Genomic characterization of Dengue virus circulation in Ethiopia.

In Ethiopia, dengue virus (DENV) infections have been reported in several regions, however, little is known about the circulating genetic diversity. Here, we conducted clinical surveillance for DENV during the 2023 nationwide outbreak and sequenced DENV whole genomes for the first time in Ethiopia. We enrolled patients at three sentinel hospital sites. Using RT-PCR, we screened serum samples for three arboviruses followed by serotyping and sequencing for DENV-positive samples (10.4% of samples). We detected two DENV serotypes (DENV1 and DENV3). Phylogenetic analysis identified one transmission cluster of DENV1 (genotype III major lineage A), and two clusters of DENV3 (genotype III major lineage B). The first showed close evolutionary relationship to the 2023 Italian outbreak and the second cluster to Indian isolates. Co-circulation of DENV1 and DENV3 in some regions of Ethiopia highlights the potential for severe dengue. Intensified surveillance and coordinated public health response are needed to address the threat of severe dengue outbreaks.

Understanding the Transmission Dynamics of the Chikungunya Virus in Africa.

The Chikungunya virus (CHIKV) poses a significant global public health concern, especially in Africa. Since its first isolation in Tanzania in 1953, CHIKV has caused recurrent outbreaks, challenging healthcare systems in low-resource settings. Recent outbreaks in Africa highlight the dynamic nature of CHIKV transmission and the challenges of underreporting and underdiagnosis. Here, we review the literature and analyse publicly available cases, outbreaks, and genomic data, providing insights into the epidemiology, genetic diversity, and transmission dynamics of CHIKV in Africa. Our analyses reveal the circulation of geographically distinct CHIKV genotypes, with certain regions experiencing a disproportionate burden of disease. Phylogenetic analysis of sporadic outbreaks in West Africa suggests repeated emergence of the virus through enzootic spillover, which is markedly different from inferred transmission dynamics in East Africa, where the virus is often introduced from Asian outbreaks, including the recent reintroduction of the Indian Ocean lineage from the Indian subcontinent to East Africa. Furthermore, there is limited evidence of viral movement between these two regions. Understanding the history and transmission dynamics of outbreaks is crucial for effective public health planning. Despite advances in surveillance and research, diagnostic and surveillance challenges persist. This review and secondary analysis highlight the importance of ongoing surveillance, research, and collaboration to mitigate the burden of CHIKV in Africa and improve public health outcomes.

SARS-CoV-2 Genomic Epidemiology Dashboards: A Review of Functionality and Technological Frameworks for the Public Health Response.

During the coronavirus disease 2019 (COVID-19) pandemic, the number and types of dashboards produced increased to convey complex information using digestible visualizations. The pandemic saw a notable increase in genomic surveillance data, which genomic epidemiology dashboards presented in an easily interpretable manner. These dashboards have the potential to increase the transparency between the scientists producing pathogen genomic data and policymakers, public health stakeholders, and the public. This scoping review discusses the data presented, functional and visual features, and the computational architecture of six publicly available SARS-CoV-2 genomic epidemiology dashboards. We found three main types of genomic epidemiology dashboards: phylogenetic, genomic surveillance, and mutational. We found that data were sourced from different databases, such as GISAID, GenBank, and specific country databases, and these dashboards were produced for specific geographic locations. The key performance indicators and visualization used were specific to the type of genomic epidemiology dashboard. The computational architecture of the dashboards was created according to the needs of the end user. The genomic surveillance of pathogens is set to become a more common tool used to track ongoing and future outbreaks, and genomic epidemiology dashboards are powerful and adaptable resources that can be used in the public health response.

Evaluating diagnostic accuracy of an RT-PCR test for the detection of SARS-CoV-2 in saliva.

BACKGROUND AND OBJECTIVE: Saliva has been proposed as a potential more convenient, cost-effective, and easier sample for diagnosing SARS-CoV-2 infections, but there is limited knowledge of the impact of saliva volumes and stages of infection on its sensitivity and specificity. METHODS: In this study, we assessed the performance of SARS-CoV-2 testing in 171 saliva samples from 52 mostly mildly symptomatic patients (aged 18 to 70 years) with a positive reference standard result at screening. The samples were collected at different volumes (50, 100, 300, and 500 µl of saliva) and at different stages of the disease (at enrollment, day 7, 14, and 28 post SARS-CoV-2 diagnosis). Imperfect nasopharyngeal (NP) swab nucleic acid amplification testing was used as a reference. We used a logistic regression with generalized estimating equations to estimate sensitivity, specificity, PPV, and NPV, accounting for the correlation between repeated observations. RESULTS: The sensitivity and specificity values were consistent across saliva volumes. The sensitivity of saliva samples ranged from 70.2% (95% CI, 49.3-85.0%) for 100 µl to 81.0% (95% CI, 51.9-94.4%) for 300 µl of saliva collected. The specificity values ranged between 75.8% (95% CI, 55.0-88.9%) for 50 µl and 78.8% (95% CI, 63.2-88.9%) for 100 µl saliva compared to NP swab samples. The overall percentage of positive results in NP swabs and saliva specimens remained comparable throughout the study visits. We observed no significant difference in cycle number values between saliva and NP swab specimens, irrespective of saliva volume tested. CONCLUSIONS: The saliva collection offers a promising approach for population-based testing.

HIV-1 pretreatment and acquired antiretroviral drug resistance before tenofovir/ /lamivudine /dolutegravir (TLD) roll-out in Mozambique.

BACKGROUND: The World Health Organization (WHO) recommends that HIV treatment scale-up is accompanied by a robust assessment of drug resistance emergence and transmission. The WHO HIV Drug Resistance (HIVDR) monitoring and surveillance strategy includes HIVDR testing in adults both initiating and receiving antiretroviral therapy (ART). Due to limited information about HIVDR in Mozambique, we conducted two nationally representative surveys of adults initiating and receiving first-line ART regimes to better inform the HIV program. METHODS: We carried out a cross-sectional study between March 2017 and December 2019. Adults (older than 15 years) living with HIV (PLHIV) initiating ART or receiving first-line ART for between 9-15 months at 25 health facilities across all eleven provinces in Mozambique were included. Genotypic HIVDR was assessed on dried blood spots (DBS) when viral loads were ≥ 1000 copies/ml. Genotypic resistance for non-nucleoside reverse transcriptase inhibitors (NNRTIs), nucleoside reverse transcriptase inhibitors (NRTIs), and protease inhibitors (PIs) was determined using the Stanford HIV database algorithm 9.5 and calibrated population resistance tool 8.1. RESULTS: Of 828 participants -enrolled, viral load (VL) testing was performed on 408 initiators and 409 ART experienced. Unsuppressed VL was found in 68.1% 419 initiators and 18.8% (77/409) of the ART experienced. Of the 278 initiators and 70 ART experienced who underwent sequencing, 51.7% (144/278) and 75.7% (53/70) were sequenced successfully. Among the new initiators, pretreatment drug resistance (PDR) for NNRTI and PI was found in 16.0% (23/144) and 1.4% (2/144) of the participants, respectively. Acquired drug resistance (ADR) was found in 56.5% (30/53) of the ART-experienced participants of whom 24.5% (13/53) were resistant to both NRTI and NNRTI. CONCLUSION: High rates of PDR and ADR for NNRTI and ADR for NRTI were observed in our study. These findings support the replacement of NNRTIs with dolutegravir (DTG) but high levels of NRTI resistance in highly treatment-experienced individuals still require attention when transitioning to new regimens. Moreover, the study underlines the need for routine VL testing and HIVDR surveillance to improve treatment management strategies.

Sustained human outbreak of a new MPXV clade I lineage in eastern Democratic Republic of the Congo.

Outbreaks of monkeypox (mpox) have historically resulted from zoonotic spillover of clade I monkeypox virus (MPXV) in Central Africa and clade II MPXV in West Africa. In 2022, subclade IIb caused a global epidemic linked to transmission through sexual contact. Here we describe the epidemiological and genomic features of an mpox outbreak in a mining region in eastern Democratic Republic of the Congo, caused by clade I MPXV. Surveillance data collected between September 2023 and January 2024 identified 241 suspected cases. Genomic analysis demonstrates a distinct clade I lineage divergent from previously circulating strains in the Democratic Republic of the Congo. Of the 108 polymerase chain reaction-confirmed mpox cases, the median age of individuals was 22 years, 51.9% were female and 29% were sex workers, suggesting a potential role for sexual transmission. The predominance of APOBEC3-type mutations and the estimated emergence time around mid-September 2023 imply recent sustained human-to-human transmission.

Performance of Softcup® menstrual cup and vulvovaginal swab samples for detection and quantification of genital cytokines.

Cytokines are important mediators of immunity in the female genital tract, and their levels may be associated with various reproductive health outcomes. However, the measurement of cytokines and chemokines in vaginal fluid samples may be influenced by a variety of factors, each with the potential to affect the sensitivity and accuracy of the assay, including the interpretation and comparison of data. We measured and compared cytokine milieu in samples collected via Softcup® menstrual cup versus vulvovaginal swabs. One hundred and eighty vulvovaginal swabs from CAPRISA 088 and 42 Softcup supernatants from CAPRISA 016 cohorts of pregnant women were used to measure the concentrations of 28 cytokines through multiplexing. Cytokines measured in this study were detectable in each of the methods however, SoftCup supernatants showed consistently, higher detectability, expression ratios, and mean concentration of cytokines than vulvovaginal swabs. While mean concentrations differed, the majority of cytokines correlated between SoftCup supernatants and vulvovaginal swabs. Additionally, there were no significant differences in a number of participants between the two sampling methods for the classification of genital inflammation. Our findings suggest that SoftCup supernatants and vulvovaginal swab samples are suitable for the collection of genital specimens to study biological markers of genital inflammatory response. However, the Softcup menstrual cup performs better for the detection and quantification of soluble biomarkers that are found in low concentrations in cervicovaginal fluid.

An Oxford Nanopore Technology-Based Hepatitis B Virus Sequencing Protocol Suitable For Genomic Surveillance Within Clinical Diagnostic Settings.

Chronic hepatitis B virus (HBV) infection remains a significant public health concern, particularly in Africa, where there is a substantial burden. HBV is an enveloped virus, with isolates being classified into ten phylogenetically distinct genotypes (A - J) determined based on full-genome sequence data or reverse hybridization-based diagnostic tests. In practice, limitations are noted in that diagnostic sequencing, generally using Sanger sequencing, tends to focus only on the S-gene, yielding little or no information on intra-patient HBV genetic diversity with very low-frequency variants and reverse hybridization detects only known genotype-specific mutations. To resolve these limitations, we developed an Oxford Nanopore Technology (ONT)-based HBV genotyping protocol suitable for clinical virology, yielding complete HBV genome sequences and extensive data on intra-patient HBV diversity. Specifically, the protocol involves tiling-based PCR amplification of HBV sequences, library preparation using the ONT Rapid Barcoding Kit, ONT GridION sequencing, genotyping using Genome Detective software, recombination analysis using jpHMM and RDP5 software, and drug resistance profiling using Geno2pheno software. We prove the utility of our protocol by efficiently generating and characterizing high-quality near full-length HBV genomes from 148 left-over diagnostic Hepatitis B patient samples obtained in the Western Cape province of South Africa, providing valuable insights into the genetic diversity and epidemiology of HBV in this region of the world.

Investing in Africa's scientific future.

Africa bears a disproportionate burden of infectious diseases, accounting for a substantial percentage of global cases. Malaria, HIV/AIDS, tuberculosis, cholera, Ebola, Lassa fever, and other tropical diseases, such as dengue and chikungunya, have had a profound impact on morbidity and mortality. Various factors contribute to the higher prevalence and incidence of infectious diseases in Africa, including socioeconomic challenges, limited access to health care, inadequate sanitation and hygiene infrastructure, climate-related factors, and endemicity of certain diseases in specific regions. A skilled workforce is crucial to addressing these challenges. Unfortunately, many countries in Africa often lack the required resources, and aspiring scientists frequently seek educational and career opportunities abroad, leading to a substantial loss of talent and expertise from the continent. This talent migration, referred to as "brain drain," exacerbates the existing training gaps and hampers the sustainability of research within Africa.

An emerging clade of Chikungunya West African genotype discovered in real-time during 2023 outbreak in Senegal.

Chikungunya (CHIKV) is a re-emerging endemic arbovirus in West Africa. Since July 2023, Senegal and Burkina Faso have been experiencing an ongoing outbreak, with over 300 confirmed cases detected so far in the regions of Kédougou and Tambacounda in Senegal, the largest recorded outbreak yet. CHIKV is typically maintained in a sylvatic cycle in Senegal but its evolution and factors contributing to re-emergence are so far unknown in West Africa, leaving a gap in understanding and responding to recurrent epidemics. We produced, in real-time, the first locally-generated and publicly available CHIKV whole genomes in West Africa, to characterize the genetic diversity of circulating strains, along with phylodynamic analysis to estimate time of emergence and population growth dynamics. A novel strain of the West African genotype, phylogenetically distinct from strains circulating in previous outbreaks, was identified. This suggests a likely new spillover from sylvatic cycles in rural Senegal and potential of seeding larger epidemics in urban settings in Senegal and elsewhere.

Insights into SARS-CoV-2 in Angola during the COVID-19 peak: Molecular epidemiology and genome surveillance.

Background: In Angola, COVID-19 cases have been reported in all provinces, resulting in >105,000 cases and >1900 deaths. However, no detailed genomic surveillance into the introduction and spread of the SARS-CoV-2 virus has been conducted in Angola. We aimed to investigate the emergence and epidemic progression during the peak of the COVID-19 pandemic in Angola. Methods: We generated 1210 whole-genome SARS-CoV-2 sequences, contributing West African data to the global context, that were phylogenetically compared against global strains. Virus movement events were inferred using ancestral state reconstruction. Results: The epidemic in Angola was marked by four distinct waves of infection, dominated by 12 virus lineages, including VOCs, VOIs, and the VUM C.16, which was unique to South-Western Africa and circulated for an extended period within the region. Virus exchanges occurred between Angola and its neighboring countries, and strong links with Brazil and Portugal reflected the historical and cultural ties shared between these countries. The first case likely originated from southern Africa. Conclusion: A lack of a robust genome surveillance network and strong dependence on out-of-country sequencing limit real-time data generation to achieve timely disease outbreak responses, which remains of the utmost importance to mitigate future disease outbreaks in Angola.

Dispersal patterns and influence of air travel during the global expansion of SARS-CoV-2 variants of concern.

The Alpha, Beta, and Gamma SARS-CoV-2 variants of concern (VOCs) co-circulated globally during 2020 and 2021, fueling waves of infections. They were displaced by Delta during a third wave worldwide in 2021, which, in turn, was displaced by Omicron in late 2021. In this study, we use phylogenetic and phylogeographic methods to reconstruct the dispersal patterns of VOCs worldwide. We find that source-sink dynamics varied substantially by VOC and identify countries that acted as global and regional hubs of dissemination. We demonstrate the declining role of presumed origin countries of VOCs in their global dispersal, estimating that India contributed <15% of Delta exports and South Africa <1%-2% of Omicron dispersal. We estimate that >80 countries had received introductions of Omicron within 100 days of its emergence, associated with accelerated passenger air travel and higher transmissibility. Our study highlights the rapid dispersal of highly transmissible variants, with implications for genomic surveillance along the hierarchical airline network.

Evaluation of miniaturized Illumina DNA preparation protocols for SARS-CoV-2 whole genome sequencing.

The global pandemic caused by SARS-CoV-2 has increased the demand for scalable sequencing and diagnostic methods, especially for genomic surveillance. Although next-generation sequencing has enabled large-scale genomic surveillance, the ability to sequence SARS-CoV-2 in some settings has been limited by the cost of sequencing kits and the time-consuming preparations of sequencing libraries. We compared the sequencing outcomes, cost and turn-around times obtained using the standard Illumina DNA Prep kit protocol to three modified protocols with fewer clean-up steps and different reagent volumes (full volume, half volume, one-tenth volume). We processed a single run of 47 samples under each protocol and compared the yield and mean sequence coverage. The sequencing success rate and quality for the four different reactions were as follows: the full reaction was 98.2%, the one-tenth reaction was 98.0%, the full rapid reaction was 97.5% and the half-reaction, was 97.1%. As a result, uniformity of sequence quality indicated that libraries were not affected by the change in protocol. The cost of sequencing was reduced approximately seven-fold and the time taken to prepare the library was reduced from 6.5 hours to 3 hours. The sequencing results obtained using the miniaturised volumes showed comparability to the results obtained using full volumes as described by the manufacturer. The adaptation of the protocol represents a lower-cost, streamlined approach for SARS-CoV-2 sequencing, which can be used to produce genomic data quickly and more affordably, especially in resource-constrained settings.

Molecular Epidemiology and Diversity of SARS-CoV-2 in Ethiopia, 2020-2022.

Ethiopia is the second most populous country in Africa and the sixth most affected by COVID-19 on the continent. Despite having experienced five infection waves, >499,000 cases, and ~7500 COVID-19-related deaths as of January 2023, there is still no detailed genomic epidemiological report on the introduction and spread of SARS-CoV-2 in Ethiopia. In this study, we reconstructed and elucidated the COVID-19 epidemic dynamics. Specifically, we investigated the introduction, local transmission, ongoing evolution, and spread of SARS-CoV-2 during the first four infection waves using 353 high-quality near-whole genomes sampled in Ethiopia. Our results show that whereas viral introductions seeded the first wave, subsequent waves were seeded by local transmission. The B.1.480 lineage emerged in the first wave and notably remained in circulation even after the emergence of the Alpha variant. The B.1.480 was outcompeted by the Delta variant. Notably, Ethiopia's lack of local sequencing capacity was further limited by sporadic, uneven, and insufficient sampling that limited the incorporation of genomic epidemiology in the epidemic public health response in Ethiopia. These results highlight Ethiopia's role in SARS-CoV-2 dissemination and the urgent need for balanced, near-real-time genomic sequencing.

Safety and pharmacokinetics of escalating doses of neutralising monoclonal antibody CAP256V2LS administered with and without VRC07-523LS in HIV-negative women in South Africa (CAPRISA 012B): a phase 1, dose-escalation, randomised controlled trial.

BACKGROUND: Young women in sub-Saharan Africa continue to bear a high burden of HIV infection. Combination anti-HIV monoclonal antibodies are a potential HIV prevention technology that could overcome adherence challenges of daily oral pre-exposure prophylaxis. In this phase 1 clinical trial we aimed to determine the safety and pharmacokinetic profile of the broadly neutralising monoclonal antibody CAP256V2LS. METHODS: CAPRISA 012B, a first-in-human dose-escalation phase 1 trial evaluated the safety, pharmacokinetics, and neutralisation activity of CAP256V2LS alone and in combination with VRC07-523LS in young HIV-negative women in Durban, South Africa. Groups 1 and 2 were open label with CAP256V2LS administered at 5 mg/kg and 10 mg/kg intravenously and 5 mg/kg, 10 mg/kg, and 20 mg/kg subcutaneously. In group 3, participants were randomly allocated to receive a combination of CAP256V2LS and VRC07-523LS at 10 mg/kg and 20 mg/kg subcutaneously comixed with ENHANZE, a recombinant human hyaluronidase. Once safety was established in the first three participants, dose escalation took place sequentially following review of safety data. Primary endpoints were the proportion of participants with mild, moderate, and severe reactogenicity or adverse events, graded as per the Division of AIDS toxicity grading. The trial is registered on the Pan African Clinical Trial Registry, PACTR202003767867253, and is recruiting. FINDINGS: From July 13, 2020, to Jan 13, 2021, 42 HIV-negative women, aged 18-45 years, were enrolled. All 42 participants, eight with intravenous and 34 with subcutaneous administration, completed the trial. There were no serious adverse events or dose-limiting toxicities. Most commonly reported symptoms following intravenous administration were headaches in seven (88%) and nausea in four (50%) participants. Commonly reported symptoms following subcutaneous administration were headache in 31 (91%), chills in 25 (74%), and malaise or fatigue in 19 (56%) participants. Adverse events included transient lymphocytopenia in eight (19%), proteinuria in nine (21%), elevated aspartate aminotransferase in ten (24%), and alanine aminotransferase in five (12%) participants. INTERPRETATION: CAP256V2LS administered alone and in combination with VRC07-523LS was safe with favourable pharmacokinetics and neutralisation activity, supporting further assessment in larger clinical studies. FUNDING: European and Developing Countries Clinical Trials Partnership, South African Medical Research Council, and South African Department of Science and Innovation.

Global Expansion of SARS-CoV-2 Variants of Concern: Dispersal Patterns and Influence of Air Travel.

In many regions of the world, the Alpha, Beta and Gamma SARS-CoV-2 Variants of Concern (VOCs) co-circulated during 2020-21 and fueled waves of infections. During 2021, these variants were almost completely displaced by the Delta variant, causing a third wave of infections worldwide. This phenomenon of global viral lineage displacement was observed again in late 2021, when the Omicron variant disseminated globally. In this study, we use phylogenetic and phylogeographic methods to reconstruct the dispersal patterns of SARS-CoV-2 VOCs worldwide. We find that the source-sink dynamics of SARS-CoV-2 varied substantially by VOC, and identify countries that acted as global hubs of variant dissemination, while other countries became regional contributors to the export of specific variants. We demonstrate a declining role of presumed origin countries of VOCs to their global dispersal: we estimate that India contributed <15% of all global exports of Delta to other countries and South Africa <1-2% of all global Omicron exports globally. We further estimate that >80 countries had received introductions of Omicron BA.1 100 days after its inferred date of emergence, compared to just over 25 countries for the Alpha variant. This increased speed of global dissemination was associated with a rebound in air travel volume prior to Omicron emergence in addition to the higher transmissibility of Omicron relative to Alpha. Our study highlights the importance of global and regional hubs in VOC dispersal, and the speed at which highly transmissible variants disseminate through these hubs, even before their detection and characterization through genomic surveillance. Highlights: Global phylogenetic analysis reveals relationship between air travel and speed of dispersal of SARS-CoV-2 variants of concern (VOCs)Omicron VOC spread to 5x more countries within 100 days of its emergence compared to all other VOCsOnward transmission and dissemination of VOCs Delta and Omicron was primarily from secondary hubs rather than initial country of detection during a time of increased global air travelAnalysis highlights highly connected countries identified as major global and regional exporters of VOCs.

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance.

Investment in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing in Africa over the past year has led to a major increase in the number of sequences that have been generated and used to track the pandemic on the continent, a number that now exceeds 100,000 genomes. Our results show an increase in the number of African countries that are able to sequence domestically and highlight that local sequencing enables faster turnaround times and more-regular routine surveillance. Despite limitations of low testing proportions, findings from this genomic surveillance study underscore the heterogeneous nature of the pandemic and illuminate the distinct dispersal dynamics of variants of concern-particularly Alpha, Beta, Delta, and Omicron-on the continent. Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve while the continent faces many emerging and reemerging infectious disease threats. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century.