Abstract
Burkholderia pseudomallei is an important human pathogen whose infection biology is still poorly understood. The bacterium is endemic to tropical regions, including South East Asia and Northern Australia, where it causes melioidosis, a serious disease associated with both high mortality and antibiotic resistance. B. pseudomallei is a Gram-negative facultative intracellular pathogen that is able to replicate in macrophages. However despite the critical nature of its interaction with macrophages, few anti-macrophage factors have been characterized to date. Here we perform a genome-wide gain of function screen of B. pseudomallei strain K96243 to identify loci encoding factors with anti-macrophage activity. We identify a total of 113 such loci scattered across both chromosomes, with positive gene clusters encoding transporters and secretion systems, enzymes/toxins, secondary metabolite, biofilm, adhesion and signal response related factors. Further phenotypic analysis of four of these regions shows that the encoded factors cause striking cellular phenotypes relevant to infection biology, including apoptosis, formation of actin ‘tails’ and multi-nucleation within treated macrophages. The detailed analysis of the remaining host of loci will facilitate genetic dissection of the interaction of this important pathogen with host macrophages and thus further elucidate this critical part of its infection cycle.
Highlights
The Gram-negative bacterium Burkholderia pseudomallei is a serious environmental pathogen of man and the causative agent of the often fatal disease melioidosis
Known bacterial factors affecting the interaction with host cells include the bacterial capsule, and effectors delivered by the type III and type VI secretion systems (T3SS and T6SS) [5]
Both of these observations are consistent with the anti-macrophage loci being associated with genomic islands (GIs) and/or being recently horizontally transferred
Summary
The Gram-negative bacterium Burkholderia pseudomallei is a serious environmental pathogen of man and the causative agent of the often fatal disease melioidosis. Australia where infections are associated with both antibiotic resistance and high mortality rates (,50%) The ability of this pathogen to infect via inhalation has necessitated its listing as a potential bio-warfare agent [4]. Different Burkholderia strains show a wide range of different interactions with human macrophages, ranging from no effect, to host cell apoptosis and capase-1-dependent lysis [3,13]. This range of different responses to macrophages suggests that the complement of antimacrophage virulence factors encoded by the genome of different strains may differ dramatically and may indicate potential functional redundancy amongst such factors. We perform a simple gain of function screen in recombinant Escherichia coli to identify the full list of loci potentially encoding toxins, or other factors, with antimacrophage activity
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