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Objectives: We assessed HIV-1 drug resistance profiles among people living with HIV (PLWH) with detectable viral load (VL) and on dolutegravir-based antiretroviral therapy (ART) in Botswana., Methods: The study utilised available 100 residual HIV-1 VL samples from unique PLWH in Francistown who had viraemia at-least 6 months after initiating ART in Botswana's national ART program from November 2023 to January 2024. Viraemia was categorized as low-level viraemia (LLV) (VL: 200-999 copies/mL) or virologic failure (VF) (VL ≥1000 copies/mL). HIV-1 protease, reverse transcriptase and integrase genes were sequenced using an in-house next-generation sequencing Oxford nanopore technology. HIV-1 drug resistance mutations (DRMs) were identified using the HIVdb Program in the Stanford HIV drug resistance database and compared between VL groups., Results: Among 100 participants, 83.0% were on dolutegravir-based, 10.0% were on non-dolutegravir-based ART and 7.0% had unknown/undocumented ART regimens. Thirty (30%) participants had LLV and 70 (70%) had VF. Among 58 successfully sequenced, 32.8% [95% Confidence Interval (CI): 21.8-46.0] had DRMs to any drug class, 33.3% (4/12) in the LLV group and 32.6% (15/46) in the VF group. Among individuals on dolutegravir-based ART, the overall HIV DRMs were 34.8% (95% CI: 22.7-49.2). By VL groups, 40.0% (95% CI: 16.8-68.7) and 33.3% (95% CI: 20.2-50.0) had DRMs at LLV and VF, respectively., Conclusions: A high but similar prevalence of any DRMs was observed among individuals with LLV and those with VF on dolutegravir-based therapy. Monitoring DRMs in individuals with detectable VL is crucial for preserving dolutegravir-based ART., (© The Author(s) 2025. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy.)

Botswana, like the rest of the world, has been significantly impacted by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In December 2022, we detected a monophyletic cluster of genomes comprising a sublineage of the Omicron variant of concern (VOC) designated as B.1.1.529.5.3.1.1.1.1.1.1.74.1 (alias FN.1, clade 22E). These genomes were sourced from both epidemiologically linked and unlinked samples collected in three close locations within the district of Greater Gaborone. In this study, we assessed the worldwide prevalence of the FN.1 lineage, evaluated its mutational profile, and conducted a phylogeographic analysis to reveal its global dispersal dynamics. Among approximately 16 million publicly available SARS-CoV-2 sequences generated by 30 September 2023, only 87 were of the FN.1 lineage, including 22 from Botswana, 6 from South Africa, and 59 from the UK. The estimated time to the most recent common ancestor of the 87 FN.1 sequences was 22 October 2022 [95% highest posterior density: 2 September 2022-24 November 2022], with the earliest of the 22 Botswana sequences having been sampled on 7 December 2022. Discrete trait reconstruction of FN.1 identified Botswana as the most probable place of origin. The FN.1 lineage is derived from the BQ.1.1 lineage and carries two missense variants in the spike protein, S:K182E in NTD and S:T478R in RDB. Among the over 90 SARS-CoV-2 lineages circulating in Botswana between September 2020 and July 2023, FN.1 was most closely related to BQ.1.1.74 based on maximum likelihood phylogenetic inference, differing only by the S:K182E mutation found in FN.1. Given the early detection of numerous novel variants from Botswana and its neighbouring countries, our study underscores the necessity of continuous surveillance to monitor the emergence of potential VOCs, integrating molecular and spatial data to identify dissemination patterns enhancing preparedness efforts., Competing Interests: None declared., (© The Author(s) 2024. Published by Oxford University Press.)

Objectives: We characterized the molecular and mutational landscape of SARS-CoV-2 recombinant strains in Botswana., Methods: We performed genomic, phylogenetic, and immunoinformatic analyses of 5254 near-complete genomes from 2020 to 2023. We assessed the presence of mutations of interested (MutOI) that may be associated with immune escape in silico ., Results: We observed a few recombinant strains in Botswana, with the majority being descendants of Omicron (XBB*), except for XV and XM. Most recombinant sequences corresponded to transmission clusters. Most recombination events occurred within the receptor-binding domain (RDB) of the spike (S) protein. We identified 16 MutOI among different proteins, with the majority occurring at a very low global prevalence (<4.8 × 10 - ⁵). We also observed S:Q474K, a MutOI in the RBD, that was predicted to escape human leukocyte antigen class I-mediated immune responses. Molecular surveillance is vital to inform early detection and response to potential variants with heightened immune and vaccine breakthrough properties., Conclusions: These results underscore the need for continued molecular surveillance to map the evolutionary landscape of SARS-CoV-2., Competing Interests: The authors declare no conflicts of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results., (© 2024 The Author(s).)

We investigated intra-host genetic evolution using two SARS-CoV-2 isolates from a fully vaccinated (primary schedule x2 doses of AstraZeneca plus a booster of Pfizer), >70-year-old woman with a history of lymphoma and hypertension who presented a SARS-CoV-2 infection for 3 weeks prior to death due to COVID-19. Two full genome sequences were determined from samples taken 13 days apart with both belonging to Pango lineage FL.2: the first detection of this Omicron sub-variant in Botswana. FL.2 is a sub-lineage of XBB.1.9.1. The repertoire of mutations and minority variants in the Spike protein differed between the two time points. Notably, we also observed deletions within the ORF1a and Membrane proteins; both regions are associated with high T-cell epitope density. The internal milieu of immune-suppressed individuals may accelerate SARS-CoV-2 evolution; hence, close monitoring is warranted., Competing Interests: Declarations of Competing Interest The authors have no competing interests to declare., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Importance: Fostemsavir (FTR) is a newly licensed antiretroviral drug that has been shown to have activity against HIV-1. The mechanism of action of FTR is different from all currently available antiretrovirals (ARVs), and as such, it offers hope for HIV-1 suppression in those people with HIV (PWH) who harbor HIV-1 variants with drug resistance mutations to currently used ARVs. Using 6,030 HIV-1 sequences covering the HIV-1 envelope from PWH in Botswana who are antiretroviral therapy (ART) naïve as well as those who are failing ART, we explored the sequences for FTR resistance-associated polymorphisms. We found the prevalence of FTR polymorphisms to be similar in both ART-naïve and ART-experienced individuals with VF in this setting, with no prior FTR exposure. Further studies on the phenotypic impact of these polymorphisms are warranted to guide how to monitor for FTR resistance., Competing Interests: The authors declare no conflict of interest.

We sought to investigate whether SARS-CoV-2 was present, and to perform full-length genomic sequencing, in a 5-year-old male crossbreed dog from Gaborone, Botswana that presented overt clinical signs (flu-like symptoms, dry hacking cough and mild dyspnoea). It was only sampled a posteriori, because three adult owners were diagnosed with SARS-CoV-2 infection. Next-generation sequencing based on Oxford Nanopore Technology (ONT) was performed on amplicons that were generated using a reverse transcriptase real-time polymerase chain reaction (RT-qPCR) of confirmed positive SARS-CoV-2 nasopharyngeal and buccal swabs, as well as a bronchoalveolar lavage with mean real cycle threshold (qCt) value of 36 based on the Nucleocapsid (N) gene. Descriptive comparisons to known sequences in Botswana and internationally were made using mutation profiling analysis and phylogenetic inferences. Human samples were not available. A near-full length SARS-CoV-2 genome (∼90% coverage) was successfully genotyped and classified under clade 20 O and Pango-Lineage AY.43 (Pango v.4.0.6 PLEARN-v1.3; 2022-04-21), which is a sublineage of the Delta variant of concern (VOC) (formerly called B.1.617.2, first detected in India). We did not identify novel mutations that may be used to distinguish SARS-CoV-2 isolates from the dog and humans. In addition to Spike (S) region mutation profiling, we performed phylogenetic analysis including 30 Delta sequences publicly available reference also isolated from dogs. In addition, we performed another exploratory analysis to investigate the phylogenetic relatedness of sequence isolated from dog with those from humans in Botswana (n = 1303) as of 31 March 2022 and of same sublineage. Expectedly, the sequence formed a cluster with Delta sublineages - AY.43, AY.116 and B.1.617.2 - circulating in same time frame. This is the first documented report of human-associated SARS-CoV-2 infection in a dog in Botswana. Although the direction of transmission remains unknown, this study further affirms the need for monitoring pets during different COVID-19 waves for possible clinically relevant SARS-CoV-2 transmissions between species., (© 2023 The Authors. Veterinary Medicine and Science published by John Wiley & Sons Ltd.)