Genomic evolution and rearrangement of CTX-Φ prophage elements in Vibrio cholerae during the 2018-2022 cholera outbreaks in The Democratic Republic of Congo

Between 2018 and 2022, we conducted whole-genome sequencing and phylogenomic analysis on 173 Vibrio cholerae bacteria (172 V. cholerae O1 and 1 V. cholerae non-O1/non-O139 isolates) from cholera patients across four provinces in the Democratic Republic of Congo (North-Kivu, South-Kivu, Tanganyika, and Kasai Oriental). All V. cholerae O1 isolates were classified into the AFR10d and AFR10e sublineages of AFR10 lineage, originating from the third wave of the seventh El Tor cholera pandemic (7PET). Compared to the strains analysed between 2014 and 2017, we observed only limited genetic changes in the core genome of both sublineages. However, AFR10e expanded across the four provinces, whereas AFR10d appeared to become extinct by the end of 2020. The V. cholerae isolates from 2022 exhibited significant rearrangement in the CTX prophage and associated phage satellites. Notably, this included the tandem repeat of a novel environmental satellite phage RS1 downstream the ctxB t oxin gene of the CTX-Φ-3 prophage on the large chromosome, and two tandemly arrayed copies of the pre-CTX-Φ prophage precursor on the small chromosome. In conclusion, while the core genome of V. cholerae O1 AFR10d and AFR10e sublineages showed minimal changes, significant alterations in the content and organisation of elements associated with the CTX-Φ and pre-CTX prophages were identified in AFR10e V. cholerae O1 isolated in 2022.