Cholera remains a serious public health problem in Uganda and Africa as a whole ^{1, 2} . It is characterized by a large disease burden, recurrent outbreaks, high case fatality rates, as well as tenacious endemicity ^{1, 2} . Over the last four decades, Uganda has experienced several cholera outbreaks ² . The detection, monitoring, and surveillance of cholera in Uganda rely upon the isolation of Vibrio cholerae using culture-based methods in microbiology laboratories ^{2, 3} . However, these methods are faced with several challenges, including: associated long turn-around times (24–48 hours); limited microbiology laboratory capacity; lack of laboratory supplies; poor laboratory infrastructure, particularly electricity necessary to operate laboratory equipment; as well as limited reliable and rapid diagnostic tests ² . Unlike culture-based methods, high-throughput sequencing, a culture-independent method, has been documented to be less affected by most of the challenges facing culture-based methods and at the same time provides an unprecedented view of pathogen biology and delivers high-resolution genomic epidemiology via rapid and cheap whole-genome sequencing ^{4– 7} . Despite this knowledge, sequencing remains a less desirable option for most scientists in Uganda and hence, data on the genomic characteristics of V. cholerae remains scarce, likely attributable to the underdeveloped bioinformatics capacity and lack of expertise necessary for analyzing whole-genome sequencing data ⁷ . This study set out to use bioinformatics approaches to analyze whole-genome sequence data obtained from V. cholerae isolates from different outbreaks in Uganda, with the aim of providing the complete array of virulence genes, pathogenicity islands, antimicrobial resistance genes, integrative and conjugative elements, and antimicrobial resistance genes associated with these elements, plasmids, and insertion sequences. In addition, this study also provided a single nucleotide polymorphism (SNP) based phylogenetic analysis of the strains.

This was a cross-sectional study that aimed at providing a comprehensive genomic analysis of 10 whole-genome sequences of V. cholerae isolates collected during three different cholera outbreaks in Uganda between 2014 and 2016, submitted by the University of Maryland (Baltimore, MD, United States) to the NCBI SRA database under a study titled, “Molecular characterization of Vibrio cholerae responsible for cholera epidemics in Uganda by PCR, MLVA and WGS”.

In the genomic analysis, this study confirmed the identity of the isolates, provided the complete array of virulence genes, pathogenicity islands, antimicrobial resistance genes, integrative and conjugative elements, and antimicrobial resistance genes associated with these elements, plasmids, and insertion sequences. In addition, this study also provided a SNP-based phylogenetic analysis of the strains.

The identity of the isolates was in tandem with what was reported by Bwire et al., 2018. In addition, the finding of this study in regards to most of the isolates belonging to the O1 serotype are consistent with other studies elsewhere; these studies attributed this to the presence of the rfbV-O1 gene in isolates classified as O1 ⁵ . Unlike a similar study done in the East African region ⁵ in which MLST revealed that their isolates belonged to a single ST (ST69), this study revealed that the isolates belonged to two STs; namely, ST69 and ST515. Strains of V. cholerae belonging to the ST515 have been reported elsewhere ¹¹ .

The virulence genes reported in this study are similar to those reported in studies elsewhere ^{5, 12– 14} . These genes included, among others, those belonging to the Type IV secretion system, those involved in adherence, Type IV pilus genes, and those involved in quorum sensing.

Accessory genetic elements, particularly pathogenicity islands previously reported to commonly occur in V. cholerae were also reported in this study; namely, VSP-1, VSP-2 as well as VPI-1 and VPI-2 ^{15, 16} . These have not only been documented to encode virulence-associated genes in V. cholerae, but have also been reported to facilitate a better understanding of the evolutionary events that lead to the emergence of pathogenic V. cholerae clones ^{15, 16} .

Despite the World Health Organization (WHO) recommendations in regards to the management of cholera with oral rehydration salts in addition to antibiotics such as streptomycin, aminoglycosides, trimethoprim, fosfomycin, fluoroquinolones, sulphonamides, chloramphenicol/florfenicol, and tetracyclines, this study reports genotypic resistance in the isolates to these same antibiotics. Similar resistance has been reported in similar studies from the East African region and elsewhere ^{5, 17– 19} .

The presence of integrative and conjugative elements (VC1786ICE), containing resistance genes associated with sulfamethoxazole and trimethoprim, chloramphenicol, and streptomycin resistance, are also reported in this study. These are genomically similar to V. cholerae ICEVchHai1 ^{17, 19} .

This study found no plasmids or intl genes. This could be attributed to the presence of integrative and conjugative elements, a factor that made them insignificant in regards to the encoding of antimicrobial resistance. Studies similar to this have reported similar findings ⁵ .

This study also reported the presence of insertion sequences IS605, IS66, IS3, and IS4. Insertion sequences have been described in various studies to be drivers of genetic variability. These studies have also alluded to them being fixed by natural selection each time that a mutation induced by these elements is selected ^{20, 21} .

The presence of T6SS genes in the study isolates could explain their antimicrobial resistance gene profile. TS66-dependent killing of other bacteria is mostly directed to neighboring cells. These consequently release their DNA, which is ultimately taken up by the killer cells and in the process, these can integrate valuable genes including those that encode antimicrobial resistance. These may consequently evolve, leading to antimicrobial resistance in the killer cells ²² .

The results obtained from the SNP-based phylogenetic analysis show the relatedness of the Ugandan V. cholerae strains and are in agreement with the results obtained by Bwire et al., 2018.

The analysis for shared SNPs among the sequences and, consequently, gene enrichment revealed the enrichment in genes that mediate transmembrane-signaling receptor activity, peptidyl-prolyl cis-trans isomerase activity, and phosphor relay response regulator activity. These play a fundamental role in quorum sensing in V. cholerae, a process of cell-cell communication that allows these bacteria to share information about cell density and adjust gene expression accordingly ^{23– 25} . Quorum sensing has been documented to regulate the expression of virulence factors in V. cholerae ^{23– 25} .

Despite the fact that bioinformatics capacity remains underdeveloped in Uganda and Africa as a whole, this study demonstrated the ability to apply bioinformatics approaches to zoom into genomes (in this case, V. cholerae genomes obtained from Uganda) to provide a comprehensive genomic analysis. This study sets a stage that encourages more sequencing work with potential public health consequences to be done in African settings. Furthermore, it also encourages the need to build bioinformatics capacity in African settings to enable analysis of whole-genome sequence data generated from the continent.