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To prevent new infections with human immunodeficiency virus type 1 (HIV-1) in sub-Saharan Africa, UNAIDS recommends targeting interventions to populations that are at high risk of acquiring and passing on the virus. Yet it is often unclear who and where these 'source' populations are. Here we demonstrate how viral deep-sequencing can be used to reconstruct HIV-1 transmission networks and to infer the direction of transmission in these networks. We are able to deep-sequence virus from a large population-based sample of infected individuals in Rakai District, Uganda, reconstruct partial transmission networks, and infer the direction of transmission within them at an estimated error rate of 16.3% [8.8-28.3%]. With this error rate, deep-sequence phylogenetics cannot be used against individuals in legal contexts, but is sufficiently low for population-level inferences into the sources of epidemic spread. The technique presents new opportunities for characterizing source populations and for targeting of HIV-1 prevention interventions in Africa.

Phylogenetic analysis of pathogens is an increasingly powerful way to reduce the spread of epidemics, including HIV. As a result, phylogenetic approaches are becoming embedded in public health and research programmes, as well as outbreak responses, presenting unique ethical, legal, and social issues that are not adequately addressed by existing bioethics literature. We formed a multidisciplinary working group to explore the ethical issues arising from the design of, conduct in, and use of results from HIV phylogenetic studies, and to propose recommendations to minimise the associated risks to both individuals and groups. We identified eight key ethical domains, within which we highlighted factors that make HIV phylogenetic research unique. In this Review, we endeavoured to provide a framework to assist researchers, public health practitioners, and funding institutions to ensure that HIV phylogenetic studies are designed, done, and disseminated in an ethical manner. Our conclusions also have broader relevance for pathogen phylogenetics.

Background Limited viral load (VL) testing in human immunodeficiency virus (HIV) treatment programs in low-income countries often delays detection of treatment failure. The impact of remaining on failing protease inhibitor (PI)–containing regimens is unclear. Methods We retrospectively tested VL in 2164 stored plasma samples from 386 patients randomized to receive lopinavir monotherapy (after initial raltegravir induction) in the Europe–Africa Research Network for Evaluation of Second-line Therapy (EARNEST) trial. Protease genotypic resistance testing was performed when VL >1000 copies/mL. We assessed evolution of PI resistance mutations from virological failure (confirmed VL >1000 copies/mL) until PI monotherapy discontinuation and examined associations using mixed-effects models. Results Median post-failure follow-up (in 118 patients) was 68 (interquartile range, 48–88) weeks. At failure, 20% had intermediate/high-level resistance to lopinavir. At 40–48 weeks post-failure, 68% and 51% had intermediate/high-level resistance to lopinavir and atazanavir; 17% had intermediate-level resistance (none high) to darunavir. Common PI mutations were M46I, I54V, and V82A. On average, 1.7 (95% confidence interval 1.5–2.0) PI mutations developed per year; increasing after the first mutation; decreasing with subsequent mutations (P <.0001). VL changes were modest, mainly driven by nonadherence (P =.006) and PI mutation development (P =.0002); I47A was associated with a larger increase in VL than other mutations (P =.05). Conclusions Most patients develop intermediate/high-level lopinavir resistance within 1 year of ongoing viral replication on monotherapy but retain susceptibility to darunavir. Viral load increased slowly after failure, driven by non-adherence and PI mutation development. Clinical Trials Registration NCT00988039. [ABSTRACT FROM AUTHOR]

The rise in COVID-19 cases in late 2021 posed a grave threat to the public health system and the economy of Jamaica. A key pillar of controlling COVID-19 includes rapid diagnosis of SARS-CoV-2 infected individuals and their contacts. Hence, we evaluated the feasibility and acceptability of weekly deployment of antigen rapid diagnostic tests (Ag-RDTs) by conducting three 6-week studies within high-risk populations in Jamaica. We enrolled 1) 287 study participants (≥18 years) from low-income communities (Study A), 2) 262 healthcare workers (Study B), and 3) 88 students (14-17 years) (Study C). Conducting these independent studies was challenging. Willingness to participate was generally low with fear of phlebotomy (42%), discomfort associated with nasal swab (39%), and lack of parental consent (35%) being the most common reasons students gave (Study C) for lack of participation. Furthermore, only 57%, 66%, and 88% of participants concluded their final study visit in studies A to C, respectively. Participants' commitment and external factors, such as severe weather and outbreaks of violence affected follow-up. Overall, a total of six participants (<1%) tested COVID-19 Ag-RDT positive during all three studies, thus the number of infections detected were too low to draw any conclusions relating to the efficacy of the testing approach. Antibodies against SARS-CoV-2 were detected in most study participants (78-94%), but vaccination rates differed significantly between communities. Understanding these differences in vaccination rates is important because, given the low participation and follow-up rate, mass vaccination may present a more suitable public health intervention than regular testing.

The rapid diagnosis of coronavirus disease 2019 (COVID-19) is critical for comprehensive public health response strategies, and self-testing with antigen rapid diagnostic tests (Ag-RDTs) presents opportunities to test in hard-to-reach communities. Therefore, we evaluated the acceptability, feasibility, and uptake of Ag-RDT self-testing at the community level in Tanzania. From June to October 2022, symptomatic individuals or those with recent contact with a known or suspected COVID-19 patient were offered assisted testing and self-testing within mining communities and at transport hubs. This study included a cross-sectional survey before and after implementation. Participants were assessed for their acceptability and uptake of the nasal Ag-RDT self-test and their preference for nasal Ag-RDT self-testing. The survey data were collected in Open Data Kit, whereas the Ag-RDT results in the community were recorded by using the COVISUSPECT Mobile Application. Data analysis was performed by using STATA and R Statistical Software. A total of 538 individuals were screened, and 454 (84.4%) consented to be tested. The preference for self-testing was relatively low (33%), and the majority of participants (67%) opted to be assisted by a healthcare professional. Of the participants who opted for testing, 149 (32.8%) were able to self-test. Generally, there was no major difference in the various assessed parameters between the baseline and end-line surveys. The results from fitting multiple logistic regression indicated that after controlling for age, participants living in Dodoma were significantly less likely to opt for self-testing (odds ratio = 0.54; P-value = 0.023) compared with those living in Dar es Salaam. There was no significant difference in self-testing between participants living in Mara and those living in Dar es Salaam (odds ratio = 0.7; P-value = 0.179). After controlling for region, older (≥40 years) participants were significantly less likely to self-test compared with participants aged 18 to <40 years (odds ratio = 0.47; P-value = 0.002). The intervention was well-accepted in all areas in which Ag-RDTs were deployed. Our findings can therefore support the Ministry of Health by increasing accessibility to severe acute respiratory syndrome coronavirus 2 testing in the hard-to-reach communities in response to the next COVID-19 wave.

During the 33rd Africa Cup of Nations (AFCON) football tournament in Cameroon, organizers and health authorities required a negative SARS-CoV-2 test result <48 hours before entry and provided free SARS-CoV-2 testing and vaccination at stadium and fan zone entrances. We describe the outcomes and implementation of mandatory SARS-CoV-2 testing at fan zones during AFCON. All consenting fan zones attendees were administered an electronic questionnaire capturing exposure factors, COVID-19-like symptoms, and COVID-19 vaccination status, before being tested for SARS-CoV-2 using an antigen rapid diagnostic test (Ag-RDT). Participants testing positive were sampled for confirmatory real-time SARS-CoV-2 polymerase chain reaction (PCR) and sequencing for variant surveillance. The case detection rate was estimated using PCR-confirmed cases, and the challenges were summarized from staff discussions and project/study documentation. In total, 4,820 fan zone attendees (median [interquartile range] age 30 [24-38], 27.7% females) were tested for SARS-CoV-2, including 1,228 (25.5%) fully vaccinated. Of 4,820 participants, 148 (3.1%) had a positive Ag-RDT result, of whom 67 consented to PCR testing and 19 of 64 (29.7%) were confirmed PCR-positive. The case detection rate was 40.1 (95% CI: 24.2-62.7) per 10,000 attendees. The Omicron variant (B.1.1.529) was found in all 11 samples successfully sequenced. The implementation of mandatory SARS-CoV-2 Ag-RDT at fan zone entrances was challenged by high attendance volume just prior to matches, lobbying of economic stakeholders, and inconsistent quality assurance when using test kits. Despite the challenges encountered, implementing mandatory SARS-CoV-2 Ag-RDT at fan zones, was a unique opportunity for SARS-CoV-2 case identification and genomic surveillance.

Mass gathering event restrictions were part of mitigation measures during the COVID-19 pandemic that were lifted as prevalence decreased and after vaccination rollout. We explored SARS-CoV-2 antigen rapid diagnostic test acceptability and positivity in community settings in Cameroon. In August-October 2022, community workers sensitized and referred individuals for COVID-19 testing to nearby testing points in Douala and Yaoundé. Participants consented to SARS-CoV-2 antigen rapid diagnostic testing, a survey, or both components. We describe the positivity rate, COVID-19-related history, and Likert-scale testing perceptions. Factors associated with testing acceptance were analyzed using logistic regression. Overall, 20.5% (2,449/11,945) of sensitized individuals visited testing points, and 1,864 (76.1%) were enrolled; 50.6% accepted the survey and testing (46.0% accepted survey only). Seven (0.7%) of 1,006 individuals tested positive. Most (71.8%; 1,292/1,800) considered community testing more accessible than hospital-based testing. Individuals accepting versus refusing testing differed in perceived COVID-19 risk (67%, 49%; P <0.001), belief in accurate test results (79%, 47%; P <0.001), and ability to test easily (96%, 55%; P <0.001). Males (adjusted odds ratio [aOR]: 1.26 [1.04-1.53]) and those over 50 years (aOR: 1.9 [1.4-2.7]), with symptoms (aOR: 1.80 [1.30-2.50]), and at least partial vaccination (aOR: 0.76 [0.58-0.99]) were significantly associated with test acceptance. Refusal reasons included lack of perceived need for testing (33.8%) and testing discomfort (26.3%). Although community-based testing was generally perceived as important, actual testing uptake was low. In future pandemics, community testing should be optimized by addressing misinformation, offering alternative testing modalities for greater comfort, creating demand, and tailoring approaches to maximize testing uptake.

During the COVID-19 pandemic, the need for making testing readily available was recognized as an important factor for individuals to help make informed decisions, including to isolate or seek care, and for policymakers to control transmission. Toward this end, FIND and the Access to COVID-19 Tools Accelerator funded 16 rapid operational research studies and one implementation project in Africa, the Caribbean, and Asia evaluating the utility, acceptability, and feasibility of different community-based SARS-CoV-2 testing approaches. Here, we discuss common factors and challenges encountered during study implementation. We note six key factors essential for success: 1) collaboration and partnerships; 2) buy-in of local stakeholders, including communities; 3) access to affordable supplies; 4) flexible financing; 5) effective approval systems; and 6) a skilled and motivated workforce. We also note various challenges that must be addressed to fully capitalize on these success factors. In particular, ethics committees are often not well equipped to assess operational research during outbreaks. Outbreaks, especially of novel pathogens, are unpredictable, and transmission dynamics are even more likely to change if the pathogen is prone to frequent mutations, such as SARS-CoV-2. Research that aims to evaluate strategies for curbing transmission must hence be easily and swiftly adaptable. This requires flexibility from researchers, funders, staff conducting the studies, and ethics and other approval committees. International guidelines for evaluating operational research protocols in outbreaks are needed to provide timely evidence to enable informed decisions by individuals, communities, and policymakers, thereby reducing both the human and the economic impact of outbreaks.

The movement of people contributes to the spread of COVID-19 between communities. Hence, we evaluated the feasibility, acceptability, and impact of offering intercity bus travelers testing prior to their departure. We conducted baseline and endline surveys to map COVID-19 prevention practices in travel agencies in western Cameroon. As interventions, buses were randomly assigned to three study arms: 1) offering systematic COVID-19 rapid diagnostic testing (RDT) to all passengers (arm A); 2) offering testing to suspected cases (arm B); or 3) no testing (arm C). All travelers were called 7-10 days after their trip to identify potential cases. Fifty-five (90.2%) of the 61 travel agencies that were reached consented to participate in a baseline survey. Although only 27 (49.1%) of the agencies implemented at least one of the recommended COVID-19 preventive measures, 39 (70.9%) agreed to host a testing station. Six agencies were selected, and 669 buses were enrolled, including 223, 224, and 222 in arms A, B, and C, respectively. A total of 31,484 departing passengers were approached and 9,594 (30.5%) agreed to participate: 1,177 (12.3%) in arm A, 4,086 (42.6%) in arm B, and 4,331 (45.1%) in arm C. In all, 1,731 tests were performed, including 1,177 in arm A and 554 in arm B. Fourteen (0.8%) tests were positive, and two participants (14.3%) agreed to postpone their travel. Offering testing with antigen RDTs in travel agencies is feasible and acceptable. One-third of passengers consented, and testing did not delay any travels. Although this approach can detect COVID-19 cases, actions are needed to increase the proportion of positive cases postponing their travels.

Robust testing strategies are an essential aspect of COVID-19 pandemic preparedness and response. In 2022, most regions of Cameroon were still below the WHO's recommended level of 10 COVID-19 tests per 10,000 population. This study aimed to detect SARS-CoV-2 cases in university settings in Cameroon using antigen rapid diagnostic tests (Ag-RDTs) to increase national testing capacity and assess the knowledge, attitudes, and practices of this population regarding COVID-19 infection. Six universities in Buea, Douala, Dschang, Maroua, Ngaoundere, and Yaounde participated in this study from June to October 2022. Nasopharyngeal swabs were collected from participants and tested for COVID-19 using Ag-RDTs. For all positive cases, high-risk contacts were also tested by Ag-RDT. Participants were administered a structured questionnaire to assess their knowledge, attitudes, and practices regarding COVID-19. A total of 7,006 participants were recruited, and 54 (0.8%) were positive for SARS-CoV-2. Among close contacts, three of 62 (4.8%) tested positive. The University of Maroua was the only site to consistently report satisfactory testing capacity, achieving the study target of 30 tests/10,000 in 94.1% of cases. Participants' knowledge of COVID-19 was moderate to good (≥50%). However, 28% were unsure about the effectiveness of the COVID-19 vaccine. Two main factors were identified that could facilitate the spread of SARS-CoV-2 in university settings, namely the lack of restrictions on entering campus without a mask (36%) and the non-respect of social distancing on campus (42.7%). The results of this study will guide future policies to better control diseases with epidemic or pandemic potential by targeting educational institutions.

Background: Millions of HIV-infected people worldwide receive antiretroviral therapy (ART) in programmes using WHO-recommended standardised regimens. Recent WHO guidelines recommend a boosted protease inhibitor plus raltegravir as an alternative second-line combination. We assessed whether this treatment option offers any advantage over the standard protease inhibitor plus two nucleoside reverse-transcriptase inhibitors (NRTIs) second-line combination after 144 weeks of follow-up in typical programme settings., Methods: We analysed the 144-week outcomes at the completion of the EARNEST trial, a randomised controlled trial done in HIV-infected adults or adolescents in 14 sites in five sub-Saharan African countries (Uganda, Zimbabwe, Malawi, Kenya, Zambia). Participants were those who were no longer responding to non-NRTI-based first-line ART, as assessed with WHO criteria, confirmed by viral-load testing. Participants were randomly assigned to receive a ritonavir-boosted protease inhibitor (lopinavir 400 mg with ritonavir 100 mg, twice per day) plus two or three clinician-selected NRTIs (protease inhibitor plus NRTI group), protease inhibitor plus raltegravir (400 mg twice per day; protease inhibitor plus raltegravir group), or protease inhibitor monotherapy (plus raltegravir induction for first 12 weeks, re-intensified to combination therapy after week 96; protease inhibitor monotherapy group). Randomisation was by computer-generated randomisation sequence, with variable block size. The primary outcome was viral load of less than 400 copies per mL at week 144, for which we assessed non-inferiority with a one-sided α of 0·025, and superiority with a two-sided α of 0·025. The EARNEST trial is registered with ISRCTN, number 37737787., Findings: Between April 12, 2010, and April 29, 2011, 1837 patients were screened for eligibility, of whom 1277 patients were randomly assigned to an intervention group. In the primary (complete-case) analysis at 144 weeks, 317 (86%) of 367 in the protease inhibitor plus NRTI group had viral loads of less than 400 copies per mL compared with 312 (81%) of 383 in the protease inhibitor plus raltegravir group (p=0·07; lower 95% confidence limit for difference 10·2% vs specified non-inferiority margin 10%). In the protease inhibitor monotherapy group, 292 (78%) of 375 had viral loads of less than 400 copies per mL; p=0·003 versus the protease inhibitor plus NRTI group at 144 weeks. There was no difference between groups in serious adverse events, grade 3 or 4 adverse events (total or ART-related), or events that resulted in treatment modification., Interpretation: Protease inhibitor plus raltegravir offered no advantage over protease inhibitor plus NRTI in virological efficacy or safety. In the primary analysis, protease inhibitor plus raltegravir did not meet non-inferiority criteria. A regimen of protease inhibitor with NRTIs remains the best standardised second-line regimen for use in programmes in resource-limited settings., Funding: European and Developing Countries Clinical Trials Partnership (EDCTP), UK Medical Research Council, Instituto de Salud Carlos III, Irish Aid, Swedish International Development Cooperation Agency, Instituto Superiore di Sanita, Merck, ViiV Healthcare, WHO., (Copyright © 2018 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Background: Cross-resistance after first-line antiretroviral therapy (ART) failure is expected to impair activity of nucleoside reverse-transcriptase inhibitors (NRTIs) in second-line therapy for patients with HIV, but evidence for the effect of cross-resistance on virological outcomes is limited. We aimed to assess the association between the activity, predicted by resistance testing, of the NRTIs used in second-line therapy and treatment outcomes for patients infected with HIV., Methods: We did an observational analysis of additional data from a published open-label, randomised trial of second-line ART (EARNEST) in sub-Saharan Africa. 1277 adults or adolescents infected with HIV in whom first-line ART had failed (assessed by WHO criteria with virological confirmation) were randomly assigned to a boosted protease inhibitor (standardised to ritonavir-boosted lopinavir) with two to three NRTIs (clinician-selected, without resistance testing); or with raltegravir; or alone as protease inhibitor monotherapy (discontinued after week 96). We tested genotypic resistance on stored baseline samples in patients in the protease inhibitor and NRTI group and calculated the predicted activity of prescribed second-line NRTIs. We measured viral load in stored samples for all patients obtained every 12-16 weeks. This trial is registered with Controlled-Trials.com (number ISRCTN 37737787) and ClinicalTrials.gov (number NCT00988039)., Findings: Baseline genotypes were available in 391 (92%) of 426 patients in the protease inhibitor and NRTI group. 176 (89%) of 198 patients prescribed a protease inhibitor with no predicted-active NRTIs had viral suppression (viral load <400 copies per mL) at week 144, compared with 312 (81%) of 383 patients in the protease inhibitor and raltegravir group at week 144 (p=0·02) and 233 (61%) of 280 patients in the protease inhibitor monotherapy group at week 96 (p<0·0001). Compared with results with no active NRTIs, 95 (85%) of 112 patients with one predicted-active NRTI had viral suppression (p=0·3) and 20 (77%) of 26 patients with two or three active NRTIs had viral suppression (p=0·08). Over all follow-up, greater predicted NRTI activity was associated with worse viral load suppression (global p=0·0004)., Interpretation: Genotypic resistance testing might not accurately predict NRTI activity in protease inhibitor-based second-line ART. Our results do not support the introduction of routine resistance testing in ART programmes in low-income settings for the purpose of selecting second-line NRTIs., Funding: European and Developing Countries Clinical Trials Partnership, UK Medical Research Council, Institito de Salud Carlos III, Irish Aid, Swedish International Development Cooperation Agency, Instituto Superiore di Sanita, WHO, Merck., (Copyright © 2017 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Objective: To determine drug resistance mutation (DRM) patterns in a large cohort of patients failing nonnucleoside reverse transcriptase inhibitor (NNRTI)-based first-line antiretroviral therapy regimens in programs without routine viral load (VL) monitoring and to examine intersubtype differences in DRMs., Design: Sequences from 787 adults/adolescents who failed an NNRTI-based first-line regimen in 13 clinics in Uganda, Kenya, Zimbabwe, and Malawi were analyzed. Multivariable logistic regression was used to determine the association between specific DRMs and Stanford intermediate-/high-level resistance and factors including REGA subtype, first-line antiretroviral therapy drugs, CD4, and VL at failure., Results: The median first-line treatment duration was 4 years (interquartile range 30-43 months); 42% of participants had VL ≥100,000 copies/mL and 63% participants had CD4 <100 cells/mm. Viral subtype distribution was A1 (40%; Uganda and Kenya), C (31%; Zimbabwe and Malawi), and D (25%; Uganda and Kenya), and recombinant/unclassified (5%). In general, DRMs were more common in subtype-C than in subtype-A and/or subtype-D (nucleoside reverse transcriptase inhibitor mutations K65R and Q151M; NNRTI mutations E138A, V106M, Y181C, K101E, and H221Y). The presence of tenofovir resistance was similar between subtypes [P (adjusted) = 0.32], but resistance to zidovudine, abacavir, etravirine, or rilpivirine was more common in subtype-C than in subtype-D/subtype-A [P (adjusted) < 0.02]., Conclusions: Non-B subtypes differ in DRMs at first-line failure, which impacts on residual nucleoside reverse transcriptase inhibitor and NNRTI susceptibility. In particular, higher rates of etravirine and rilpivirine resistance in subtype-C may limit their potential utility in salvage regimens.

Objective: To assess neurocognitive function at the first-line antiretroviral therapy failure and change on the second-line therapy., Design: Randomized controlled trial was conducted in 5 sub-Saharan African countries., Methods: Patients failing the first-line therapy according to WHO criteria after >12 months on non-nucleoside reverse transcriptase inhibitors-based regimens were randomized to the second-line therapy (open-label) with lopinavir/ritonavir (400 mg/100 mg twice daily) plus either 2-3 clinician-selected nucleoside reverse transcriptase inhibitors, raltegravir, or as monotherapy after 12-week induction with raltegravir. Neurocognitive function was tested at baseline, weeks 48 and 96 using color trails tests 1 and 2, and the Grooved Pegboard test. Test results were converted to an average of the 3 individual test z-scores., Results: A total of 1036 patients (90% of those >18 years enrolled at 13 evaluable sites) had valid baseline tests (58% women, median: 38 years, viral load: 65,000 copies per milliliter, CD4 count: 73 cells per cubic millimeter). Mean (SD) baseline z-score was -2.96 (1.74); lower baseline z-scores were independently associated with older age, lower body weight, higher viral load, lower hemoglobin, less education, fewer weekly working hours, previous central nervous system disease, and taking fluconazole (P < 0.05 in multivariable model). Z-score was increased by mean (SE) of +1.23 (0.04) after 96 weeks on the second-line therapy (P < 0.001; n = 915 evaluable), with no evidence of difference between the treatment arms (P = 0.35)., Conclusions: Patients in sub-Saharan Africa failing the first-line therapy had low neurocognitive function test scores, but performance improved on the second-line therapy. Regimens with more central nervous system-penetrating drugs did not enhance neurocognitive recovery indicating this need not be a primary consideration in choosing a second-line regimen.