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To study the structure and function of the Legionella flagellum, we screened a genomic L. micdadei library in Escherichia coli for expression of the flagellin (Fla) subunit. One recombinant clone, JM105 (pHI5588), producing a truncated Fla protein of 40.5 kDa was identified. The plasmid pHI5588 carried a L. micdadei DNA insert of 5 kb, containing ca 95% of the fla gene. The complete DNA sequence of the L. micdadei fla gene was obtained by combining sequence data from pHI5588 with results using a polymerase chain reaction-based system for genome walking (vectorette PCR). The L. micdadei fla gene shared a high degree of homology with other flagellin genes in the amino- and carboxy termini, whereas the central region was found to be nonconserved. The fla sequence will facilitate the cloning of Fla proteins from other Legionella species and the study of flagella in the pathogenesis of Legionnaires' disease.

To study individual Legionella antigens, a Legionella micdadei genomic library in Escherichia coli SC181 was established. Partially Sau3A digested L. micdadei DNA fragments (15–25 kilobase pairs (kb)) were cloned into the tetracycline resistance gene of the cosmid vector pHC79. Four thousand ampicillin resistant recombinants were obtained; seven hundred were screened for expression of Legionella antigens in Western blot analysis with a polyspecific E. coli -absorbed anti-L. micdadei rabbit antibody. One of the positive clones expressed a 60 kilodalton (K) antigen, which reacted strongly with a monospecific rabbit antiserum raised against L. micdadei“common antigen” (CA), and an additional 13 K L. micdadei protein. The region encoding these two proteins from the 17 kb recombinant plasmid (pBA 2) was subcloned in pBGS 18+. The DNA sequence of the CA encoding region in the 2.7 kb subcloned fragment will provide important information with respect to genetic vs. antigenic relatedness among Legionellae and other Gram-negative species, as well as to CA structure and possible function.