Ampicillin tolerance of Legionella pneumophila for counter-selection of transconjugants in heterospecific matings with Escherichia coli donors

✍ Scribed by Bunty Jain; Larisa Bender; P. Christian Lück; Dr. Manfred Ott

• Publisher: John Wiley and Sons
• Year: 1992
• Tongue: English
• Weight: 416 KB
• Volume: 32
• Category: Article
• ISSN: 0233-111X
• DOI: 10.1002/jobm.3620320616

✦ Synopsis

Legioizellu yrieiri~zoplzilo, the causative agent of Legionaires' disease is semiti\ c to ampicillin. However, the slowly growing bacteria are not killed even by high doses of this antibiotic. This natural tolerancc was used for counter-selection of truns-conjugants in heterospecific matings with r k / ~r k / i ; ~ c,o/i as donor. This approach is useful for gentic manipulations in Lrgiorwlltr. a s it avoids the use of antibiotic-resistant variants, which have to be tested for full virulencc bel'ore usc. Genetic attempts have been carried out to study pathogenicity factors of Le,qioric//tr piieur?zopl~ilcr (CIANCIOTTO ef d. 1989 a). DNA transfer from E . d w i k h i t r cw/i donors to L. pi?ez/i?70~/7j/~/ recipients was achieved by conjugation. Broad host range plasmids of the incompatibility groups inrP and iizcQ were shown to replicate stably in I-. ~~/ 7 c , z / / ? 7 ~~~~~7 i ~[ / .

For counter-selection the L. piiezmiop/zi/rr host in heterospecific niatings. spontaneously arising streptomycin or rifampicin resistant variants were used ( SZETO and SHUMAN 1990, CIANCIOTTO et al. 1989a, 1989b). It has been reported, that L. p7cwmp/zi/t/ strains lose their virulence properties spontaneously, and that avirulent clones cannot revert to the virulent phase, even not after passage through animals (CAI-RENICII and JOHNSON 1988). Such variants must therefore be tested for full virulence before use in genetic manipulations. In this report we introduce an alternative approach of counter-selection recipients by using the natural ampicillin tolerance of L. pneiwiophilrr, thus avoiding the use of antibiotic resistant variants.