Antigenic, Immunologic and Genetic Characterization of Rough Strains B.abortus RB51, B.melitensis B115 and B.melitensis B18

Rosanna Adone,Giuseppina La Rosa,Michela Tarantino,Michele Muscillo,Massimiliano Francia,Laurence Van Melderen

doi:10.1371/journal.pone.0024073

Abstract

The lipopolysaccharide (LPS) is considered the major virulent factor in Brucella spp. Several genes have been identified involved in the synthesis of the three LPS components: lipid A, core and O-PS. Usually, Brucella strains devoid of O-PS (rough mutants) are less virulent than the wild type and do not induce undesirable interfering antibodies. Such of them proved to be protective against brucellosis in mice. Because of these favorable features, rough strains have been considered potential brucellosis vaccines. In this study, we evaluated the antigenic, immunologic and genetic characteristics of rough strains B.abortus RB51, B.melitensis B115 and B.melitensis B18. RB51 derived from B.abortus 2308 virulent strain and B115 is a natural rough strain in which the O-PS is present in the cytoplasm. B18 is a rough rifampin-resistan mutant isolated in our laboratory.The surface antigenicity of RB51, B115 and B18 was evaluated by testing their ability to bind antibodies induced by rough or smooth Brucella strains. The antibody response induced by each strain was evaluated in rabbits. Twenty-one genes, involved in the LPS-synthesis, were sequenced and compared with the B.melitensis 16M strain.The results indicated that RB51, B115 and B18 have differences in antigenicity, immunologic and genetic properties. Particularly, in B115 a nonsense mutation was detected in wzm gene, which could explain the intracellular localization of O-PS in this strain.Complementation studies to evaluate the precise role of each mutation in affecting Brucella morphology and its virulence, could provide useful information for the assessment of new, attenuated vaccines for brucellosis.

Highlights

In Brucella spp., as in many other gram-negative bacteria, the smooth lipopolysaccharide (S-LPS) is an important component of the outer membrane, strongly involved in pathogenesis mechanisms
In Complement Fixation Tests (CFTs) performed on whole-cell antigen suspensions of RB51, B18 and B115, B.abortus 99 smooth strain, tested as control, bound only anti-O-PS antibodies
RB51 and B115 bound antibodies induced by the rough strain RB51, and did not react with antiOPS antibodies induced by the smooth strain B.abortus 99

Summary

Introduction

In Brucella spp., as in many other gram-negative bacteria, the smooth lipopolysaccharide (S-LPS) is an important component of the outer membrane, strongly involved in pathogenesis mechanisms. Its precise role as a virulence factor is not yet clear. It has been suggested, that the LPS molecule may play a key role in the invasion and intracellular multiplication of Brucella spp. as well as in protecting the cell against complement-mediated lysis. The LPS is the immunodominant antigen to which the majority of antibodies resulting from either infection or vaccination are directed [1,2,3,4]. The S-LPS molecule has three sections: the lipid A, the core oligosaccharide and the distal O-polysaccharide chain (O-PS or O-antigen). The O-PS is a homopolymer of N-formyl-perosamine

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Conclusion