Tracing the environmental footprint of theBurkholderia pseudomalleilipopolysaccharide genotypes in the tropical 'Top End' of the Northern Territory, Australia

The Tier 1 select agentBurkholderia pseudomalleiis an environmental bacterium that causes melioidosis, a high mortality disease. Variably present genetic markers used to elucidate strain origin, relatedness and virulence inB. pseudomalleiinclude theBurkholderiaintracellular motility factor A (bimA) and filamentous hemagglutinin 3 (fhaB3) gene variants. Three lipopolysaccharide (LPS) O-antigen types inB. pseudomalleihave been described, which vary in proportion between Australian and Asian isolates. However, it remains unknown if these LPS types can be used as genetic markers for geospatial analysis within a contiguous melioidosis-endemic region. Using a combination of whole-genome sequencing (WGS), statistical analysis and geographical mapping, we examined if the LPS types can be used as geographical markers in the Northern Territory, Australia. The clinical isolates revealed that LPS A prevalence was highest in the Darwin and surrounds (n = 660; 96% being LPS A and 4% LPS B) and LPS B in the Katherine and Katherine remote and East Arnhem regions (n = 79; 60% being LPS A and 40% LPS B). Bivariate logistics regression of 999 clinicalB. pseudomalleiisolates revealed that the odds of getting a clinical isolate with LPS B was highest in East Arnhem in comparison to Darwin and surrounds (OR 19.5, 95% CI 9.1 – 42.0;pbimAgenotypebimABmhas a similar remote region geographical footprint to that of LPS B. In addition, correlation of LPS type with multi-locus sequence typing (MLST) was strong, and where multiple LPS types were identified within a single sequence type, WGS confirmed homoplasy of the MLST loci. The clinical, sero-diagnostic and vaccine implications of geographically-basedB. pseudomalleiLPS types, and their relationships to regional and global dispersal of melioidosis, require global collaborations with further analysis of larger clinically and geospatially-linked datasets.Author SummaryBurkholderia pseudomalleiis a pathogenic soil bacterium that causes the disease melioidosis, which occurs in many tropical regions globally and in recent years has emerged in non-tropical regions. Melioidosis has been predicted to affect 165,000 people every year resulting in an estimated 89,000 deaths. Person to person transmission is rare with most cases linked to exposure to the bacterium from the environment. The genetic background ofB. pseudomalleihas been well studied and variably present genes have been linked to distinct melioidosis disease states and geographic regions, however we still need a stronger understanding of the association of genes with geography. Three lipopolysaccharide types exist inB. pseudomalleiand the prevalence of the lipopolysaccharide genes vary between melioidosis endemic regions, but it is unknown if the lipopolysaccharide genes can be used as geographical markers in a single melioidosis-endemic region. In this study, we used a combination of whole-genome sequencing, statistics and geographical mapping to elucidate if the three lipopolysaccharide genes can be used as geographical markers within the Northern Territory, Australia. We show that the three LPS types have distinct but overlapping geographical footprints within a single melioidosis region and can be used as geographic markers alongside a number of other important variably presentB. pseudomalleigenes.