Molecular phylogeny of Burkholderia pseudomallei
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Structures of bacterial populations
In order to understand the evolution, diversity and reproduction of living organisms some knowledge of their genetic structure is necessary. This provides a means of accounting for the observed differences between populations of the organism and placing these in the context of the degree of genetic diversity associated with the species. Some bacterial species are ubiquitous in nature and will contain innumerable strains while others are restricted to particular habitats and contain few strains.
Burkholderia in nature
The genus Burkholderia like many gram-negative saprophytic pseudomonads multiply in the rhizosphere which is defined as the region of soil modified as a result of the uptake and deposition of substances by a growing plant root. Here, bacteria growing in close association with root nodules assist in the fixation of nitrogen and promote uptake of minerals, particularly phosphate, by the plant. Organisms such as B. cepacia produce a wide range of antifungal and other compounds which are able to
Ribotypes of B. pseudomallei
The sequences of 16S and 23S rRNA molecules are most often specific for a single species. Their function is constant and the sequence changes at a rate consistent with the evolutionary rate of the organism. These elements can therefore be used to establish the most ancient relationships between organisms. The choice of restriction endonuclease utilized for ribotyping is of critical importance. A restriction enzyme recognizes specific sequences which are distributed around and within the rRNA
Clinical associations
The clinical features of 113 melioidosis patients infected with B. pseudomallei strains of different ribotypes were investigated to assess whether certain strains or groups were associated with a particular clinical picture. Ribotypes 1, 4 and 15 were markedly associated with fatal infections with the highest mortality observed for ribotype 4 (OR=3.36; 95% CI=0.58–20.58). The lowest mortality was associated with ribotypes 27 and 34. The risk of relapse with isolates of ribotype 1 was more than
Environmental strains
Two groups of ribotypes were observed among 139 environmental strains and 228 human isolates. Group I comprised 37 different ribotype patterns represented among the 44 ribotypes so far identified. All of these ribotypes were identified among the clinical isolates and 18 of them were also found among 59 environmental isolates. Group II was represented by 12 BamHI patterns of 1–5 bands of 9 kb to >23 kb. All but one of the 73 isolates in this group grew on a minimal medium supplemented with
DNA macrorestriction analysis and ribotype
DNA macrorestriction analysis probably allows at least 50% of the genome to be visualised as opposed to about 0.1% for ribotyping. As for ribotyping, the fragment positions can be subjected to numerical analysis to give statistical estimates of sequence similarity. Pulsed-field gel electrophoresis (PFGE) of XbaI digests of B. pseudomallei DNA proved to be highly discriminating and divided the 350 isolates of 44 ribotype patterns into 226 PFGE profile types. In general 15–24 bands were resolved
Clonality of B. pseudomallei
The considerable heterogeneity in the ribotypes of B. pseudomallei reflects the high level of diversity in rrn loci in the species. Three-quarters of the species fell in one of nine ribotypes and one-half were segregated into one of two ribotypes. Those groups of isolates which share both genotypic and epidemiological characteristics strongly suggest a common ancestral origin and indicates that the structure of B. pseudomallei at the rRNA level is clonal. An alternative view is that none of the
Population insights from other molecular studies
There is a high degree of homology between B. cepacia and B. pseudomallei flagellin gene sequences and B. cepacia flagellin gene probes can be used to identify B. pseudomallei flagellin genes in gene libraries. Winstanley et al. (1998) cloned and sequenced the flagellin genes from three clinical and one environmental isolate of B. pseudomallei and found them to be 100% identical or differing by only one base pair. Flagellin genotyping by PCR/RFLP revealed no differences between four clinical
Conclusions
B. pseudomallei is a heterogeneous species with extensive diversity in rrn loci. Two ribotypes predominate in nature and there is some evidence of geographical association in that ribotype 3 is prevalent in Asian countries while ribotype 1 is more wide-spread. Some disease association is apparent for strains of four ribotypes but there is no correlation between ribotype and underlying disease. DNA macrorestriction analysis reveals an almost limitless number of strain types. A distinct
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