HIV Diagnostics and Vaccines: It Takes Two to Tango.

Current serologic tests for HIV screening and confirmation of infection present challenges to the adoption of HIV vaccines. The detection of vaccine-induced HIV-1 antibodies in the absence of HIV-1 infection, referred to as vaccine-induced seropositivity/seroreactivity, confounds the interpretation of test results, causing misclassification of HIV-1 status with potential affiliated stigmatization. For HIV vaccines to be widely adopted with high community confidence and uptake, tests are needed that are agnostic to the vaccination status of tested individuals (ie, positive only for true HIV-1 infection). Successful development and deployment of such tests will require HIV vaccine developers to work in concert with diagnostic developers. Such tests will need to match today's high-performance standards (accuracy, cost-effectiveness, simplicity) for use in vaccinated and unvaccinated populations, especially in low- and middle-income countries with high HIV burden. Herein, we discuss the challenges and strategies for developing modified serologic HIV tests for concurrent deployment with HIV vaccines.

Prioritizing interventions for preventing COVID-19 outbreaks in military basic training.

Like other congregate living settings, military basic training has been subject to outbreaks of COVID-19. We sought to identify improved strategies for preventing outbreaks in this setting using an agent-based model of a hypothetical cohort of trainees on a U.S. Army post. Our analysis revealed unique aspects of basic training that require customized approaches to outbreak prevention, which draws attention to the possibility that customized approaches may be necessary in other settings, too. In particular, we showed that introductions by trainers and support staff may be a major vulnerability, given that those individuals remain at risk of community exposure throughout the training period. We also found that increased testing of trainees upon arrival could actually increase the risk of outbreaks, given the potential for false-positive test results to lead to susceptible individuals becoming infected in group isolation and seeding outbreaks in training units upon release. Until an effective transmission-blocking vaccine is adopted at high coverage by individuals involved with basic training, need will persist for non-pharmaceutical interventions to prevent outbreaks in military basic training. Ongoing uncertainties about virus variants and breakthrough infections necessitate continued vigilance in this setting, even as vaccination coverage increases.

Transmission of SARS-CoV-2 among recruits in a US Army training environment: a brief report.

BACKGROUND: In 2020, preventive measures were implemented to mitigate the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) among 600-700 recruits arriving weekly at a basic combat training (BCT) facility in the southern United States. Trainees were sorted into companies and platoons (cocoons) at arrival, tested, quarantined for 14 days with daily temperature and respiratory-symptom monitoring and retested before release into larger groups for training where symptomatic testing was conducted. Nonpharmaceutical measures, such as masking, and social distancing, were maintained throughout quarantine and BCT. We assessed for SARS-CoV-2 transmission in the quarantine milieu. METHODS: Nasopharyngeal (NP) swabs were collected at arrival and at the end of quarantine and blood specimens at both timepoints and at the end of BCT. Epidemiological characteristics were analyzed for transmission clusters identified from whole-genome sequencing of NP samples. RESULTS: Among 1403 trainees enrolled from 25 August to 7 October 2020, epidemiological analysis identified three transmission clusters (n = 20 SARS-CoV-2 genomes) during quarantine, which spanned five different cocoons. However, SARS-CoV-2 incidence decreased from 2.7% during quarantine to 1.5% at the end of BCT; prevalence at arrival was 3.3%. CONCLUSIONS: These findings suggest layered SARS-CoV-2 mitigation measures implemented during quarantine minimized the risk of further transmission in BCT.

Virological and Serological Assessment of US Army Trainees Isolated for Coronavirus Disease 2019.

BACKGROUND: Laboratory screening for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a key mitigation measure to avoid the spread of infection among recruits starting basic combat training in a congregate setting. Because viral nucleic acid can be detected persistently after recovery, we evaluated other laboratory markers to distinguish recruits who could proceed with training from those who were infected. METHODS: Recruits isolated for coronavirus disease 2019 (COVID-19) were serially tested for SARS-CoV-2 subgenomic ribonucleic acid (sgRNA), and viral load (VL) by reverse-transcriptase polymerase chain reaction (RT-PCR), and for anti- SARS-CoV-2. Cluster and quadratic discriminant analyses of results were performed. RESULTS: Among 229 recruits isolated for COVID-19, those with a RT-PCR cycle threshold >30.49 (sensitivity 95%, specificity 96%) or having sgRNA log10 RNA copies/mL <3.09 (sensitivity and specificity 96%) at entry into isolation were likely SARS-CoV-2 uninfected. Viral load >4.58 log10 RNA copies/mL or anti-SARS-CoV-2 signal-to-cutoff ratio <1.38 (VL: sensitivity and specificity 93%; anti-SARS-CoV-2: sensitivity 83%, specificity 79%) had comparatively lower sensitivity and specificity when used alone for discrimination of infected from uninfected. CONCLUSIONS: Orthogonal laboratory assays used in combination with RT-PCR may have utility in determining SARS-CoV-2 infection status for decisions regarding isolation.

Pretreatment and Acquired Antiretroviral Drug Resistance Among Persons Living With HIV in Four African Countries.

BACKGROUND: Emerging HIV drug resistance (HIVDR) could jeopardize the success of standardized HIV management protocols in resource-limited settings. We characterized HIVDR among antiretroviral therapy (ART)-naive and experienced participants in the African Cohort Study (AFRICOS). METHODS: From January 2013 to April 2019, adults with HIV-1 RNA >1000 copies/mL underwent ART history review and HIVDR testing upon enrollment at 12 clinics in Uganda, Kenya, Tanzania, and Nigeria. We calculated resistance scores for specific drugs and tallied major mutations to non-nucleoside reverse transcriptase inhibitors (NNRTIs), nucleoside reverse transcriptase inhibitors (NRTIs), and protease inhibitors (PIs) using Stanford HIVDB 8.8 and SmartGene IDNS software. For ART-naive participants, World Health Organization surveillance drug resistance mutations (SDRMs) were noted. RESULTS: HIVDR testing was performed on 972 participants with median age 35.7 (interquartile range [IQR] 29.7-42.7) years and median CD4 295 (IQR 148-478) cells/mm3. Among 801 ART-naive participants, the prevalence of SDRMs was 11.0%, NNRTI mutations 8.2%, NRTI mutations 4.7%, and PI mutations 0.4%. Among 171 viremic ART-experienced participants, NNRTI mutation prevalence was 83.6%, NRTI 67.8%, and PI 1.8%. There were 90 ART-experienced participants with resistance to both efavirenz and lamivudine, 33 (36.7%) of whom were still prescribed these drugs. There were 10 with resistance to both tenofovir and lamivudine, 8 (80.0%) of whom were prescribed these drugs. CONCLUSIONS: Participants on failing ART regimens had a high burden of HIVDR that potentially limited the efficacy of standardized first- and second-line regimens. Management strategies that emphasize adherence counseling while delaying ART switch may promote drug resistance and should be reconsidered.

A Recombinant Vesicular Stomatitis Virus Ebola Vaccine.

BACKGROUND: The worst Ebola virus disease (EVD) outbreak in history has resulted in more than 28,000 cases and 11,000 deaths. We present the final results of two phase 1 trials of an attenuated, replication-competent, recombinant vesicular stomatitis virus (rVSV)-based vaccine candidate designed to prevent EVD. METHODS: We conducted two phase 1, placebo-controlled, double-blind, dose-escalation trials of an rVSV-based vaccine candidate expressing the glycoprotein of a Zaire strain of Ebola virus (ZEBOV). A total of 39 adults at each site (78 participants in all) were consecutively enrolled into groups of 13. At each site, volunteers received one of three doses of the rVSV-ZEBOV vaccine (3 million plaque-forming units [PFU], 20 million PFU, or 100 million PFU) or placebo. Volunteers at one of the sites received a second dose at day 28. Safety and immunogenicity were assessed. RESULTS: The most common adverse events were injection-site pain, fatigue, myalgia, and headache. Transient rVSV viremia was noted in all the vaccine recipients after dose 1. The rates of adverse events and viremia were lower after the second dose than after the first dose. By day 28, all the vaccine recipients had seroconversion as assessed by an enzyme-linked immunosorbent assay (ELISA) against the glycoprotein of the ZEBOV-Kikwit strain. At day 28, geometric mean titers of antibodies against ZEBOV glycoprotein were higher in the groups that received 20 million PFU or 100 million PFU than in the group that received 3 million PFU, as assessed by ELISA and by pseudovirion neutralization assay. A second dose at 28 days after dose 1 significantly increased antibody titers at day 56, but the effect was diminished at 6 months. CONCLUSIONS: This Ebola vaccine candidate elicited anti-Ebola antibody responses. After vaccination, rVSV viremia occurred frequently but was transient. These results support further evaluation of the vaccine dose of 20 million PFU for preexposure prophylaxis and suggest that a second dose may boost antibody responses. (Funded by the National Institutes of Health and others; rVSV∆G-ZEBOV-GP ClinicalTrials.gov numbers, NCT02269423 and NCT02280408 .).

Anorectal and Urogenital Mycoplasma genitalium in Nigerian Men Who Have Sex With Men and Transgender Women: Prevalence, Incidence, and Association With HIV.

Among 413 Nigerian men who have sex with men and transgender women, retrospective testing for Mycoplasma genitalium revealed mostly asymptomatic infections of the anorectum (prevalence, 36.8%; incidence, 18.4 cases/100 person-years) and urogenital tract (12.4%, 4.0 cases/100 person-years). Risk factors included HIV and increasing number of sex partners.

Decreased Seroreactivity in Individuals Initiating Antiretroviral Therapy during Acute HIV Infection.

Antiretroviral therapy (ART) during acute HIV infection (AHI) interrupts viral dynamics and may delay the emergence of serological markers targeted by current HIV screening and confirmatory assays, thus creating challenges for correctly classifying HIV infection status. The performance of three HIV antigen/antibody combination (HIV Ag/Ab Combo) assays (the Bio-Rad GS, Abbott Architect, and Bio-Rad BioPlex 2200 assays) was evaluated with samples collected from RV254/South East Asia Research Collaboration in HIV 010 (RV254/SEARCH010) study (Bangkok, Thailand) participants at weeks 12 and 24 following the initiation of ART at Fiebig stage I (FI) (n = 23), FII (n = 39), or FIII/IV (n = 22). Supplemental, confirmatory testing was performed by the Geenius HIV 1/2 and HIV-1 Western blot assays (Bio-Rad). Samples from 30 untreated, HIV-1-infected individuals demonstrated robust HIV Ag/Ab Combo assay reactivity with well-developed HIV-1 Western blotting profiles by 24 weeks after infection. In contrast, 52.2% of samples from individuals initiating ART at FI, 7.7% of samples from individuals initiating ART at FII, and 4.5% of samples from individuals initiating ART at FIII/IV were nonreactive by the HIV Ag/Ab Combo assays, with 36.4 to 39.1% of samples having low signal-to-cutoff (S/CO) results by the Architect and BioPlex assays (S/CO < 10). Seroreversion from a reactive to a nonreactive status was observed in 10 individuals initiating ART at FII and 3 individuals initiating ART at FIII/IV. The Geenius and HIV-1 Western blot assay results were negative or indeterminate for 73.9% and 69.6% of individuals, respectively, treated at FI; 50.0% and 26.3% of individuals, respectively, treated at FII; and 54.5% and 40.9% of individuals, respectively, treated at FIII/IV. Virologic suppression of HIV-1 by ART during AHI impedes seroconversion to biomarkers of infection, limiting the utility of HIV Ag/Ab Combo and supplemental, confirmatory assays for infection status determination.

Impact of Early Antiretroviral Therapy on Detection of Cell-Associated HIV-1 Nucleic Acid in Blood by the Roche Cobas TaqMan Test.

The Roche Cobas AmpliPrep/Cobas TaqMan HIV-1 test, v2.0 (the CAP/CTM assay), was used to quantify cell-associated HIV-1 (CAH) nucleic acid in peripheral blood mononuclear cells (PBMC) from well-characterized clinical specimens from HIV-1-infected individuals on antiretroviral therapy (ART). Chronically infected individuals on ART with no detectable plasma HIV-1 RNA demonstrated average CAH burdens of 3.2 HIV-1 log10 copies/million cells. Assay sensitivity and specificity were 98.9% and 100%, respectively, with the positive and negative predictive values being 100% and 98.6%, respectively. The CAH burden was also measured at weeks 0, 1, 2, 8, and 60 in 37 participants (RV254/SEARCH010, Bangkok, Thailand) stratified by Fiebig stage (Fiebig stage I [FI] to FVI) at ART initiation. Prior to ART initiation, the average CAH burden was 1.4, 4.1, and 3.6 log10 copies/million PBMCs for individuals who initiated ART at FI, FII, and FIII to FVI, respectively. Initiation of ART resulted in a rapid decline of CAH in all individuals, with the greatest decrease being observed in individuals who initiated ART at FI to FIII. By week 60, 100% (FI), 71.8% (FII/FIII), and 20.5% (FIV to FVI) of samples from individuals initiating treatment were at or near the limit of quantitation. Residual CAH was detectable at 60 weeks in most individuals who initiated ART at later stages (FIV to FVI) and averaged 1.9 ± 0.7 log10 copies/million PBMCs. The modified Roche CAP/CTM assay provides a convenient, standardized approach to measure residual HIV in blood and may be useful for monitoring patients under therapy or those participating in HIV remission studies.

Stability of Human Immunodeficiency Virus Serological Markers in Samples Collected as HemaSpot and Whatman 903 Dried Blood Spots.

Dried blood spots (DBS) are frequently used in clinical testing for biosurveillance, infectious disease and confirmatory testing, and clinical trials, particularly for populations in remote areas. The HemaSpot-HF blood collection device (HS) provides an alternative format to the Whatman 903 cards (903) to simplify sample collection and processing. In this study, the performance of the HS was compared to that of the 903 using previously characterized clinical specimens and HIV seroconversion panels known to exhibit markers of early human immunodeficiency virus (HIV) infection. HS and 903 samples were prepared and tested by Bio-Rad GS HIV Combo Ag/Ab enzyme immunoassay (EIA), GS HIV-1/-2 Plus O EIA, GS HIV-1 Western blot, and HIV-1 Geenius assays. Both HS and 903 performed well for up to 6 months at room temperature, but a marked loss of Western blot and low titer antibody signals from early infection samples was observed in samples stored for 180 days at elevated (37 to 45°C) temperatures and high humidity (95%). HemaSpot samples placed in sealed bags with additional desiccant were protected from degradation and showed improved signal recovery relative to that of the 903. HS was easier to use than the 903 and showed higher sensitivity and reproducibility for early infection samples and improved stability.