Conjugation-mediated genetic exchange in Legionella pneumophila

Abstract

Genetic exchange mechanisms, to our knowledge, have not been reported for Legionella pneumaphila, and consequently, studies on the genetic organization of L. pneumophila have not appeared in the literature. Here, we describe gene transfer mediated by broad host range conjugative plasmids in Legionella spp. Escherichia coli strains carrying plasmids RP1 and R68.45 (IncPl), S-a (IncW), and R40a (IncC), but not plasmids of incompatability groups Fl, FII, and FV, served as donors in matings with L. pneumophila Knoxville 1 (LPK-1). Transconjugants selected by resistance to kanamycin (RP1, R68.45, and S-a) and carbenicillin (R40a) were observed at frequencies of 6.6 x 10-, 4.7 x 10-, 2.2 x 10-, and 5.4 x 10-, respectively. Plasmid transfer was not affected by DNase added to the mating medium. After plasmid transfer, LPK-1 stably maintained RP1, R68.45, and S-a, but not R40a. Plasmid-containing LPK-1 isolates also served as donors in agar plate matings with E. coli W1485-1 and naladixic acid-resistant mutants of LPK-1, Legionella micdadei, and Legionella longbeachii. Recombinational exchange of a chromosomal trait was demonstrated when a thymidine auxotroph of L. pneumophila was repaired by R68.45-mediated chromosomal mobilization of a prototrophic donor strain.

Role of the 25 kDa major outer membrane protein ofLegionella pneumophilain attachment to U‐937 cells and its potential as a virulence factor for chick embryos

“…Introduction of genetic material into the legionellae has proved problematic. Studies have shown that under limited conditions, Legionella can accept foreign DNA through conjugal transfer and that this DNA can remain either in the cytoplasm, or in some instances, can be delivered into the chromosome (Chen et al 1984 ;Dreyfus and Iglewski 1985). Successful triparental matings using plasmids constructed with Leg.…”

Broad-host-range plasmid pRK340 delivers Tn5 into the Legionella pneumophila chromosome

“…We have recently added Tn3 (ampicillin resistance) to pRK340 and are presently evaluating the use of ampicillin resistance as a marker for the plasmid. Studies with R68.45 indicate that ampicillin is a strong selectable marker in L. pneumophila (8).…”

Anaerobic expression of thegyrAgene demonstrated by β-galactosidase activity ingyrA::Mu-lacfusion derivatives ofEscherichia coli

“…We first transformed E. coli C600Alac with pakl plasmid and selected for chloramphenicol resistance (15 ~g/ml). Using its tetracycline resistance marker, the helper conjugative plasmid R68.45, which carrys antibiotic resistance genes for ampicillin, kanamycin, neomycin and tetracycline [8], was placed by conjugation into E. coli C600Alac pakl. A gyrA::Mu-lac fusion strain was made resistant to rifampicin (25/tg/ml).…”

“…Bott. The helper conjugative plasmid, R68.45, capable of transferring other plasrnids to various bacterial species was sent to us by Dr. L.A. Dreyfus[8]. Introduction of B. subtilis gyrA gyrB region into the gyrA: :Mu-lac fusion strain ofE.…”