Molecular investigations of a locally acquired case of melioidosis in Southern AZ, USA

Abstract

Melioidosis is caused by Burkholderia pseudomallei, a Gram-negative bacillus, primarily found in soils in Southeast Asia and northern Australia. A recent case of melioidosis in non-endemic Arizona was determined to be the result of locally acquired infection, as the patient had no travel history to endemic regions and no previous history of disease. Diagnosis of the case was confirmed through multiple microbiologic and molecular techniques. To enhance the epidemiological analysis, we conducted several molecular genotyping procedures, including multi-locus sequence typing, SNP-profiling, and whole genome sequence typing. Each technique has different molecular epidemiologic advantages, all of which provided evidence that the infecting strain was most similar to those found in Southeast Asia, possibly originating in, or around, Malaysia. Advancements in new typing technologies provide genotyping resolution not previously available to public health investigators, allowing for more accurate source identification.

Figure 1. Population assignment of case isolate based on MLST sequence type.

An estimated population assignment of case isolate (ST426) compared to 640 other MLST sequence types was developed using Structure . Each thin vertical line represents one sequence type (ST) and is divided into 2 portions that represent the likelihood of assignment into each population. STs are sorted by likelihood of assignment into two populations. STs from strains collected in Australia dominate the left hand population and STs from strains collected in Southeast Asia dominate the right hand population. The case isolate was placed in the second population in 85% of 100,000 iterations.

Figure 2. Maximum parsimony phylogenetic comparison of case isolate sequence to all available B. pseudomallei sequences.

The case isolate whole genome sequence (WGS) was compared to 22 B. pseudomallei and 10 B. mallei sequences, using 54,135 shared SNPs (including 31,202 parsimony informative SNPs). MEGA5 was used to conduct maximum parsimony analysis of all SNP loci common to the genome sequences. The percentages of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) are shown next to the branches. The tree is drawn to scale, with branch lengths calculated using the average pathway method and are in the units of the number of changes over the whole sequence.