Abstract
ABSTRACTBrucella microti was isolated a decade ago from wildlife and soil in Europe. Compared to the classical Brucella species, it exhibits atypical virulence properties such as increased growth in human and murine macrophages and lethality in experimentally infected mice. A spontaneous rough (R) mutant strain, derived from the smooth reference strain CCM4915T, showed increased macrophage colonization and was non-lethal in murine infections. Whole-genome sequencing and construction of an isogenic mutant of B. microti and Brucella suis 1330 revealed that the R-phenotype was due to a deletion in a single gene, namely wbkE (BMI_I539), encoding a putative glycosyltransferase involved in lipopolysaccharide (LPS) O-polysaccharide biosynthesis. Complementation of the R-strains with the wbkE gene restored the smooth phenotype and the ability of B. microti to kill infected mice. LPS with an intact O-polysaccharide is therefore essential for lethal B. microti infections in the murine model, demonstrating its importance in pathogenesis.
Highlights
Brucellae are Gram-negative facultative intracellular coccobacilli causing brucellosis, a major bacterial zoonosis with 500,000 human cases globally reported every year [1]
Whole-genome sequencing and construction of an isogenic mutant of B. microti and Brucella suis 1330 revealed that the R-phenotype was due to a deletion in a single gene, namely wbkE (BMI_I539), encoding a putative glycosyltransferase involved in lipopolysaccharide (LPS) O-polysaccharide biosynthesis
The lethal phenotype in mice depends on the type IV secretion system VirB [12] and is a general unambiguous criterion to establish if a specific Brucella gene plays a role in virulence of B. microti, as wild-type bacteria kill the murine host at the infection dose of 105 CFU; in contrast, in classical species virulence has been correlated to the capacity of a strain to establish or maintain various degrees of chronic infection of the spleen and/ or the liver, necessitating repeated bacterial enumeration in these organs to follow up the course of infection
Summary
Brucellae are Gram-negative facultative intracellular coccobacilli causing brucellosis, a major bacterial zoonosis with 500,000 human cases globally reported every year [1]. New species of Brucella, such as Brucella microti, Brucella inopinata and isolates from Australian rodents and amphibians, have been described [2] These strains are metabolically more active, acidresistant and fast-growing when compared to the wellknown classical, human-pathogenic Brucella species, which include Brucella abortus, Brucella melitensis, Brucella suis and Brucella canis [2,3,4,5,6,7]. B. microti was isolated from common vole, red fox, wild boar, and soil in Central Europe and, more recently, from a domestic marsh frog farm [8,9] This species is closer to those pathogenic for human and livestock than to the group of newly described atypical species/strains [2,10]. Lethality in mice was later demonstrated for B. inopinata BO1 and supplemental data for this article can be accessed here
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