Unprecedented Melioidosis Cases in Northern Australia Caused by an Asian Burkholderia pseudomallei Strain Identified by Using Large-Scale Comparative Genomics.

Melioidosis is a disease of humans and animals that is caused by the saprophytic bacterium Burkholderia pseudomallei. Once thought to be confined to certain locations, the known presence of B. pseudomallei is expanding as more regions of endemicity are uncovered. There is no vaccine for melioidosis, and even with antibiotic administration, the mortality rate is as high as 40% in some regions that are endemic for the infection. Despite high levels of recombination, phylogenetic reconstruction of B. pseudomallei populations using whole-genome sequencing (WGS) has revealed surprisingly robust biogeographic separation between isolates from Australia and Asia. To date, there have been no confirmed autochthonous melioidosis cases in Australia caused by an Asian isolate; likewise, no autochthonous cases in Asia have been identified as Australian in origin. Here, we used comparative genomic analysis of 455 B. pseudomallei genomes to confirm the unprecedented presence of an Asian clone, sequence type 562 (ST-562), in Darwin, northern Australia. First observed in Darwin in 2005, the incidence of melioidosis cases attributable to ST-562 infection has steadily risen, and it is now a common strain in Darwin. Intriguingly, the Australian ST-562 appears to be geographically restricted to a single locale and is genetically less diverse than other common STs from this region, indicating a recent introduction of this clone into northern Australia. Detailed genomic and epidemiological investigations of new clinical and environmental B. pseudomallei isolates in the Darwin region and ST-562 isolates from Asia will be critical for understanding the origin, distribution, and dissemination of this emerging clone in northern Australia.

Tracing melioidosis back to the source: using whole-genome sequencing to investigate an outbreak originating from a contaminated domestic water supply.

Melioidosis, a disease of public health importance in Southeast Asia and northern Australia, is caused by the Gram-negative soil bacillus Burkholderia pseudomallei. Melioidosis is typically acquired through environmental exposure, and case clusters are rare, even in regions where the disease is endemic. B. pseudomallei is classed as a tier 1 select agent by the Centers for Disease Control and Prevention; from a biodefense perspective, source attribution is vital in an outbreak scenario to rule out a deliberate release. Two cases of melioidosis within a 3-month period at a residence in rural northern Australia prompted an investigation to determine the source of exposure. B. pseudomallei isolates from the property's groundwater supply matched the multilocus sequence type of the clinical isolates. Whole-genome sequencing confirmed the water supply as the probable source of infection in both cases, with the clinical isolates differing from the likely infecting environmental strain by just one single nucleotide polymorphism (SNP) each. For the first time, we report a phylogenetic analysis of genomewide insertion/deletion (indel) data, an approach conventionally viewed as problematic due to high mutation rates and homoplasy. Our whole-genome indel analysis was concordant with the SNP phylogeny, and these two combined data sets provided greater resolution and a better fit with our epidemiological chronology of events. Collectively, this investigation represents a highly accurate account of source attribution in a melioidosis outbreak and gives further insight into a frequently overlooked reservoir of B. pseudomallei. Our methods and findings have important implications for outbreak source tracing of this bacterium and other highly recombinogenic pathogens.

Distribution of Burkholderia pseudomallei in northern Australia, a land of diversity.

Burkholderia pseudomallei is a Gram-negative soil bacillus that is the etiological agent of melioidosis and a biothreat agent. Little is known about the biogeography of this bacterium in Australia, despite its hyperendemicity in the northern region of this continent. The population structure of 953 Australian B. pseudomallei strains representing 779 and 174 isolates of clinical and environmental origins, respectively, was analyzed using multilocus sequence typing (MLST). Bayesian population structure and network SplitsTree analyses were performed on concatenated MLST loci, and sequence type (ST) diversity and evenness were examined using Simpson's and Pielou's indices and a multivariate dissimilarity matrix. Bayesian analysis found two B. pseudomallei populations in Australia that were geographically distinct; isolates from the Northern Territory were grouped mainly into the first population, whereas the majority of isolates from Queensland were grouped in a second population. Differences in ST evenness were observed between sampling areas, confirming that B. pseudomallei is widespread and established across northern Australia, with a large number of fragmented habitats. ST analysis showed that B. pseudomallei populations diversified as the sampling area increased. This observation was in contrast to smaller sampling areas where a few STs predominated, suggesting that B. pseudomallei populations are ecologically established and not frequently dispersed. Interestingly, there was no identifiable ST bias between clinical and environmental isolates, suggesting the potential for all culturable B. pseudomallei isolates to cause disease. Our findings have important implications for understanding the ecology of B. pseudomallei in Australia and for potential source attribution of this bacterium in the event of unexpected cases of melioidosis.

In-house testing for COVID-19: effects on length of stay, isolation and the need for inpatient rehabilitation.

Objective The COVID-19 pandemic has strained healthcare worldwide. Its direct complications, management and prognosis have been described. Downstream effects, including length of hospital stay (LOS), implications on discharge planning, and effect of in-house testing require formal study. Methods A retrospective cohort study of patients suspected of COVID-19 infection admitted at a metropolitan Australian hospital was conducted. Outcomes before and after availability of in-house COVID-19 testing were compared. Results A total of 129 admissions were analysed. Indications for COVID-testing were dyspnoea (61.2%), fever (19.3%) and delirium (10.8%). All tested negative for COVID-19. Prior to in-house testing, mean LOS was 7.17 days (s.d. ± 4.2), and mean isolation of 1.8 days (s.d. ± 0.8). After availability of in-house testing, mean LOS was 4.78 days (s.d. ± 4.3) with mean isolation of 1.3 days (s.d. ± 0.9), both statistically significant differences. There were five inpatient falls, equivalent to 14.8 falls per 1000 patient/days. Twenty-two patients (17%) required subsequent sub-acute admission, 15 before in-house testing and five after (P = 0.058); however, a sub-group analysis for age >65 years was performed, and the results were significant (P < 0.05), showing all patients who required subacute admissions were aged >65. Conclusion In-house COVID-19 testing is suggested to significantly reduce the duration patients spend in isolation and overall LOS in hospital. A shorter period of isolation and hospital LOS may reduce the need for subacute transfer in patients aged greater than 65 years, as well as the rates of inpatient falls. Large scale studies are needed to further elucidate these findings.

Suspected cases of intracontinental Burkholderia pseudomallei sequence type homoplasy resolved using whole-genome sequencing.

Burkholderia pseudomallei is a Gram-negative environmental bacterium that causes melioidosis, a disease of high mortality in humans and animals. Multilocus sequence typing (MLST) is a popular and portable genotyping method that has been used extensively to characterise the genetic diversity of B. pseudomallei populations. MLST has been central to our understanding of the underlying phylogeographical signal present in the B. pseudomallei genome, revealing distinct populations on both the intra- and the inter-continental level. However, due to its high recombination rate, it is possible for B. pseudomallei isolates to share the same multilocus sequence type (ST) despite being genetically and geographically distinct, with two cases of 'ST homoplasy' recently reported between Cambodian and Australian B. pseudomallei isolates. This phenomenon can dramatically confound conclusions about melioidosis transmission patterns and source attribution, a critical issue for bacteria such as B. pseudomallei that are of concern due to their potential for use as bioweapons. In this study, we used whole-genome sequencing to identify the first reported instances of intracontinental ST homoplasy, which involved ST-722 and ST-804 B. pseudomallei isolates separated by large geographical distances. In contrast, a third suspected homoplasy case was shown to be a true long-range (460 km) dispersal event between a remote Australian island and the Australian mainland. Our results show that, whilst a highly useful and portable method, MLST can occasionally lead to erroneous conclusions about isolate origin and disease attribution. In cases where a shared ST is identified between geographically distant locales, whole-genome sequencing should be used to resolve strain origin.

Burkholderia pseudomallei Genotype Distribution in the Northern Territory, Australia.

Melioidosis is a tropical disease of high mortality caused by the environmental bacterium, Burkholderia pseudomallei. We have collected clinical isolates from the highly endemic Northern Territory of Australia routinely since 1989, and animal and environmental B. pseudomallei isolates since 1991. Here we provide a complete record of all B. pseudomallei multilocus sequence types (STs) found in the Northern Territory to date, and distribution maps of the eight most common environmental STs. We observed surprisingly restricted geographic distributions of STs, which is contrary to previous reports suggesting widespread environmental dissemination of this bacterium. Our data suggest that B. pseudomallei from soil and water does not frequently disperse long distances following severe weather events or by migration of infected animals.

Melioidosis from contaminated bore water and successful UV sterilization.

Two cases of melioidosis at a residence in rural northern Australia were linked to the unchlorinated domestic bore (automated well) water supply, which was found to have a high concentration of Burkholderia pseudomallei. Using multilocus sequence typing, clinical B. pseudomallei isolates from both cases were identical to an isolate from the bore water supply. A simple UV sterilizer reduced B. pseudomallei from the domestic water supply to undetectable levels. We have shown that UV treatment is highly effective for remediation of water contaminated with B. pseudomallei and recommend its consideration in households where individuals may be at heightened risk of contracting melioidosis.