Genomic epidemiology of SARS-CoV-2 in Sudan: A retrospective analysis 2020-2022 (preprint)

SARS-CoV-2 was first detected in Sudan on 13th March 2020. Here, we describe the genomic epidemiology of SARS-CoV-2 in Sudan between May 2020 and April 2022 to understand the introduction and transmission of SARS-CoV-2 variants in the country. A total of 667 SARS-CoV-2 positive samples were successfully sequenced using the nCoV-19 Artic protocol on the Oxford Nanopore Technology ([≥]70% genome completeness). The genomes were compared with a select contemporaneous global dataset to determine genetic relatedness and estimate import/export events. The genomes were classified into 37 Pango lineages within the ancestral strain (107 isolates across 13 Pango lineages), Eta variant of interest (VOI) (78 isolates in 1 lineage), Alpha variant of concern (VOC) (10 isolates in 2 lineages), Beta VOC (26 isolates in 1 lineage), Delta VOC (171 isolates across 8 lineages) and Omicron VOC (242 isolates across 12 lineages). We estimated a total of 144 introductions of the observed variants from different countries across the globe. Multiple introductions of the Eta VOI, Beta VOC and Omicron VOC were observed in Sudan mainly from Europe and Africa. These findings suggest a need for continuous genomic surveillance of SARS-CoV-2 to monitor their introduction and spread consequently inform public health measures to combat SARS-CoV-2 transmission.

SARS-CoV-2 Omicron variant of concern in the Seychelles: Introduction and spread

Background: The emergence of the Omicron variant of concern in late 2021 led to a resurgence of SARS-CoV-2 infections globally. By September 2022, Seychelles had experienced two major surges of SARS-CoV-2 infections driven by the Omicron variant. Here, we examine the genomic epidemiology of Omicron in the Seychelles between November 2021 and September 2022. Methods: : We analysed 618 SARS-CoV-2 Omicron genomes identified in the Seychelles between November 2021 and September 2022 to infer virus introductions and local transmission patterns using phylogenetics and the ancestral state reconstruction approach. We then evaluated the impact of government coronavirus 2019 (COVID-19) countermeasures on the estimated number of viral introductions during the study period. Results: : The genomes classified into 43 distinct Pango lineages. The first surge in Omicron cases (beginning November 2021 and peaking in January 2022) was predominated by the BA.1.1 lineage (59%) co-circulating with 11 other Omicron lineages. In the second surge (between April and June 2022), four lineages (BA.2, BA.2.10, BA.2.65 and BA.2.9) co-circulated and these were swiftly replaced by BA.5 subvariants in July 2022, which remained predominant through to September 2022. In the latter period, sporadic detections of BA.5 subvariants BQ.1, BE and BF were observed. We estimated 109 independent Omicron importations into Seychelles over the 11-month period, most of which occurred between December 2021 and March 2022 when strict government restrictions (SI>50%) were still in force. The districts Anse Royale, and Baie St. Anne Praslin appeared to be the major dispersal points fuelling local transmission. Conclusions: : Our results suggest that the waves of Omicron infections in the Seychelles were driven by multiple lineages and multiple virus introductions. The introductions were followed by substantial local spread and successive lineage displacement that mirrored the global patterns.

Genomic epidemiology of SARS-CoV-2 within households in coastal Kenya: a case ascertained cohort study (preprint)

Background Analysis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomic sequence data from household infections should aid its detailed epidemiological understanding. Using viral genomic sequence data, we investigated household SARS-CoV-2 transmission and evolution in coastal Kenya households. Methods We conducted a case-ascertained cohort study between December 2020 and February 2022 whereby 573 members of 158 households were prospectively monitored for SARS-CoV-2 infection. Households were invited to participate if a member tested SARS-CoV-2 positive or was a contact of a confirmed case. Follow-up visits collected a nasopharyngeal/oropharyngeal (NP/OP) swab on days 1, 4 and 7 for RT-PCR diagnosis. If any of these were positive, further swabs were collected on days 10, 14, 21 and 28. Positive samples with an RT-PCR cycle threshold of <33.0 were subjected to whole genome sequencing followed by phylogenetic analysis. Ancestral state reconstruction was used to determine if multiple viruses had entered households. Results Of 2,091 NP/OP swabs that were collected, 375 (17.9%) tested SARS-CoV-2 positive. Viral genome sequences (>80% coverage) were obtained from 208 (55%) positive samples obtained from 61 study households. These genomes fell within 11 Pango lineages and four variants of concern (Alpha, Beta, Delta and Omicron). We estimated 163 putative transmission events involving members of the sequenced households, 40 (25%) of which were intra-household transmission events while 123 (75%) were infections that likely occurred outside the households. Multiple virus introductions (up-to-5) were observed in 28 (47%) households with the 1-month follow-up period. Conclusions We show that a considerable proportion of SARS-CoV-2 infections in coastal Kenya occurred outside the household setting. Multiple virus introductions frequently occurred into households within the same infection wave in contrast to observations from high income settings, where single introduction appears to be the norm. Our findings suggests that control of SARS-CoV-2 transmission by household member isolation may be impractical in this setting.

The genomic epidemiology of SARS-CoV-2 variants of concern in Kenya (preprint)

The emergence and establishment of SARS CoV 2 variants of concern presented a major global public health crisis across the world. There were six waves of SARS CoV 2 cases in Kenya that corresponded with the introduction and eventual dominance of the major SARS-COV-2 variants of concern, excepting the first 2 waves that were both wildtype virus. We estimate that more than 1000 SARS CoV 2 introductions occurred in the two-year epidemic period (March 2020 to September 2022) and a total of 930 introductions were associated with variants of concern namely Beta (n=78), Alpha(n=108), Delta(n=239) and Omicron (n=505). A total of 29 introductions were associated with A.23.1 variant that circulated in high frequencies in Uganda and Rwanda. The actual number of introductions is likely to be higher than these conservative estimates due to limited genomic sequencing. Our data suggested that cryptic transmission was usually underway prior to the first real-time identification of a new variant, and that multiple introductions were responsible. Following emergence of each VOC and subsequent introduction, transmission patterns were associated with hotspots of transmission in Coast, Nairobi and Western Kenya and follows established land and air transport corridors. Understanding the introduction and dispersal of major circulating variants and identifying the sources of new introductions is important to inform public health control strategies within Kenya and the larger East-African region. Border control and case finding reactive to new variants is unlikely to be a successful control strategy.

Genomic Epidemiology of SARS-CoV-2 in Seychelles, 2020-2021.

Seychelles, an archipelago of 155 islands in the Indian Ocean, had confirmed 24,788 cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by the 31st of December 2021. The first SARS-CoV-2 cases in Seychelles were reported on the 14th of March 2020, but cases remained low until January 2021, when a surge was observed. Here, we investigated the potential drivers of the surge by genomic analysis of 1056 SARS-CoV-2 positive samples collected in Seychelles between 14 March 2020 and 31 December 2021. The Seychelles genomes were classified into 32 Pango lineages, 1042 of which fell within four variants of concern, i.e., Alpha, Beta, Delta and Omicron. Sporadic cases of SARS-CoV-2 detected in Seychelles in 2020 were mainly of lineage B.1 (lineage predominantly observed in Europe) but this lineage was rapidly replaced by Beta variant starting January 2021, and which was also subsequently replaced by the Delta variant in May 2021 that dominated till November 2021 when Omicron cases were identified. Using the ancestral state reconstruction approach, we estimated that at least 78 independent SARS-CoV-2 introduction events occurred in Seychelles during the study period. The majority of viral introductions into Seychelles occurred in 2021, despite substantial COVID-19 restrictions in place during this period. We conclude that the surge of SARS-CoV-2 cases in Seychelles in January 2021 was primarily due to the introduction of more transmissible SARS-CoV-2 variants into the islands.

Transmission networks of SARS-CoV-2 in Coastal Kenya during the first two waves: A retrospective genomic study.

Background: Detailed understanding of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) regional transmission networks within sub-Saharan Africa is key for guiding local public health interventions against the pandemic. Methods: Here, we analysed 1139 SARS-CoV-2 genomes from positive samples collected between March 2020 and February 2021 across six counties of Coastal Kenya (Mombasa, Kilifi, Taita Taveta, Kwale, Tana River, and Lamu) to infer virus introductions and local transmission patterns during the first two waves of infections. Virus importations were inferred using ancestral state reconstruction, and virus dispersal between counties was estimated using discrete phylogeographic analysis. Results: During Wave 1, 23 distinct Pango lineages were detected across the six counties, while during Wave 2, 29 lineages were detected; 9 of which occurred in both waves and 4 seemed to be Kenya specific (B.1.530, B.1.549, B.1.596.1, and N.8). Most of the sequenced infections belonged to lineage B.1 (n = 723, 63%), which predominated in both Wave 1 (73%, followed by lineages N.8 [6%] and B.1.1 [6%]) and Wave 2 (56%, followed by lineages B.1.549 [21%] and B.1.530 [5%]). Over the study period, we estimated 280 SARS-CoV-2 virus importations into Coastal Kenya. Mombasa City, a vital tourist and commercial centre for the region, was a major route for virus imports, most of which occurred during Wave 1, when many Coronavirus Disease 2019 (COVID-19) government restrictions were still in force. In Wave 2, inter-county transmission predominated, resulting in the emergence of local transmission chains and diversity. Conclusions: Our analysis supports moving COVID-19 control strategies in the region from a focus on international travel to strategies that will reduce local transmission. Funding: This work was funded by The Wellcome (grant numbers: 220985, 203077/Z/16/Z, 220977/Z/20/Z, and 222574/Z/21/Z) and the National Institute for Health and Care Research (NIHR), project references: 17/63/and 16/136/33 using UK Aid from the UK government to support global health research, The UK Foreign, Commonwealth and Development Office. The views expressed in this publication are those of the author(s) and not necessarily those of the funding agencies.

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

Investment in Africa over the past year with regards to SARS-CoV-2 genotyping has led to a massive increase in the number of sequences, exceeding 100,000 genomes generated to track the pandemic on the continent. Our results show an increase in the number of African countries able to sequence within their own borders, coupled with a decrease in sequencing turnaround time. Findings from this genomic surveillance underscores the heterogeneous nature of the pandemic but we observe repeated dissemination of SARS-CoV-2 variants within the continent. Sustained investment for genomic surveillance in Africa is needed as the virus continues to evolve, particularly in the low vaccination landscape. These investments are very crucial for preparedness and response for future pathogen outbreaks.

An optimization of four SARS-CoV-2 qRT-PCR assays in a Kenyan laboratory to support the national COVID-19 rapid response teams.

Background: The COVID-19 pandemic relies on real-time polymerase chain reaction (qRT-PCR) for the detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), to facilitate roll-out of patient care and infection control measures. There are several qRT-PCR assays with little evidence on their comparability. We report alterations to the developers' recommendations to sustain the testing capability in a resource-limited setting. Methods: We used a SARS-CoV-2 positive control RNA sample to generate several 10-fold dilution series that were used for optimization and comparison of the performance of the four qRT-PCR assays: i) Charité Berlin primer-probe set, ii) European Virus Archive - GLOBAL (EVAg) primer-probe set, iii) DAAN premixed commercial kit and iv) Beijing Genomics Institute (BGI) premixed commercial kit. We adjusted the manufacturer- and protocol-recommended reaction component volumes for these assays and assessed the impact on cycle threshold (Ct) values. Results: The Berlin and EVAg E gene and RdRp assays reported mean Ct values within range of each other across the different titrations and with less than 5% difference. The DAAN premixed kit produced comparable Ct values across the titrations, while the BGI kit improved in performance following a reduction of the reaction components. Conclusion: We achieved a 2.6-fold and 4-fold increase in the number of tests per kit for the commercial kits and the primer-probe sets, respectively. All the assays had optimal performance when the primers and probes were used at 0.375X, except for the Berlin N gene assay. The DAAN kit was a reliable assay for primary screening of SARS-CoV-2 whereas the BGI kit's performance was dependent on the volumes and concentrations of both the reaction buffer and enzyme mix. Our recommendation for SARS-CoV-2 diagnostic testing in resource-limited settings is to optimize the assays available to establish the lowest volume and suitable concentration of reagents required to produce valid results.

An optimisation of four SARS-CoV-2 qRT-PCR assays in a Kenyan laboratory to support the national COVID-19 rapid response teams

Background: The global COVID-19 outbreak relies on a quantitative real-time polymerase chain reaction (qRT-PCR) for the detection of severe acute respiratory syndrome coronavirus (SARS-CoV-2), to facilitate the roll-out of patient care and infection control measures. There are several qRT-PCR assays with little evidence on their comparability. We report alterations to the developers' recommendations to sustain the testing capability in our setting, where the supply of testing reagents is limited. Methods: Standards generated from a serially-diluted positive control and previously identified positive/negative samples were used to determine the optimal volumes of the qRT-PCR reagents and to evaluate the validity and performance of four assays: Charité Berlin and European Virus Archive - GLOBAL (EVAg) primer-probe sets, and DAAN and Beijing Genomics Institute (BGI) premixed commercial kits. A multiplex and singleplex RT-PCR kit was used with the two primer-probe sets and the recommended assay volumes of the two premixed kits were altered. Results: In comparison to the multiplex RT-PCR kit, the singleplex RT-PCR kit combined with the primer-probe sets yielded consistent cycle threshold (Ct) values across the different titrations tested. The DAAN premixed kit produced comparable Ct values across the titrations, while the BGI kit showed incomparable Ct values and inconsistent results between batches using the manufacturer's recommended volumes. Conclusion: We achieved a 2.5-fold and 4-fold increase in the number of tests/kit for the premixed kits and the primer-probe sets, respectively. The primer-probe set assays were reliable and consistent, and we preferred a combination of an EVAg and a Berlin target. Any inconclusive result was repeated by different individuals following the same protocol. DAAN was a consistent and reliable assay even at lower concentrations from the stated recommendations. BGI in contrast, required dilution to improve its performance and was hence an assay that was used in combination with EVAg or Berlin targets.

Genomic Epidemiology of SARS-CoV-2 in Seychelles, 2020-2021 (preprint)

By 31st December 2021, Seychelles, an archipelago of 115 islands in the Indian Ocean, had confirmed 24,788 cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The first SARS-CoV-2 cases in Seychelles were reported on 14th March 2020, but cases remained low until January 2021, when a surge of SARS-CoV-2 cases was observed on the islands. Here, we investigated the potential drivers of the surge by genomic analysis 1,056 SARS-CoV-2 positive samples collected in Seychelles between 14th March 2020 and 31st December 2021. The Seychelles genomes were classified into 32 Pango lineages, 1,042 of which fell within four variants of concern i.e., Alpha, Beta, Delta and Omicron. Sporadic of SARS-CoV-2 detected in Seychelles in 2020 were mainly of lineage B.1 (Europe origin) but this lineage was rapidly replaced by Beta variant starting January 2021, and which was also subsequently replaced by the Delta variant in May 2021 that dominated till November 2021 when Omicron cases were identified. Using ancestral state reconstruction approach, we estimated at least 78 independent SARS-CoV-2 introduction events into Seychelles during the study period. Majority of viral introductions into Seychelles occurred in 2021, despite substantial COVID-19 restrictions in place during this period. We conclude that the surge of SARS-CoV-2 cases in Seychelles in January 2021 was primarily due to introduction of the more transmissible SARS-CoV-2 variants into the islands.

Tracking the introduction and spread of SARS-CoV-2 in coastal Kenya.

Genomic surveillance of SARS-CoV-2 is important for understanding both the evolution and the patterns of local and global transmission. Here, we generated 311 SARS-CoV-2 genomes from samples collected in coastal Kenya between 17th March and 31st July 2020. We estimated multiple independent SARS-CoV-2 introductions into the region were primarily of European origin, although introductions could have come through neighbouring countries. Lineage B.1 accounted for 74% of sequenced cases. Lineages A, B and B.4 were detected in screened individuals at the Kenya-Tanzania border or returning travellers. Though multiple lineages were introduced into coastal Kenya following the initial confirmed case, none showed extensive local expansion other than lineage B.1. International points of entry were important conduits of SARS-CoV-2 importations into coastal Kenya and early public health responses prevented established transmission of some lineages. Undetected introductions through points of entry including imports from elsewhere in the country gave rise to the local epidemic at the Kenyan coast.

Genomic surveillance reveals the spread patterns of SARS-CoV-2 in coastal Kenya during the first two waves (preprint)

The transmission networks of SARS-CoV-2 in sub-Saharan Africa remain poorly understood. We analyzed 684 genomes from samples collected across six counties in coastal Kenya during the first two waves (March 2020 - February 2021). Up to 32 Pango lineages were detected in the local sample with six accounting for 88.0% of the sequenced infections: B.1 (60.4%), B.1.1 (8.9%), B.1.549 (7.9%), B.1.530 (6.4%), N.8 (4.4%) and A (3.1%). In a contemporaneous global sample, 571 lineages were identified, 247 for Africa and 88 for East Africa. We detected 262 location transition events comprising: 64 viral imports into Coastal Kenya; 26 viral exports from coastal Kenya; and 172 inter-county import/export events. Most international viral imports (61%) and exports (88%) occurred through Mombasa, a key coastal touristic and commercial center; and many occurred prior to June 2020, when stringent local COVID-19 restriction measures were enforced. After this period, local transmission dominated, and distinct local phylogenies were seen. Our analysis supports moving control strategies from a focus on international travel to local transmission.

A year of genomic surveillance reveals how the SARS-CoV-2 pandemic unfolded in Africa (preprint)

The progression of the SARS-CoV-2 pandemic in Africa has so far been heterogeneous and the full impact is not yet well understood. Here, we describe the genomic epidemiology using a dataset of 8746 genomes from 33 African countries and two overseas territories. We show that the epidemics in most countries were initiated by importations, predominantly from Europe, which diminished following the early introduction of international travel restrictions. As the pandemic progressed, ongoing transmission in many countries and increasing mobility led to the emergence and spread within the continent of many variants of concern and interest, such as B.1.351, B.1.525, A.23.1 and C.1.1. Although distorted by low sampling numbers and blind-spots, the findings highlight that Africa must not be left behind in the global pandemic response, otherwise it could become a breeding ground for new variants.

Tracking the introduction and spread of SARS-CoV-2 in coastal Kenya (preprint)

We generated 274 SARS-CoV-2 genomes from samples collected during the early phase of the Kenyan pandemic. Phylogenetic analysis identified 8 global lineages and at least 76 independent SARS-CoV-2 introductions into Kenyan coast. The dominant B.1 lineage (European origin) accounted for 82.1% of the cases. Lineages A, B and B.4 were detected from screened individuals at the Kenya-Tanzania border or returning travellers but did not lead to established transmission. Though multiple lineages were introduced in coastal Kenya within three months following the initial confirmed case, none showed extensive local expansion other than cases characterised by lineage B.1, which accounted for 45 of the 76 introductions. We conclude that the international points of entry were important conduits of SARS-CoV-2 importations. We speculate that early public health responses prevented many introductions leading to established transmission, but nevertheless a few undetected introductions were sufficient to give rise to an established epidemic.