Re-emergence of dengue, chikungunya, and Zika viruses in 2021 after a 10-year gap in Gabon

Highlights • DENV-1, CHIKV, and ZIKV were detected in Gabon in 2021.• DENV appeared to switch to serotype 1 from serotype 2 and 3 since 2010.• Aedes albopictus‒adapted CHIKV appears to circulate repeatedly in Central Africa.• The recent Gabonese ZIKV strain was genetically different from the previous strain.


Introduction
Infections by mosquito-borne viruses, including dengue virus (DENV), chikungunya virus (CHIKV), and Zika virus (ZIKV), are caused primarily by the bite of Aedes mosquitoes infected with these viruses ( Gubler, 2001 ). Outbreaks of arboviral diseases have been consistently reported in Africa and the disease burden is increasing ( Wilder-Smith et al., 2017 ;Africa CDC, 2022 ). Recent examples were outbreaks of chikungunya in the Democratic Republic of the Congo in 2019 and dengue in Burkina Faso in 2017. Sporadic molecular and serological surveys have been conducted on DENV, CHIKV, and ZIKV in Gabon since the first documentation of the outbreaks or occurrence of infectious diseases caused by DENV-2, CHIKV, andZIKV in 2007 ( Leroy et al., 2009 ;Grard et al., 2014 ;Abe et al., 2020 ;Lim et al., 2021 ;Ushijima et al., 2021 ). However, little information is available on the genetic diversity and spatiotemporal dynamics of these viruses. Therefore, we in-

Results and Discussion
Of the 1060 samples, two (SYMAV-H0408, two-year-old female in Lambaréné; SYMAV-H0983, 29-year-old female in Lambaréné), one (SYMV-H0915, 6-year-old male in Sindara), and one (SYMAV-H0931, 12-year-old male in Ndjolé) were positive for DENV-1, CHIKV, and ZIKV, respectively, as analyzed by RT-qPCR. All patients visited the hospital 2 -3 days after the onset of fever and presented with a body temperature of 38°C or higher, but it was difficult to diagnose them with specific clinical manifestations.
To investigate the phylogeny of the detected viruses, we determined the sequences of the DENV-1 envelope (E) gene from two samples, and the complete genome sequence was determined for SYMAV-H0983 with a Ct value of 16.7 (GenBank accession nos. LC707378 and LC707382 ). Phylogenetic analysis inferred that these two samples belonged to the African group of genotype V and were closely related to the Gabon/2012 strain ( Figure 1 A and Supplemental Figure S1). The phylogenetic tree also showed that the clade of the Gabonese strains was separated from that of the Angola/2013 strain, suggesting that DENV-1 has circulated persistently in Gabon since its last appearance. Considering previous reports from 2015 -2017 showing the detection of both DENV-2 and 3 in the same area ( Abe et al., 2020 ;Lim et al., 2021 ), the serotype has likely now switched to DENV-1. The emergence of severe dengue cases should be carefully monitored in the area because secondary infections by other serotypes increase the risk of developing severe dengue ( Vaughn et al., 2000 ).
A phylogenetic tree of the CHIKV envelope 1 (E1) gene (GenBank accession no. LC707379 ) revealed that SYMAV-H0915 belonged to the East/Central/South African (ECSA) lineage and fell within the same group as the Cameroon/2018 and Gabon/2007 strains harboring E1-A226V, which confers a fitness advantage of CHIKV in Aedes albopictus ( Tsetsarkin et al., 2007 ) ( Figure 1 B). Bayesian phylogeny suggested that the recent Gabonese CHIKV strain was branched off from the strain detected in the Republic of Congo in 2011 and diverged earlier than the Cameroon isolates in 2016 and 2018 (Supplemental Figure S3). CHIKV appeared to circulate repeatedly in Cameroon, Gabon, and the Republic of the Congo. The CHIKV detected in the Republic of the Congo in 2019 exhibited the same E1-A226V mutation as the Gabonese strains, suggesting a vector-host switch event from Aedes aegypti to Aedes albopictus ( Fritz et al., 2019 ). Therefore, the spread of a new strain with A226V mutation could be a threat to Gabon and other Ae. albopictus -dominated regions ( Kraemer et al., 2019 ).
In Gabon, ZIKV was only reported in 2007, and its genome information was very limited, except for partial sequences of the E and nonstructural 3 (NS3) genes from one strain ( Grard et al., 2014 ). In this study, the sequences of the full-length NS3 gene and 1,065 bp E genes of SYMAV-H0931 were determined (GenBank accession nos. LC707380 and LC707381 ). A phylogenetic tree using the NS3 gene showed that the strain belonged to the African lineage and was located in the Central African clade, which contained old strains detected in the 1960 -1980s ( Figure 1 C), unlike the previous Gabonese strain belonging to the West African clade ( Figure 2 A and B). These results indicate that the recent ZIKV has either been newly introduced after 2007 or the traditional strain present in Central Africa since 1970-1980s has been circulating in Gabon for a long time without detection.
In conclusion, this study revealed ongoing DENV-1, CHIKV, and ZIKV circulation around Lambaréné and Gabon (Supplemental Figure S6). Considering that these viral infections are often subclinical, requiring no hospital visit, and that the preferred habitat (urban, rural, and so on) varies with the type of mosquito, active surveys based on communitybased samples and expanded study areas will help understand the entire country's situation.