Genomic and morphological analysis of human clinical Burkholderia pseudomallei isolates from New Caledonia (2021−2022)

Melioidosis is an infectious disease caused by the bacterium Burkholderia pseudomallei that is prevalent in New Caledonia. This study aimed to characterize recent human cases, evaluate the genomic diversity of circulating strains, and develop a rapid genotyping tool for local isolates. Thirteen human cases were identified between 2021 and 2022, all of whom survived. Most cases occurred between February and April, coinciding with extreme weather events associated with La Niña. The clinical isolates were characterized using morphological assessments, whole-genome sequencing, multilocus sequence typing (MLST), and single-nucleotide polymorphism (SNP) analysis. Morphological and genomic analyses revealed substantial strain diversity. ST292 remained predominant, but several previously unreported MLST sequence types, differing by a single SNP at one or a few loci, were detected, particularly along the west coast. Although New Caledonian strains are closely related to other Australasian strains, SNP-based phylogenomic analyses clearly distinguish them from other regional isolates. Within New Caledonia, strains cluster phylogenetically into several well-supported branches. To rapidly discriminate these branches, we developed a high-resolution melting (HRM) PCR assay targeting 13 SNPs, enabling accurate assignment of strains. Analyses of morphologically distinct subcultures from individual patients revealed subtle variations, including one subculture (22-9817_4 R#1) with a reduced genome and deletions of key virulence genes, highlighting the presence of distinct bacterial subpopulations within a single patient. These findings confirm the endemicity of B. pseudomallei in New Caledonia, underscore the importance of analysing multiple colonies, and support integrated One Health strategies combining human, animal, environmental, and meteorological data to anticipate and mitigate future cases.