The COVID-19 pandemic, caused by SARS-CoV-2, has resulted in millions of infections and deaths globally. Genomic surveillance is crucial for understanding virus transmission and evolution, particularly with the emergence of Variants of Concern (VOCs) exhibiting increased transmissibility and reduced vaccine susceptibility. VOCs such as B.1.1.7 (UK), B.1.351 (South Africa), and P.1 (Brazil) have raised significant concerns. This study focuses on Uganda, where SARS-CoV-2 was first detected in March 2020. Initial infections were among international travelers, followed by transmission via truck drivers and community spread. By March 2021, Uganda had reported over 40,000 cases and 334 deaths. This research aims to investigate the emergence and spread of a new SARS-CoV-2 sublineage, A.23.1, which has gained dominance in the Ugandan epidemic. Understanding its genetic characteristics and potential impact on vaccine performance is paramount.

Previous studies have highlighted the importance of genomic surveillance in tracking the evolution and spread of SARS-CoV-2. The development of a detailed classification system (Pango lineages) has aided in monitoring the virus's evolution. Research has shown that several VOCs have emerged with mutations that increase transmissibility and decrease vaccine efficacy. Prior work in Uganda documented initial SARS-CoV-2 introductions and genomic diversity, showing a mixture of lineage A and B viruses initially, with later spread predominantly via land routes involving truck drivers. Studies in other countries have explored the evolution of specific lineages and their spread globally. This current work builds upon previous genomic surveillance efforts in Uganda and globally to characterize the newly identified A.23.1 lineage.

SARS-CoV-2 positive samples were collected from various clinical and surveillance locations across Uganda from March 2020 to January 2021. A total of 322 complete, high-coverage SARS-CoV-2 genomes were sequenced using the MinION platform. Phylogenetic analysis was performed using maximum-likelihood methods to determine the evolutionary relationships between the Ugandan sequences and those globally. Pango lineages were assigned using the pangolin module, and Nextstrain clades were determined using Nextclade. The spike protein sequences were analyzed to identify amino acid changes compared to the reference strain. To assess changes in other viral proteins, a profile Hidden Markov Model (pHMM) approach was used, comparing the Ugandan A.23.1 genomes to other VOCs (B.1.1.7, B.1.351, P.1). This involved translating open reading frames into amino acid peptides, clustering them, and then using HMMER to compare to a lineage B reference genome.

The study identified the emergence of SARS-CoV-2 lineage A.23.1 in Uganda. Lineage A.23.1 was first detected in late October 2020 and became the dominant lineage by December 2020-January 2021, accounting for 90% of the sequenced genomes. Lineage A.23.1 arose from lineage A.23. Initially observed in prison outbreaks, A.23 later spread across Uganda. A.23.1 exhibited several amino acid changes in the spike protein, including changes at positions 613 and 681 (similar to changes observed in VOCs). These changes may increase infectivity and furin cleavage. Additional changes were observed in the N-terminal domain, known to be a target of immune selection. Importantly, some A.23.1 sequences carried the E484K mutation in the receptor-binding domain, potentially impacting vaccine efficacy. Analysis of non-spike proteins (nsp6, ORF8, and ORF9) also revealed changes similar to those seen in VOCs. The A.23.1 lineage was also detected in 26 other countries, suggesting its potential for global spread.

The emergence and rapid spread of A.23.1 in Uganda mirrors the patterns observed with other VOCs, suggesting that local evolution and subsequent spread might be a common feature of SARS-CoV-2. The similarities in spike protein changes (particularly at positions 613 and 681 and the N-terminal domain), as well as changes in non-spike proteins (nsp6, ORF8, ORF9) between A.23.1 and VOCs warrant further investigation into potential functional consequences. The presence of the E484K mutation in some A.23.1 sequences is concerning due to its potential to compromise vaccine efficacy. The findings highlight the importance of ongoing genomic surveillance to detect and characterize emerging variants and assess their potential impact on public health strategies.

This study demonstrates the emergence and rapid spread of a novel SARS-CoV-2 lineage A.23.1 in Uganda with characteristics similar to other VOCs. The findings underscore the need for continuous genomic surveillance to monitor viral evolution and assess the potential for immune escape and reduced vaccine efficacy. Further research is needed to fully elucidate the clinical implications of A.23.1 and its response to current COVID-19 vaccines. This should include detailed epidemiological studies linking genetic changes to clinical outcomes and functional studies characterizing the effects of identified mutations on viral infectivity, immune evasion, and vaccine efficacy.

This study has limitations due to resource constraints in the study region. The number of sequenced genomes (322) represents a small fraction of the total number of cases in Uganda (40,000+). While the sampling covered multiple geographical locations across the country, limitations in testing capacity and surveillance might have resulted in an underrepresentation of certain lineages. While stringent quality control measures were implemented, the use of MinION sequencing technology has its inherent error profile, although it is now widely used for SARS-CoV-2 genome sequencing. The evolutionary pathway proposed may not reflect the full picture, as the data may not cover the full evolutionary landscape in the studied region. Alternative evolutionary pathways in other unsampled countries might have also contributed to the observed lineage spread.