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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 ...

Monoclonal hybridoma antibodies directed against a 65-kilodalton (kDa) mycobacterial protein could detect similarly sized antigens in many other bacterial species. In Pseudomonas aeruginosa, the cross-reacting protein corresponded to a 62-kDa antigen that has been called Common Antigen. The mycobacterial 65-kDa antigen and Common Antigen are similar in that both (i) are highly immunoreactive molecules, (ii) contain species-specific and genus-specific epitopes in addition to the broadly cross-reactive epitopes, (iii) can be isolated as homomultimers of greater than 240 kDa, and (iv) have similar amino acid compositions. In Escherichia coli, the cross-reactive protein corresponded to the GroEL protein. Both the GroEL protein and the mycobacterial 65-kDa protein are expressed as heat shock proteins.

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A DNA fragment located on the 3' side of the CoxieUla burnetii htpAB operon was determined by Southern blotting to exist in approximately 19 copies in the Nine Mile I genome. The DNA sequences of this htpAB-associated repetitive element and two other independent copies were analyzed to determine the size and nature of the element. The three copies of the element were 1,450, 1,452, and 1,458 bp long, with less than 2% divergence among the three sequences. Several features characteristic of bacterial insertion sequences were discovered. These included a single significant open reading frame that would encode a 367-amino-acid polypeptide which was predicted to be highly basic, to have a DNA-binding helix-turn-helix motif, to have a leucine zipper motif, and to have homology to polypeptides found in several other bacterial insertion sequences. Identical 7-bp inverted repeats were found at the ends of all three copies of the element. However, duplications generated by many bacterial mobile elements in the recipient DNA during insertion events did not flank the inverted repeats of any of the three C. burnetii elements examined. A second pair of inverted repeats that flanked the open reading frame was also found in all three copies of the element. Most of the divergence among the three copies of the element occurred in the region between the two inverted repeat sequences in the 3' end of the element. Despite the sequence changes, all three copies of the element have retained significant dyad symmetry in this region.