the past year, persistence was 80% and was lower among MSM who were younger, had lower education, and had fewer sex partners. Of 302 MSM who took PrEP in the past month, adherence at ≥4 doses/week was 80% and adherence at 7 doses/week was 66%. Adherence was lower among MSM who were younger, were Black, and had fewer sex partners. CONCLUSIONS: Although persistence and adherence among MSM were high, 1 in 5 past-year PrEP users were not persistent and 1 in 5 past-month PrEP users were not adherent at levels that would effectively protect them from acquiring HIV (i.e., ≥4 doses/week). Efforts to support PrEP persistence and adherence should include MSM who are young, are Black, and have less education.

OBJECTIVE: To determine whether a clinician-directed acute respiratory tract infection (ARI) intervention was associated with improved antibiotic prescribing and patient outcomes across a large US healthcare system. DESIGN: Multicenter retrospective quasi-experimental analysis of outpatient visits with a diagnosis of uncomplicated ARI over a 7-year period. PARTICIPANTS: Outpatients with ARI diagnoses: sinusitis, pharyngitis, bronchitis, and unspecified upper respiratory tract infection (URI-NOS). Outpatients with concurrent infection or select comorbid conditions were excluded. INTERVENTION(S): Audit and feedback with peer comparison of antibiotic prescribing rates and academic detailing of clinicians with frequent ARI visits. Antimicrobial stewards and academic detailing personnel delivered the intervention; facility and clinician participation were voluntary. MEASURE(S): We calculated the probability to receive antibiotics for an ARI before and after implementation. Secondary outcomes included probability for a return clinic visits or infection-related hospitalization, before and after implementation. Intervention effects were assessed with logistic generalized estimating equation models. Facility participation was tracked, and results were stratified by quartile of facility intervention intensity. RESULTS: We reviewed 1,003,509 and 323,023 uncomplicated ARI visits before and after the implementation of the intervention, respectively. The probability to receive antibiotics for ARI decreased after implementation (odds ratio [OR], 0.82; 95% confidence interval [CI], 0.78-0.86). Facilities with the highest quartile of intervention intensity demonstrated larger reductions in antibiotic prescribing (OR, 0.69; 95% CI, 0.59-0.80) compared to nonparticipating facilities (OR, 0.89; 95% CI, 0.73-1.09). Return visits (OR, 1.00; 95% CI, 0.94-1.07) and infection-related hospitalizations (OR, 1.21; 95% CI, 0.92-1.59) were not different before and after implementation within facilities that performed intensive implementation. CONCLUSIONS: Implementation of a nationwide ARI management intervention (ie, audit and feedback with academic detailing) was associated with improved ARI management in an intervention intensity-dependent manner. No impact on ARI-related clinical outcomes was observed.

OBJECTIVES: The Core Elements of Outpatient Antibiotic Stewardship provides a framework to improve antibiotic use, but cost-effectiveness data on implementation of outpatient antibiotic stewardship interventions are limited. We evaluated the cost-effectiveness of Core Element implementation in the outpatient setting. METHODS: An economic simulation model from the health-system perspective was developed for patients presenting to outpatient settings with uncomplicated acute respiratory tract infections (ARI). Effectiveness was measured as quality-adjusted life years (QALYs). Cost and utility parameters for antibiotic treatment, adverse drug events (ADEs), and healthcare utilization were obtained from the literature. Probabilities for antibiotic treatment and appropriateness, ADEs, hospitalization, and return ARI visits were estimated from 16,712 and 51,275 patient visits in intervention and control sites during the pre- and post-implementation periods, respectively. Data for materials and labor to perform the stewardship activities were used to estimate intervention cost. We performed a one-way and probabilistic sensitivity analysis (PSA) using 1,000,000 second-order Monte Carlo simulations on input parameters. RESULTS: The proportion of ARI patient-visits with antibiotics prescribed in intervention sites was lower (62% vs 74%) and appropriate treatment higher (51% vs 41%) after implementation, compared to control sites. The estimated intervention cost over a 2-year period was $133,604 (2018 US dollars). The intervention had lower mean costs ($528 vs $565) and similar mean QALYs (0.869 vs 0.868) per patient compared to usual care. In the PSA, the intervention was dominant in 63% of iterations. CONCLUSIONS: Implementation of the CDC Core Elements in the outpatient setting was a cost-effective strategy.

BACKGROUND: HIV exposure to penile tissues provides a risk of acquisition among men, yet studies evaluating penile antiretroviral (ARV) drug distribution have been lacking. We measured ARVs on urethral and glans surface swabs collected following a dose of tenofovir alafenamide, emtricitabine, elvitegravir, darunavir and cobicistat. METHODS: Thirty-five HIV-negative male participants provided urethral swabs, glans swabs, rectal swabs, blood and urine up to 96 h following a single dose of tenofovir alafenamide/emtricitabine/elvitegravir/cobicistat and darunavir. ARVs were measured by liquid chromatography-mass spectrometry with a lower limit of detection (LOD) of 1 ng/swab for swabs and 10 ng/mL for plasma and urine. Concentrations are reported as median and range. RESULTS: Urethral swab emtricitabine and darunavir concentrations peaked at 4 h for emtricitabine (36 ng/swab; 3-307 ng/swab) and 8 h for darunavir (25 ng/swab; 2-52 ng/swab). Glans swab emtricitabine and darunavir concentrations peaked 24 h after dosing (emtricitabine 14 ng/swab, <LOD-328 ng/swab; darunavir 6 ng/swab, <LOD-149 ng/swab). Estimated peak urethral secretion emtricitabine and darunavir concentrations are between 10 and 20 μg/mL, similar to rectal secretions, 4-fold greater than in plasma, but 2-fold lower than in urine. Tenofovir and elvitegravir were detected on less than 20% of urethral or glans swabs collected within 24 h of dosing. CONCLUSIONS: We document ARV dosing in the urethra and on the glans surface with high drug concentrations noted for emtricitabine and darunavir and lower tenofovir and elvitegravir concentrations. Data suggest a potential protective role of urethral emtricitabine or darunavir against penile HIV acquisition.

Point of care (POC) tests for antiretroviral drugs (ARVs) could help improve individual adherence. This study sought to define the utility of urine, blood, and buccal swabs as minimally invasive specimens amenable to development of POC tests for ARVs. Urine, dried blood spots (DBS) and buccal swabs were collected from 35 HIV-negative men between 2 and 96 hours following a single dose of tenofovir alafenamide (TAF)/emtricitabine (FTC)/elvitegravir (EVG)/cobicistat and darunavir (DRV). ARV concentrations were measured by high performance liquid chromatography-mass spectrometry. High concentrations of FTC, DRV and TFV were detectable in urine at least 24 hours after dosing. FTC, DRV and EVG remained detectable in DBS at least 24 hours post dose. FTC and DRV were detectable on buccal swabs up to 2- and 24-hours post dose, respectively. TFV was not detectable in DBS or buccal swabs collected between 2 and 96 hours after dosing. Variable distribution of ARVs in minimally invasive specimens highlights the challenge of developing POC assays for recent ARV exposure.

BACKGROUND: The Core Elements of Outpatient Antibiotic Stewardship provide a framework to improve antibiotic use, but evidence supporting safety are limited. We report the impact of Core Elements implementation within Veterans Health Administration sites. METHODS: A quasi-experimental controlled study assessed the effects of an intervention targeting antibiotic prescription for uncomplicated acute respiratory tract infections (ARI). Outcomes included per-visit antibiotic prescribing, treatment appropriateness, potential benefits and complications of reduced antibiotic treatment, and change in ARI diagnoses over a 3-year pre-implementation and 1-year post implementation period. Logistic regression adjusted for covariates [OR (95% CI)] and a difference-in-differences analysis compared outcomes between intervention and control sites. RESULTS: From 2014-2019, there were 16,712 and 51,275 patient-visits in 10 intervention and 40 control sites, respectively. Antibiotic prescribing rates pre-post implementation in intervention sites were 59.7% and 41.5%, respectively; in control sites they were 73.5% and 67.2%, respectively [difference-in-differences p<0.001]. The intervention site pre-post implementation odds ratio to receive appropriate therapy increased [1.67 (1.31, 2.14)] which remained unchanged within control sites [1.04 (0.91, 1.19)]. There was no difference in ARI-related return visits post-implementation [(-1.3% vs. -2.0%; difference-in-differences p=0.76] but all-cause hospitalization was lower within intervention sites [(-0.5% vs. -0.2%); difference-in-differences p=0.02]. The odds ratio to diagnose upper respiratory tract infection not otherwise specified compared to other non-ARI diagnosis increased post-implementation for intervention [1.27(1.21,1.34)] but not control [0.97(0.94,1.01)] sites. CONCLUSIONS: Implementation of the Core Elements was associated with reduced antibiotic prescribing for uncomplicated ARIs and a reduction in hospitalizations. ARI diagnostic coding changes were observed.

BACKGROUND: Daily oral pre- or post-exposure prophylaxis (PrEP or PEP) is highly effective in preventing HIV infection. However, many people find it challenging to adhere to a daily oral regimen. Chemoprophylaxis with single oral doses of antiretroviral drugs taken before or after sex may better adapt to changing or unanticipated sexual practices and be a desirable alternative to daily PrEP or PEP. We investigated willingness to use a single oral pill before or after sex among men who have sex with men (MSM) and assessed the biological efficacy of a potent antiretroviral combination containing elvitegravir (EVG), emtricitabine (FTC), and tenofovir alafenamide (TAF). METHODS: Data on willingness to use single-dose PrEP or PEP were obtained from the 2017 cycle of the American Men's Internet Survey (AMIS), an annual online behavioral surveillance survey of MSM in the United States. Antiretroviral drug levels were measured in humans and macaques to define drug distribution in rectal tissue and identify clinically relevant doses for macaque modeling studies. The biological efficacy of a single dose of FTC/TAF/EVG as PrEP or PEP was investigated using a repeat-challenge macaque model of rectal HIV infection. FINDINGS: Through pharmacokinetic assessment in humans and macaques we found that EVG penetrates and concentrates in rectal tissues supporting its addition to FTC/TAF to boost and extend chemoprophylactic activity. Efficacy estimates for a single oral dose given to macaques 4h before or 2h after SHIV exposure was 91•7%[35•7%-98•9%] and 100%, respectively, compared to 80•1%[13•9%-95•4%] and 64•6%[-19•4%-89•5%] when single doses were given 6 and 24h post challenge, respectively. A two-dose regimen at 24h and 48h after exposure was also protective [77•1%[1•7%-94•7%]. INTERPRETATION: Informed by user willingness, human and macaque pharmacokinetic data, and preclinical efficacy we show that single-dose prophylaxis before or after sex is a promising HIV prevention strategy. Carefully designed clinical trials are needed to determine if any of these strategies will be effective in humans. FUNDING: Funded by CDC intramural funds, CDC contract HCVJCG2-2016-03948 (to CFK), and a grant from the MAC AIDS Fund and by the National Institutes of Health [P30AI050409] - the Emory Center for AIDS Research (to MZ and TS).

CONTEXT: Antiretroviral therapy (ART) to treat and pre-exposure prophylaxis (PrEP) to prevent HIV infection are effective tools to help end the HIV epidemic. However, their use could affect HIV transfusion-transmission risk. OBJECTIVES: Three different ART/PrEP prevalence analyses in blood donors were conducted. METHODS: First, blood samples from HIV-positive and a comparison group of infection-nonreactive donors were tested under blind using liquid chromatography-tandem mass spectrometry for ART. Second, blood donor samples from infection-nonreactive, 18-45 year-old, male, first-time blood donors in six US locations were tested for emtricitabine and tenofovir. Third, in men who have sex with men (MSM) participating in the 2017 CDC National HIV Behavioral Surveillance (NHBS) from five US cities self-reported PrEP use proximate to donation was assessed. FINDINGS: In blind testing, no ART was detected in 300 infection-nonreactive donor samples, but in 299 HIV-confirmed infected donor samples, 46 (15.4%, 95% CI 11.5 - 20.0%) had evidence of ART. Of the 1,494 samples tested from first-time, male donors, 9 (0.6%, 95% CI 0.03 - 1.1%) had tenofovir and emtricitabine. In the NHBS MSM survey, 27 of 591 respondents (4.8%, 95% CI 3.2 - 6.9%) reported donating blood in 2016 or 2017 and PrEP use within the same time frame as blood donation. CONCLUSIONS: Persons who are HIV-positive and taking ART and persons taking PrEP to prevent HIV infection are donating blood. Both situations could lead to increased risk of HIV transfusion transmission if blood screening assays are unable to detect HIV in donations from infected donors.

BACKGROUND: While prior epidemiologic studies have suggested that injectable progestin-based contraceptive depot medroxyprogesterone acetate (DMPA) use may increase a woman's risk of acquiring HIV, recent data have suggested that DMPA users may be at a similar risk for HIV acquisition as users of the copper intrauterine device and levonorgestrel implant. Use of the etonogestrel Implant (Eng-Implant) is increasing but there are currently no studies evaluating its effect on HIV acquisition risk. OBJECTIVE: Evaluate the potential effect of the Eng-Implant use on HIV acquisition risk by analyzing HIV target cells and cytokine profiles in the lower genital tract and blood of adult premenopausal HIV-negative women using the Eng-Implant. METHODS: We prospectively obtained paired cervicovaginal lavage (CVL) and blood samples at 4 study visits over 16 weeks from women between ages 18-45, with normal menses (22-35 day intervals), HIV uninfected with no recent hormonal contraceptive or copper intrauterine device (IUD) use, no clinical signs of a sexually transmitted infection at enrollment and who were medically eligible to initiate Eng-Implant. Participants attended pre-Eng-Implant study visits (week -2, week 0) with the Eng-Implant inserted at the end of the week 0 study visit and returned for study visits at weeks 12 and 14. Genital tract leukocytes (enriched from CVL) and peripheral blood mononuclear cells (PBMC) from the study visits were evaluated for markers of activation (CD38, HLA-DR), retention (CD103) and trafficking (CCR7) on HIV target cells (CCR5+CD4+ T cells) using multicolor flow cytometry. Cytokines and chemokines in the CVL supernatant and blood plasma were measured in a Luminex assay. We estimated and compared study endpoints among the samples collected before and after contraception initiation with repeated-measures analyses using linear mixed models. RESULTS: Fifteen of 18 women who received an Eng-Implant completed all 4 study visits. The percentage of CD4+ T cells in CVL was not increased after implant placement but the percentage of CD4+ T cells expressing the HIV co-receptor CCR5 did increase after implant placement (p = 0.02). In addition, the percentage of central memory CD4+ T-cells (CCR7+) in CVL increased after implant placement (p = 0.004). The percentage of CVL CD4+, CCR5+ HIV target cells expressing activation markers after implant placement was either reduced (HLA-DR+, p = 0.01) or unchanged (CD38+, p = 0.45). Most CVL cytokine and chemokine concentrations were not significantly different after implant placement except for a higher level of the soluble lymphocyte activation marker (sCD40L; p = 0.04) and lower levels of IL12p70 (p = 0.02) and G-CSF (p<0.001). In systemic blood, none of the changes noted in CVL after implant placement occurred except for decreases in the percentage CD4 T-cells expressing HLA-DR+ T cells (p = 0.006) and G-CSF (p = 0.02). CONCLUSIONS: Eng-Implant use was associated with a moderate increase in the availability of HIV target cells in the genital tract, however the percentage of these cells that were activated did not increase and there were minimal shifts in the overall immune environment. Given the mixed nature of these findings, it is unclear if these implant-induced changes alter HIV risk.

BACKGROUND: Urine provides a minimally invasive specimen that may allow for development of rapid tests to detect antiretroviral drugs (ARVs) and provide opportunities to improve individual adherence. This study sought to determine if urine could provide a biomarker of adherence for currently approved PrEP and HIV treatment regimens. METHODS: Urine and blood were collected from 34 HIV-negative men who have sex with men aged 18-49 years enrolled in a clinical trial comparing 2 ARV regimens. Specimens were collected 4 and 24 hours after a single oral dose of tenofovir disoproxil fumarate (TDF)/emtricitabine (FTC) (n=10) or tenofovir alafenamide (TAF)/FTC/cobicistat (COBI)/elvitegravir (EVG) (n=8), or after 4 and 10 days of daily oral TDF/FTC (n=9) or TAF/FTC/COBI/EVG (n=7). Tenofovir (TFV), FTC, and EVG were measured by high performance liquid chromatography-mass spectrometry. RESULTS: Median urine FTC concentrations at 4 and 24 hours were similar between men receiving TDF/FTC (4 hours 147 microg/mL; 24 hours 10 microg/mL) and men receiving TAF/FTC/COBI/EVG (4 hours 333 microg/mL, p=0.173; 24 hours 13 microg/mL, p=0.681). Median urine TFV concentrations were lower among men receiving TAF/FTC/COBI/EVG (4 hours 1.2 microg/mL; 24 hours 0.8 microg/mL) compared to men receiving TDF/FTC (4 hours 17 microg/mL, p<0.001; 24 hours 7 microg/mL, p=0.001). Urine TFV concentrations remained reduced among men receiving TAF/FTC/COBI/EVG compared to men receiving TDF/FTC following daily dosing. EVG was not consistently measureable in urine. CONCLUSION: High urine FTC and TFV concentrations could provide an indication of adherence to daily oral dosing with TDF or TAF-based regimens used for treatment and prevention.

OBJECTIVE: As a proxy for undiagnosed HIV, CDC's National HIV Behavioral Surveillance (NHBS) monitors participants who report being unaware of their infection, defined as self-reporting an HIV-negative or unknown status during the interview but testing positive for HIV infection. We validated the NHBS measure of awareness among men who have sex with men (MSM) in 2014. DESIGN: We tested dried blood spots (DBS) from MSM who reported being unaware of their infection for seven antiretrovirals (ARVs). MSM unaware with >/=1 ARV detected were defined as misreporters. METHODS: Weighted percentages and 95% confidence intervals were calculated to compare characteristics among misreporters, non-misreporters, and those who self-reported as HIV-positive. Viral load (VL) was quantified with a validated assay using DBS. RESULTS: Of 1,818 HIV-positive MSM, 299 (16%) self-reported as HIV-negative or unknown infection status. Of these 299, 145 (49%) were considered misreporters based on ARV detection. Among the unaware, misreporters were more likely than non-misreporters to be older and have health insurance. Compared to self-reported HIV-positive MSM, misreporters were more likely to be black, be bisexual, and have perceived discrimination. Of 138 misreporters with VL data, 116 (84%) had an undetectable VL. CONCLUSIONS: ARV testing revealed that half of MSM classified as unaware of their infection misreported their status. While off-label PrEP use might explain the presence of ARVs, it is unlikely since many misreporters were virally suppressed, suggesting they were on HIV therapy. Biomarker validation of behavioral data can improve data quality and usefulness in NHBS and other studies.

INTRODUCTION: Oral pre-exposure prophylaxis (PrEP) with tenofovir disoproxil fumarate (TDF) and emtricitabine (FTC) is highly effective in preventing HIV infection among men who have sex with men (MSM). The effects of consistent personal lubricant use in the rectum on tissue PrEP drug concentrations and the rectal microbiota are unknown. We investigated rectal PrEP drug concentrations and the microbiota in MSM before and after repeated rectal application of a hyperosmolar lubricant. METHODS: We randomized 60 HIV-negative MSM to apply 4 mL of hyperosmolar rectal lubricant daily (n = 20), take daily oral TDF/FTC (n = 19), or both (n = 21) for seven days. Blood, rectal biopsies and rectal secretions were collected via rigid sigmoidoscopy before and on day 8 after product use. Tenofovir (TFV) and FTC as well as their intracellular metabolites tenofovir-diphosphate (TFV-DP), FTC-triphosphate (FTC-TP) were measured by HPLC-mass spectrometry. Rectal mucosal microbiota was sequenced with 16S rRNA sequencing using Illumina MiSeq. RESULTS: Seven days of lubricant application was not associated with differences in PrEP drug concentrations in rectal tissue or secretions. Lubricant use was associated with a decrease in the relative abundance of the Bacteroides genus (p = 0.01) and a non-significant increase in the Prevotella genus (p = 0.09) in the rectum. PrEP drug concentrations in rectal tissue and secretions were not associated with microbiota composition or diversity either before or after lubricant use. CONCLUSIONS: Repeated rectal application of a hyperosmolar lubricant does not affect mucosal PrEP drug concentrations but is associated with changes in the rectal microbiome.

Clinical trials demonstrated intermittent preventive treatment in pregnancy (IPTp) with mefloquine (MQ) reduced malaria rates among pregnant women, yet an unexpected higher risk of mother to child transmission (MTCT) of HIV among HIV-positive women receiving MQ has also been observed. To determine if interactions between antiretroviral drugs (ARVs) and MQ could contribute to the increased MTCT observed in women receiving MQ, we performed a retrospective cross-sectional analysis of ARVs in peripheral blood plasma (maternal plasma) and cord blood plasma (cord plasma) collected at delivery from 186 mothers participating in a randomized clinical trial of MQ compared to placebo in Kenya. Plasma zidovudine (AZT), lamivudine (3TC) and nevirapine (NVP) concentrations were measured by high-performance liquid chromatography-tandem mass spectrometry. ARVs were detected in maternal plasma and cord plasma specimens in similar proportions between the two study arms. Median concentrations of AZT and 3TC were not significantly lower in the MQ arm compared to the placebo arm for maternal plasma and cord plasma (p > 0.05). However, median NVP concentrations were significantly lower in the MQ study arm compared to the placebo study arm in both maternal plasma (1597 ng/mL vs. 2353 ng/mL, Mann-Whitney Rank Sum, p = 0.023) and cord plasma (2038 ng/mL vs. 2434 ng/mL, p = 0.048). Reduced NVP concentrations in maternal and cord plasma of women receiving MQ suggest MQ may affect NVP metabolism for both mother and infant. These results highlight the need to evaluate potential drug-drug interactions between candidate antimalarials and ARVs for use in pregnant women.

Background: The female genital tract (FGT) microbiome may affect vaginal pH and other factors that influence drug movement into the vagina. We examined the relationship between the microbiome and antiretroviral concentrations in the FGT. Methods: Over one menstrual cycle, 20 human immunodeficiency virus (HIV)-infected women virologically suppressed on tenofovir (TFV) disoproxil fumarate/emtricitabine and ritonavir-boosted atazanavir (ATV) underwent serial paired cervicovaginal and plasma sampling for antiretroviral concentrations using high-performance liquid chromatography-tandem mass spectrometry. Analysis of 16S ribosomal RNA gene sequencing of cervicovaginal lavage clustered each participant visit into a unique microbiome community type (mCT). Results: Participants were predominantly African American (95%), with a median age of 38 years. Cervicovaginal lavage sequencing (n = 109) resulted in a low-diversity mCT dominated by Lactobacillus (n = 40), and intermediate-diversity (n = 28) and high-diversity (n = 41) mCTs with abundance of anaerobic taxa. In multivariable models, geometric mean FGT:plasma ratios varied significantly by mCT for all 3 drugs. For both ATV and TFV, FGT:plasma was significantly lower in participant visits with high- and low-diversity mCT groups (all P < .02). For emtricitabine, FGT:plasma was significantly lower in participant visits with low- vs intermediate-diversity mCT groups (P = .002). Conclusions: Certain FGT mCTs are associated with decreased FGT antiretroviral concentrations. These findings are relevant for optimizing antiretrovirals used for biomedical HIV prevention in women.

Clinical trials of preexposure prophylaxis (PrEP) using antiretroviral medications have demonstrated success in preventing HIV infection among at-risk populations. Oral PrEP regimens containing the nucleotide and nucleoside reverse transcriptase inhibitors tenofovir disoproxil fumarate (TDF) and emtricitabine (FTC) are highly effective among adherent study participants [1–4]. Studies of dosing patterns from clinical trials indicate increased PrEP efficacy is associated with greater adherence to oral daily dosing regimens [5,6]. However, measuring adherence for persons taking oral PrEP regimens is challenging and has relied primarily on self-report [7]. Laboratory methods of determining adherence to antiretroviral drug regimens involve mass spectrometry analyses of accumulated drug metabolites in hair or blood specimens [8,9]. These assays provide reliable measures of cumulative drug exposure predictive of protection against HIV, yet are technologically complex and expensive. The availability of rapid, inexpensive, and noninvasive methods to assess adherence to PrEP regimens would allow for opportunities to provide timely feedback regarding individual adherence. TDF and FTC are metabolized and excreted primarily in urine [10], and urine tenofovir (TFV) has recently been used to indicate PrEP adherence in an individual using chewed Truvada (TDF/FTC) [11]. Therefore, we sought to determine whether urine might provide a specimen type amenable to development of noninvasive methods to measure PrEP drug exposure. | Urine and peripheral blood were collected from 23 MSM on day 8 after taking daily oral Truvada (Gilead Sciences, Inc., Foster City, California, USA) for 7 consecutive days, and 10 MSM not taking Truvada as controls. TFV and FTC were measured in blood plasma and urine specimens by liquid chromatography mass spectrometry using methodology similar to that previously described with a lower limit of quantification of 10 ng/ml [12]. Standard curves were prepared by spiking solutions of known TFV and FTC concentration into normal human urine diluted 1 : 100 in 0.2% formic acid. Participant urine specimens were diluted 1 : 100 in 0.2% formic acid to obtain measurements within the linear range of the standard curve. Intracellular metabolites tenofovir diphosphate and FTC triphosphate were measured in methanol extracts of peripheral blood mononuclear cells (PBMC) as previously described [13].

OBJECTIVE: This study sought to measure residual contraceptive hormone levels in vaginal rings as an adherence marker for monitoring product use in clinical trials. STUDY DESIGN: Residual etonogestrel and ethinyl estradiol levels from used NuvaRings(R) of 26 self-reported adherent women enrolled in a clinical trial of vaginal ring acceptability were compared to those from 16 women who used NuvaRing(R) as their contraceptive choice. RESULTS: Twenty-one (81%) clinical trial rings had contraceptive hormone levels within the range of those used as a contraceptive choice. Five returned rings had unused or discordant levels of residual contraceptive hormones. CONCLUSION: Residual vaginal ring drug levels could help assess adherence in clinical trials.

Successful clinical trials of antiretroviral pre-exposure prophylaxis (PrEP) to prevent HIV infection have been reported in heterosexual men and women, men who have sex with men (MSM), and injection drug users1. A daily oral drug regimen containing nucleoside reverse transcriptase inhibitors (NRTIs) tenofovir disoproxil fumarate (TDF) alone or in combination with emtricitabine (FTC) have been effective when participant adherence is high1. Analysis of PrEP dosing patterns estimate taking at least four TDF doses per week provides 96% protection from HIV infection and at least two doses per week provides 76% protection in MSM and transgender women2, though some estimate more frequent dosing in heterosexual women may be needed3. TDF and FTC require phosphorylation in HIV target cells to tenofovir-diphosphate (TFV-DP) and emtricitabine-triphosphate (FTC-TP) to provide PrEP protection as competitive analogs of naturally occurring deoxyadenosine triphosphate (dATP) and deoxycytidine triphosphate (dCTP), respectively. However, it is unclear if physiological conditions that increase dNTP concentrations can affect pharmacokinetics (PK) and pharmacodynamics (PD) of NRTIs. This may be particularly relevant when cellular deoxynucleoside triphosphate (dNTP) pools in HIV target cells increase in response to immune activation. Decreased ratios of phosphorylated NRTIs to their respective dNTPs have been associated with cell activation in vitro and in nonhuman primate studies4,5, yet this observation remains unexplored in PK and PD studies that measured intracellular drug6-8. The study presented here compared dATP and dCTP concentrations to TFV-DP and FTC-TP in peripheral blood mononuclear cell (PBMCs) of persons using daily oral Truvada™ during a successful HIV PrEP study. We further examined if variations among intracellular drug:dNTP ratios were related to lymphocyte activation.

The female genital tract (FGT) provides a means of entry to pathogens, including HIV, yet immune cell populations at this barrier between host and environment are not well defined. We initiated a study of healthy women to characterize resident T cell populations in the lower FGT from lavage and patient-matched peripheral blood to investigate potential mechanisms of HIV sexual transmission. Surprisingly, we observed FGT CD4 T cell populations were primarily CCR7hi, consistent with a central memory or recirculating memory T cell phenotype. In addition, roughly half of these CCR7hi CD4 T cells expressed CD69, consistent with resident memory T cells, whereas the remaining CCR7hi CD4 T cells lacked CD69 expression, consistent with recirculating memory CD4 T cells that traffic between peripheral tissues and lymphoid sites. HIV susceptibility markers CCR5 and CD38 were increased on FGT CCR7hi CD4 T cells compared with blood, yet migration to the lymphoid homing chemokines CCL19 and CCL21 was maintained. Infection with GFP-HIV showed that FGT CCR7hi memory CD4 T cells are susceptible HIV targets, and productive infection of CCR7hi memory T cells did not alter chemotaxis to CCL19 and CCL21. Variations of resident CCR7hi FGT CD4 T cell populations were detected during the luteal phase of the menstrual cycle, and longitudinal analysis showed the frequency of this population positively correlated to progesterone levels. These data provide evidence women may acquire HIV through local infection of migratory CCR7hi CD4 T cells, and progesterone levels predict opportunities for HIV to access these novel target cells.

The genetic diversity among human immunodeficiency virus (HIV) subtypes as well as the variability of viral sequences found in HIV-infected individuals presents a number of difficult obstacles for the development of universally effective HIV treatment and prevention methods. Here, we present a brief summary of recent developments in the analysis of viral genetics and human genomics to provide insight into future methods for HIV treatment and prevention. Recent studies have mined viral sequences found in newly infected individuals to identify common features of all transmitted viruses that could provide potential targets for HIV vaccine development. Analysis of human immunogenetics has identified specific alleles associated with reduced virus loads in HIV-infected individuals providing valuable information that may influence individual responses to treatment and prevention methods. Increased sensitivity of antiretroviral drug resistance testing has improved the detection of hidden drug resistant virus but also highlighted the potential for drug resistant viruses to reduce the effectiveness of clinical treatment regimens. The rapidly expanding amount of data generated by studies of viral genetics and human immunogenetics will provide valuable information to guide the design of new strategies to improve clinical treatment and enhance HIV vaccine development.

There is a need to improve in vitro testing to evaluate topical microbicide candidates that prevent acquisition of HIV. Many vaginal microbicides with different anti-HIV activities have undergone preclinical testing and a few of those have been selected for clinical safety and efficacy testing. Clinical efficacy trials of vaginal microbicide gels have yielded mixed results. Initial nonspecific entry inhibitors were shown to be ineffective in clinical efficacy trials,1–4 whereas more recent testing of microbicide gels containing the nucleoside reverse transcriptase inhibitor, tenofovir, has shown some level of protection in 1 of 2 clinical trials.5,6 Yet, nearly all preclinical testing outcomes predicted that products should significantly reduce sexual acquisition of HIV when used appropriately in a clinical trial. However, a recent report using ex vivo testing of the microbicide Pro2000 demonstrated that this product loses anti-HIV activity after vaginal application and sexual intercourse.7 Changes in anti-HIV activity over time after vaginal application have not been studied for most candidate vaginal microbicides. | We investigated the carrageenan-based vaginal gel Carraguard, an HIV entry inhibitor that failed in a clinical efficacy trial,1 for degradation and loss of anti-infective activity after vaginal application. Cervicovaginal lavages (CVLs) were collected from 16 HIV-infected Thai women participating in a randomized, controlled, 3-treatment (Carraguard, methylcellulose placebo, no product) crossover safety trial.8 Women applied each treatment daily for 7 days after menses. The order of the 3 treatments was randomized. CVLs (5 mL) were collected on the first clinic visit in each treatment cycle before gel application (T0), 15 minutes after gel application (T15min), and on day 7 clinic visit which was 8–24 hours after the final gel application (T8–24hr). Self-reported adherence was 98% overall, and participants reported that vaginal application of the gel was highly acceptable.9

The potent non-nucleoside reverse transcriptase inhibitor UC781 has been safety tested as a vaginal microbicide gel formulation for prevention of HIV-1 sexual transmission. To investigate whether UC781 retained anti-infective activity following exposure to the female genital tract, we conducted an ex vivo analysis of the UC781 levels and antiviral activity in cervicovaginal lavages (CVL) from 25 Thai women enrolled in a 14-day safety trial of twice-daily vaginal application of two concentrations of UC781 microbicide gel. CVL samples were collected from women in 0.1% (n=5), 0.25% (n=15) and placebo (n=5) gel arms following the first application of gel (T(15min)) and 8-24 hours after the final application (T(8-24hr)), and separated into cell-free (CVL-s) and pelletable fractions (CVL-p). As UC781 is highly hydrophobic, there were significantly higher levels of UC781 in the CVL-p samples compared to CVL-s for the UC781 gel arms. In T(8-24hr) CVL-p samples, 2/5 and 13/15 samples collected from the 0.1% and 0.25% UC781 gel arms, respectively, efficiently blocked infection with ≥4 log(10) TCID(50) of a CCR5-tropic CRF01_AE HIV-1 virus stock. Independent of arm, the 11 CVL-p samples with UC781 levels ≥5 mcg/CVL reduced infectious HIV ≥4 log(10) TCID(50). Our results suggest that the levels and anti-infective activity of UC781 gel formulations are likely to be associated with a cellular or pelletable component in CVL samples. Therefore, cellular and pelletable fractions should be assayed for drug levels and anti-infective activity in preclinical studies of candidate microbicides.

This study compared HIV-1 genotypes shed over time (≤ 3.5 years) in the vaginal secretions (VS) and blood plasma (BP) of 15 chronically infected women. Analysis of predicted coreceptor tropism (CCR5 = R5, CXCR4 = X4) for quasispecies shedding revealed three patterns: (1) viral quasispecies shed in both VS and BP were restricted to R5-tropism at all time points, (2) quasispecies shed in VS were restricted to R5-tropism at all time points but X4 quasispecies were identified in the BP at one or more time points, (3) quasispecies shed in matched VS and BP both contained X4-tropic viruses. Overall, the frequency of X4 quasispecies circulation in VS was 2-fold less than in BP and detection of X4 virus in VS was more likely to occur when X4 quasispecies comprised more than 50% of BP viruses (p = 0.01) and when declines in blood CD4+ lymphocyte levels were the greatest (p = 0.038). Additionally, the mean number of predicted N-glycosylation sites between matched VS and BP samples were strongly correlated (r = 0.86, p < 0.0001) with glycosylation densities in the following order (VS R5 = BP R5 > BP X4 > VS X4). The X4 glyscosylation densities may result from compartmentalization pressures in the female genital tract or the delayed appearance of these viruses in VS. Our results suggest that the presence of X4 virus in VS is associated with a threshold population of X4 quasispecies in BP which are increasing during the HIV-induced failure of the human immune system.

BACKGROUND: Among human immunodeficiency virus (HIV)-infected men who have sex with men (MSM) taking combination antiretroviral therapy (cART), the impact of rectal sexually transmitted infections (STIs) on rectal HIV-1 shedding is unknown. METHODS: Human immunodeficiency virus type 1 (HIV-1) RNA was quantified from rectal swabs collected for Neisseria gonorrhoeae (GC) and Chlamydia trachomatis (CT) screening of HIV-1-infected MSM. Correlations of STIs with rectal viral load were explored using multinomial regression modeling. HIV-1 coreceptor tropism was predicted from sequencing in a subset of men. RESULTS: Thirty-one (39%) of 80 men (59 prescribed combination antiretroviral therapy [cART]) had HIV detected in 38 (42%) of 91 rectal swabs. Rectal HIV detection was associated with plasma virus loads above 3.15 log(10) copies/mL (95% confidence limit [CL] 2.73, 3.55) and paired rectal viral loads and plasma viral loads were correlated (Kendall's tau [tau] 0.68, Spearman rho [P] = .77). Rectal STIs and abnormal anal cytology were not associated with rectal viral load. HIV coreceptor distribution was very similar between the plasma and rectum in 3 of 4 men. CONCLUSIONS: Plasma and rectal viral load were correlated, and rectal STIs did not increase the likelihood of detecting HIV in the rectal secretions in MSM, including those with low or undetectable plasma viral load. Suppressing plasma viral load is likely to reduce risk of HIV transmission to insertive partners.