Summary.A restriction fragment length polymorphism (RFLP) typing method for Legionella pneumophila serogroup 1 was developed. The method depended upon the use of cloned EcoRl fragments from L. pneumophila (Knoxville-1) probing Neil restriction fragments of chromosomal DNA. Examination of strains of L. pneumophila which were apparently unrelated showed that inter-strain RFLPs were common, and these formed the basis of the typing scheme. The technique was found to be highly reproducible and discriminatory. When the RFLP data were compared to that obtained by monoclonal antibody (MAb) subgrouping both methods of strain differentiation gave consistent results. The isolates examined by either method were also sub-divided by the alternative technique. The analysis of RFLPs by cloned probes should be of considerable epidemiological value.
SUMMARYOne hundred and seventy-nine isolates of Legionella pneumophila serogroup 1, obtained from a site associated with an outbreak of Legionnaires' disease, were examined by monoclonal antibody subgrouping, restriction fragment length polymorphism typing, restriction endonuclease analysis and plasmid content. Nine distinct phenotypes were detected but at the genotypic level all strains were closely related. The data presented indicate that phenotypic variation of a single parent strain can occur within an environmental site. The implications of these findings are discussed in relation to the investigation of outbreaks of Legionnaires' disease.
Summary.The relationship between serogroup and ,genotype of Legionella pneumophila strains was investigated by restriction fragment length polymorphism (RFLP) typing with a previously standardised method. Of the 51 RFLP types identified, 19 comprised strains of more than one serogroup. Several RFLP types included strains of five or more serogroups. To determine if sharing the same RFLP type indicates that strains are genotypically indistinguishable or merely that they are superficially similar, 31 strains were selected for further analysis with an extended range of restriction endonucleases and nucleic acid probes. In some cases, strains of a particular RFLP type were indistinguishable, while in others the restriction fragment patterns showed minor differences. It is possible that in the latter case the strains are diverging representatives of a parent clone. We conclude that analysis of restriction fragment patterns, either probed or unprobed, provides a more accurate measure of the ancestral relationship between strains than can be obtained with serological methods.
A typing method based on analysis of restriction fragment length polymorphisms has previously been developed for Legionella pneumophila serogroup 1. Here data are presented demonstrating the utility of this method for typing strains of all other L. pneumophila serogroups described to date. The method, which is highly discriminatory, should be of considerable value in epidemiological investigations of legionella infections.
Summary. The use of probes derived from rRNA sequences to detect restriction fragment length polymorphisms (RFLPs) associated with the ribosomal RNA genes for epidemiological typing (ribotyping) is a powerful and readily applicable tool. Different probes and enzymes for ribotyping were compared for a series of 73 unrelated Legionella pneumophila serogroup 1 strains. The probes compared were cDNAs, transcribed from L. pneumophila or Escherichia coli rRNA subunits, and a cloned L. pneumophila rRNA gene. The cloned rRNA gene probe gave the best discrimination and this probe was further compared with cloned probes comprised of randomly selected (non-rRNA) parts of the L. pneumophila chromosome. In this instance the greatest discrimination was achieved when one of the non-ribosomal RNA gene probes was employed. The overall discrimination of RFLP typing was enhanced by combining the data obtained with both rRNA and non-rRNA probes.
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