COVID-19 has been characterized by several epidemiological waves and the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants from the ancestral strain in Wuhan, China [1] . This virus poses one of the greatest threats to mankind, due to its ability to evolve in the population and rapidly progress through variants [2] [3] . Variants have been classified, according to their degree of importance, as variants of high consequence (HCV), variants of concern (VOC), variants of interest (VOI) or variants under surveillance (VUM) [4] [5] . VOCs have been identified as the driving force behind viral circulation and dispersal, with higher rates in Northern Europe, Central America and sub-Saharan Africa [6] [7] . The emergence of variants with high pathogenicity and transmissibility has modified approaches to preventive measures, travel restrictions and the adoption of quarantine measures [8] [9] . In Africa, the spread of SARS-CoV-2 has been gradual and varied from country to country [10] . In Central Africa, the first case was confirmed in Gabon in March 2020 and around Chad, the first case of SARS-CoV-2 (COVID-19) infection was recorded in Cameroon on March 6, 2020, and from May 2, 2020, a total of 2069, cases and 61 deaths were recorded [6] [11] . In Chad, the first case was confirmed on 19 March 2020, in N’Djamena on a passenger arriving from Douala in Cameroon [12] . From 2020 to 2021, three epidemic peaks were recorded, with 6183 cases in the 23 provinces, and 184 deaths. Cases were concentrated in the capital, with 4373 cases and 107 deaths [13] . Data on variants were not available due to the absence of trained laboratory staff, lack of infrastructure and sequencing equipment. In addition, strategies for shipping samples to specialized laboratories outside the country were not implemented. The present study was undertaken to contribute to the knowledge base on the SARS-CoV-2 epidemic, and to provide preliminary data on some of the variants that circulated from residual samples from N’Djamena, Chad.

Of the 4373 RT PCR-positive SARS-CoV-2, 100 samples were taken and sequenced. Seventy-seven (77) samples produced sequences and 23 were altered and could not be sequenced, giving a success rate of 73% and a failure rate of 23% ( Figure 1 ).

Our study had a reasonable sample size, with over 70% of samples sequenced. However, around 20% of the samples analyzed were failures. These failures could be linked to the fact that samples were not shipped within the two days initially planned for shipment from N’Djamena to Kinshasa but took 4 days to Addis Ababa. This long transit time led to a break in the cold chain, which would have led to the degradation of the nuclear materials, resulting in the deterioration of 23 samples.

The study revealed SARS CoV-2 variants circulating in N’Djamena, with higher proposals in males than females. The maximum number of people exposed were aged between 26 - 35 and 36 - 45 years, and particularly among people not vaccinated against COVID-19 and residents of N’Djamena. In view of these results, the trend in the sex ratio in favor of men has been reported in studies carried out in Iran and Algeria [17] . The age ranges observed in this study corroborate the results of Maiga in Mali in 2021 [18] . These results could be explained by the fact that they constitute the most active group of people, and therefore more exposed to the risk of contamination. Women are generally not very mobile in Chad, largely in the informal sector [19] . Sequences of SARS-CoV-2 variants were found in both vaccinated and unvaccinated samples. A similar situation was reported in a study carried out in Tunisia, which revealed fifteen cases of COVID-19 infection in healthcare personnel correctly vaccinated against SARS-CoV-2 who had experienced two episodes of infection [20] . These data confirm that vaccination cannot prevent reinfection, but it can mitigate its severity. Moreover, these variants can escape the immune system, reducing the efficacy of vaccines [21] . SARS-CoV-2 variants have been identified in travelers as well as in the community. This observation could be explained by an exogenous and endogenous origin of the variants [22] . However, once the pathogen has been introduced, community spread takes over and develops significantly [23] . Reports of variants of concern (VOCs) in various parts of the world, notably the UK, South Africa, and Brazil [23] [24] .

Viral sub-lineages from the ancestral Wuhan 19B strain to the variant of concern (Alpha, Delta, Omicron), the variant of interest (Eta) and the variant under surveillance (B.1.640) have been observed to circulate [25] . The presence of the Wuhan sequence and variants previously reported in other countries indicates a probable importation of the virus, because the first confirmed case of COVID-19 in N’Djamena was a passenger from Douala/Cameroon [10] . A similar result was obtained in Bamako, Mali [18] . In the same period, co-circulation of the Alpha and Eta variants was reported in Nigeria, a country bordering Chad [26] . In China, the wild Wuhan strain was sequenced as early as January 2020 [27] . Although samples had been collected, the first sequences of SARS-CoV-2 were not known until July 2021. Maiga’s work in Mali in 2021 [18] reported only the Delta variant, whereas in our case the Alpha, Delta and Omicron variants were identified in our sequences. In addition, the Wuhan strain was present throughout the study period. As of October 7, 2021, 51 AU (African Union) member states reported the presence of variants of concern (VOC) Alpha (45 member states), Beta (39 member states), Delta (40 member states) and/or Gamma (2 member states). In addition, the beta variant circulated in Cameroon. The absence of certain variants could also be explained by the country’s difficulties with diagnostic tools [28] .

Our study shows that the country experienced three epidemic peaks between 2020 and 2023, characterized by early circulation of SARS CoV-2 variants of the Wuhan ancestral strain viral sublineage, followed by the Alpha variant in the last quarter of 2020, Alpha associated with Eta and B.1.640 in the first half of 2021. The Delta variant associated with B.1.640 from July to November 2021. And the Omicron variant in December 2021 was associated with the Delta variant. This gradual appearance of variants can be explained by the plasticity of the virus genome and its ability to produce variants rapidly. Numerous countries also experienced peaks, with similar situations recorded between January and February, with an average of 2544 daily cases. Burkina Faso, Nigeria, and Sierra Leone saw their peak in January 2021, with 36.3%, 42.2% and 55.1% of cases respectively. Ghana, Benin, Guinea Bissau, and Senegal reached their peak in February 2021. These countries accounted for 1.8%, 15.4%, 0.5% and 8.7% respectively of the total number of COVID-19 cases recorded in this second wave [29] .

In this study, the ancestral Wuhan strain, Alpha, and Delta-Omicron variants were implicated in epidemiological peaks, with peaks in March 2020, January 2021, and November 2021, associated with deaths and high case-fatality rates (8.51%) during circulation of the Wuhan strain and a decrease to 1% with the other variants. This finding seems to differ from the literature, which attributes the high case fatality associated with the circulation of the SARS-CoV-2 variants of concern, notably the Alpha, Beta, Gamma, and Omicron variants, which are highly transmissible and have a higher disease severity than the wild strain [30] . This very high case-fatality rate at the start of the pandemic may be linked to inadequate disease management capacity at the beginning of the pandemic, and the lack of appropriate treatment/therapeutic protocols in Chad.

Our study highlighted the circulation of SARS-CoV-2 and its variants in N’Djamena, Chad, as elsewhere in the world. These variants were at the origin of the outbreaks. Lethality was higher at the peak of the first wave than at the peaks of the last two waves. The variants were detected in both travelers and residents, as well as in immunized and non-immunized individuals. In view of the study’s limitations, the results cannot be extrapolated to a national scale. It would be necessary to extend SARS-CoV-2 genomic surveillance to the entire country. Also, between the end of the study period on December 31, 2021, and the declaration of the end of the pandemic by the WHO in January 2023, the genomic profile of the virus is not known in Chad. Investigations in this direction could be envisaged.

We sincerely thank all the participants in this study, in particular the Head of the Directorate of Disease Control and Epidemiological Surveillance of the Ministry of Public Health of Chad. We would also like to thank the medical and laboratory staff who helped us carry out the investigations and diagnosis of SARS CoV-2: Adam Abba Alkhali, Houlkeurbe NgamDaita, Djamal Hachim, Mahamat Adoum, ZitaAleyo, Narbe Maxim, and Boubacar Hassan for their technical assistance and the technical staff of the Institute National de Recherche’s Biomedicals’ (INRB) in Kinshasa for the sequencing of SARS CoV-2.

All authors contributed to the concept of this study. H.M., F.H.Y. and A.M.M. designed the study and wrote the initial manuscript. H.M and FHY analyzed data. F.H.Y and A.M.M. revised the manuscript and approved the final version. All authors have read and agreed to the published version of the manuscript.