Chromosomal DNA Deletions Explain Phenotypic Characteristics of Two Antigenic Variants, Phase II and RSA 514 (Crazy), of theCoxiella burnetiiNine Mile Strain

Abstract

After repeated passages through embyronated eggs, the Nine Mile strain of Coxiella burnetii exhibits antigenic variation, a loss of virulence characteristics, and transition to a truncated lipopolysaccharide (LPS) structure. In two independently derived strains, Nine Mile phase II and RSA 514, these phenotypic changes were accompanied by a large chromosomal deletion (M. H. Vodkin and J. C. Williams, J. Gen. Microbiol. 132:2587-2594, 1986). In the work reported here, additional screening of a cosmid bank prepared from the wild-type strain was used to map the deletion termini of both mutant strains and to accumulate all the segments of DNA that comprise the two deletions. The corresponding DNAs were then sequenced and annotated. The Nine Mile phase II deletion was completely nested within the deletion of the RSA 514 strain. Basic alignment and homology studies indicated that a large group of LPS biosynthetic genes, arranged in an apparent O-antigen cluster, was deleted in both variants. Database homologies identified, in particular, mannose pathway genes and genes encoding sugar methylases and nucleotide sugar epimerase-dehydratase proteins. Candidate genes for addition of sugar units to the core oligosaccharide for synthesis of the rare sugar 6-deoxy-3-C-methylgulose (virenose) were identified in the deleted region. Repeats, redundancies, paralogous genes, and two regions with reduced G؉C contents were found within the deletions.Coxiella burnetii is an obligately parasitic bacterium that replicates within phagolysosomes of eukaryotic hosts (1,16,27). This organism is the causative agent of Q fever in humans. It is endemic in many species of domestic animals, especially sheep, goats, and cattle. C. burnetii is extremely infectious; inhalation of one organism is enough to initiate an acute illness in a guinea pig (32). In humans, the disease usually is a moderate to severe flu-like illness with headache, fever and chills, malaise, myalgia, and anorexia or an atypical pneumonia with a dry cough (26). Recrudescence and chronic illness, usually in the form of hepatitis or intractable endocarditis, are the most serious manifestations of the disease (24,25,28,55). Humans usually contract Q fever by inhaling contaminated dust, often in barnyard, stockyard, or abattoir settings (34, 45).Some strains of C. burnetii have been observed to undergo variation in surface antigens (18,40). Continual passage of the Nine Mile strain (the tick-derived United States prototype strain) through immunocompetent hosts (306 successive passages in guinea pigs) apparently maintained the organism in its original, native (wild-type) antigenic form. However, eight subsequent passages through embryonated eggs resulted in the emergence of organisms that were easily distinguished serologically from the original isolate (40). The new phenotype reacted strongly with complement-fixing antibody found in acute-phase serum, compared with the lack of a reaction demonstrated by the guinea pig-passaged antigenic form (40). The two antigenic types differed ...

VirulentCoxiella burnetiidoes not activate human dendritic cells: Role of lipopolysaccharide as a shielding molecule

“…Recent microarray analysis of the genomes of NMI and NMII has confirmed that the only difference between these strains, at the gene-deletion level, is a Ϸ26-kb deletion in NMII that eliminates a number of genes involved in LPS biosynthesis (ref. 3 and P. Beare and R.A.H., unpublished work). It would therefore seem logical to hypothesize that truncated NMII LPS directly stimulates DC, whereas full-length NMI LPS does not.…”

“…C. burnetii strains of decreased or no virulence in animal models have been generated through serial in vitro passage. These isogenic strains have undergone a ''phase variation'' event in which a chromosomal mutation affecting LPS biosynthesis has occurred (3). Avirulent or ''phase II'' bacteria, typified by the Nine Mile phase II (NMII) strain, produce an LPS that lacks the unique branched terminal sugar-containing O-polysaccharide chain that is characteristic of the LPS of virulent or ''phase I'' organisms, such as the Nine Mile phase I (NMI) strain (4).…”

Is Q Fever an emerging or re-emerging zoonosis?

“…Yet the LPS of C. burnetii has an endotoxic activity 100 to 1 000 times lower than that of the LPS of Enteriobacteriaceae [4], but it induces the production of inflammatory cytokines in murine and human macrophages [35,171]. Avirulent phase II bacteria are produced by spontaneous mutations or large genetic rearrangements that result in the synthesis of truncated forms of LPS, which lack the branched-chain sugars virenose and dihydrohydroxystreptose present in phase I LPS [64,65,164]. This phase variation is observed when Coxiella organisms are propagated in non-immunocompetent cell cultures or hen eggs and is irreversible [122].…”

Galleria mellonella as an alternative model of Coxiella burnetii infection

“…Phase I bacteria are virulent, in contrast to phase II bacteria which are avirulent in immunocompetent animal models of disease (Moos & Hackstadt, 1987;Andoh et al, 2007;Marmion, 2007). The difference in virulence of these strains has been defined as a consequence of differential O-antigen expression (Hoover et al, 2002). One potential reason for our poor understanding of C. burnetii pathobiology is a historical lack of molecular tools for genetic manipulation of the organism.…”