Identification and cloning of a 27-kDa Coxiella burnetii immunoreactive protein

Patients with acute Q fever develop Coxiella burnetii-specific antibody, and chronic disease patients often develop extremely high levels of C. burnetii-specific antibody. Antibody-reactive LPS has been well characterized, but only a few immunoreactive proteins have been identified. An immunoreactive ca. 62-kDa protein has been cloned, sequenced, and shown to be related to an Escherichia coli heat-shock protein. A ca. 27-kDa immunoreactive outer membrane protein(s) has also been identified. We have begun characterizing C. burnetii immunoreactive proteins by gene-cloning methods. A gene bank of total C. burnetii Nine Mile phase 1 (acute strain) DNA was created using the lambda vector EMBL3. This bank was screened for plaques which reacted with E. coli-pre-absorbed rabbit antisera specific for purified, Nine Mile C. burnetii whole cells. Twenty-three immunoreactive plaques were identified from a screening of 3000 plaques. Twenty-two plaques produced a ca. 60-65-kDa immunoreactive protein. One plaque produced a ca. 27-kDa immunoreactive protein, as well as a ca. 60-65-kDa immunoreactive protein. Phage extracts of this plaque were used to enrich the antisera to produce a ca. 27-kDa-specific antisera. This enriched sera reacted with a ca. 27-kDa protein in all C. burnetii isolates tested, comprising isolates from both chronic and acute strains. The DNA contained in the immunoreactive clone was C. burnetii-specific, as shown by DNA hybridization. We are currently subcloning the ca. 27-kDa protein-coding region for sequencing to determine if this gene encodes the previously identified ca. 27-kDa immunodominant protein. This protein may ultimately have both diagnostic and vaccinogenic potential.