Identification of Legionella species by random amplified polymorphic DNA profiles

Abstract

Random amplified polymorphic DNA (RAPD) was used for the identification of Legionella species. Primer SK2 (5'-CGGCGGCGGCGG-3') and standardized RAPD conditions gave the technique a reproducibility of 93 to 100%, depending on the species tested. Species-specific patterns corresponding to the 42 Legionella species were consequently defined by this method; the patterns were dependent on the recognition of a core of common bands for each species. This specificity was demonstrated by testing 65 type strains and 265 environmental and clinical isolates. No serogroup-specific profiles were obtained. A number of unidentified Legionella isolates potentially corresponding to new species were clustered in four groups. RAPD analysis appears to be a rapid and reproducible technique for identification of Legionella isolates to the species level without further restriction or hybridization.

FIG. 1

RAPD amplification patterns of Legionella type strains. Lanes: M, molecular weight marker VI (Boehringer Mannheim); 1 and 16, L. pneumophila serogroup 1 strain ATCC 33152; 2, L. pneumophila serogroup 3 strain ATCC 33155; 3, L. micdadei ATCC 33218; 4, L. maceachernii ATCC 35300; 5, L. anisa ATCC 35292; 6, L. bozemanii serogroup 1 strain ATCC 33217; 7, L. parisiensis ATCC 35299; 8, L. gratiana ATCC 49413; 9, L. tucsonensis ATCC 49180; 10, L. longbeachae serogroup 1 strain ATCC 33462; 11, L. longbeachae serogroup 2 strain ATCC 33484; 12, L. erythra serogroup 1 strain ATCC 35303; 13, L. erythra serogroup 2 strain LC217; 14, L. rubrilucens ATCC 35304; 15, L. spiritensis serogroup 1 strain ATCC 49655.

FIG. 2

Schematic core of species- and subspecies-specific patterns of Legionella species. Schematic of RAPD patterns constructed with Taxotron software. The bands represented are those always present in a given species; bands are represented according to their intensities. a, L. pneumophila subsp. pneumophila; b, L. pneumophila subsp. fraseri; c, L. pneumophila subsp. pascullei.

FIG. 3

Visual identification of Legionella isolates by RAPD patterns. Lanes: M, molecular weight marker VI (Boehringer Mannheim); 1, L. erythra serogroup 1 strain ATCC 35302; 2, L. erythra serogroup 2 strain LC217; 3, L. erythra serogroup 2 strain Aix96VII2-1; 4, L. rubrilucens aix56; 5, L. rubrilucens ATCC 35304; 6, L. bozemanii serogroup 1 strain GAPH; 7, L. bozemanii serogroup 1 strain Bruxelles D; 8, L. bozemanii serogroup 1 strain DignB111; 9, L. bozemanii serogroup 2 strain Paris-96010250; 10, L. bozemanii serogroup 2 strain ATCC 35545; 11, L. feeleii serogroup 1 strain ATCC 35072; 12, L. feeleii serogroup 1 strain Ly126.92a; 13, L. londiniensis ATCC 49505; 14, L. londiniensis IP8-1; 15, L. jordanis ATCC 33623; 16, L. jordanis Ly95.96.