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PMID: 9488389 Published · ppublish English Journal Article Research Support, U.S. Gov't, Non-P.H.S.

Transposon-derived Brucella abortus rough mutants are attenuated and exhibit reduced intracellular survival.

Infection and immunity ·Vol. 66 ·No. 3 ·1998-03-00 ·Pages 1008-16

Allen CA, Adams LG, Ficht TA

Abstract

The O antigen of Brucella abortus has been described as a major virulence determinant based on the attenuated survival of fortuitously isolated rough variants. However, the lack of genetic definition of these mutants and the virulence of naturally occurring rough species, Brucella ovis and Brucella canis, has confused interpretation. To better characterize the role of O antigen in virulence and survival, transposon mutagenesis was used to generate B. abortus rough mutants defective in O-antigen presentation. Sequence analysis of DNA flanking the site of Tn5 insertion was used to verify insertion in genes encoding lipopolysaccharide (LPS) biosynthetic functions. Not surprisingly, each of the rough mutants was attenuated for survival in mice, but unexpected differences among the mutants were observed. In an effort to define the basis for the observed differences, the structure of the rough LPS and the sensitivity of these mutants to individual killing mechanisms were examined in vitro. All of the B. abortus rough mutants exhibited a 4- to 5-log-unit increase, compared to the smooth parental strain, in sensitivity to complement-mediated lysis. Little change was evident in the sensitivity of these organisms to hydrogen peroxide, consistent with an inability of O antigen to exclude relatively small molecules. Sensitivity to polymyxin B, which was employed as a model cationic, amphipathic peptide similar to defensins found in phagocytic cells, revealed survival differences among the rough mutants similar to those observed in the mouse. One mutant in particular exhibited hypersensitivity to polymyxin B and reduced survival in mice. This mutant was characterized by a truncated rough LPS. DNA sequence analysis of this mutant revealed a transposon interruption in the gene encoding phosphomannomutase (pmm), suggesting that this activity may be required for the synthesis of a full-length core polysaccharide in addition to O antigen. B. abortus O antigen appears to be essential for extra- and intracellular survival in mice.

MeSH Terms
Amino Acid Sequence Animals Brucella abortus/genetics,immunology,pathogenicity Complement System Proteins/immunology DNA Transposable Elements Mice Mice, Inbred BALB C Molecular Sequence Data Mutation O Antigens/biosynthesis Phagocytosis Polymyxin B/pharmacology Virulence
Chemicals
DNA Transposable Elements O Antigens Complement System Proteins Polymyxin B
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Allen C A
Department of Veterinary Pathobiology, Texas A&M University, College Station 77843-4467, USA.
Adams L G
Ficht T A
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Article Info
Journal
Infection and immunity
Abbr.
Infect Immun
ISSN
0019-9567
DOI
10.1128/IAI.66.3.1008-1016.1998
Published
1998-03-00
Pages
1008-16
Language
English
Region
United States
NLM ID
0246127
PMCID
PMC108009
Subset
IM
External Links
PubMed source DOI: 10.1128/IAI.66.3.1008-1016.1998 PMC Full Text View journal details
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