Sets of EcoRI fragments containing ribosomal RNA sequences are conserved among different strains of Listeria monocytogenes.

Abstract

To classify Listeria monocytogenes using taxonomic characters derived from the rRNA operons and their flanking sequences, we studied a sample of 1346 strains within the taxon. DNA from each strain was digested with a restriction endonuclease, EcoRI. The fragments were separated by gel electrophoresis, immobilized on a membrane, and hybridized with a labeled rRNA operon from Escherichia coli. The pattern of bands, positions, and intensities of hybridized fragments were electronically captured. Software was used to normalize the band positions relative to standards, scale the signal intensity, and reduce the background so that each strain was reproducibly represented in a data base as a pattern. With these methods, L. monocytogenes was resolved into 50 pattern types differing in the length of at least one polymorphic fragment. Pattern types representing multiple strains were recorded as the mathematical average of the strain patterns. Pattern types were arranged by size polymorphisms of assigned rRNA regions into subsets, which revealed the branching genetic structure of the species. Subtracting the polymorphic variants of a specific assigned region from the pattern types and averaging the types within each subset resulted in reduced sets of conserved fragments that could be used to recognize strains of the species. Pattern types and reduced sets of conserved fragments were conserved among different strains of L. monocytogenes but were not observed in total among strains of other species.Strains of Listeria monocytogenes are classified into the taxon based on genotypic and phenotypic similarities (1, 2). A general method for classification and identification of strains by using DNA restriction fragments containing portions of rRNA operons has been described (3, 4). This method has been applied to the genus Listeria (5) and to L. monocytogenes (6), demonstrating its utility for classifying, identifying, and typing strains.We have described a standard method for species description by using conserved sets of species-specific rRNA gene restriction endonuclease-derived fragments (7). In the present study, > 1000 strains ofL. monocytogenes were characterized by using EcoRI fragments containing sequences complementary to an rRNA operon from Escherichia coli. The pattern structure of the species was described in detail by the use of fixed electrophoretic conditions, fragment standards, electronic imaging, and software for mobility normalization. In addition, we introduced the use of continuous-scale relative intensity in recording patterns from -9000 strains of -200 species. The L. monocytogenes patterns were arranged into the taxonomic structure by the use of squared correlation values (8) and visual assessment. We assigned letter names to the rRNA regions, each containing a given part of a given operon, and differentiated strains by restriction fragment length polymorphisms ofThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "adverti...

Genetic and phenotypic characterization of Listeria monocytogenes lineage III

“…Fifty-two L. monocytogenes isolates in our strain collection (www.pathogentracker.net) were identified as belonging to lineage III based on their EcoRI ribotypes and hly PCR-RFLP types (Bruce et al, 1995). Forty-six of those isolates were chosen for inclusion in this study (Table 1); six isolates were eliminated because their sources of isolation (e.g.…”

Comparison of ribotyping and arbitrarily primed PCR for molecular typing of Salmonella enterica and relationships between strains on the basis of these molecular markers

“…1988). This has also been proposed in E. coli (Caugant et al 1986). Relevant to this hypothesis of horizontal transfer, it has been suggested that the relative positions of the y f b gene cluster (genes for the biosynthesis and assembly of the 0 lipopolysaccharide) and the H1 and H2 genes (flagellar antigens), being separated by three map units on the chromosome (Sanderson and Hurley 1987), could be transferred by conjugation, in which case only a few recombinational events would be required to account for the observed distribution of serotypes (Beltran et al 1988).…”

“…The Riboprinter automated system was straightforward to use and generated profiles of eight samples within 4 h. This type of automated ribotyping has been reported to provide discrimination between different strains of Listeria monocytogenes (Bruce et al 1995) and Stuphylococcus sciuri (Webster etal. 1994).…”

Halomonas alkaliphila sp. nov., a novel halotolerant alkaliphilic bacterium isolated from a salt pool in Campania (Italy)

“…The digestion with EcoRI restriction endonuclease resulted in resolved patterns consisting of at least 6 bands for Halomonas venusta DSM 4743 T and 18bAG T , which were well suited for comparison (Fig. 2) (Bruce et al, 1995). The patterns of the strains were highly different and showed a pattern similarity of only 0.50, data that differentiate strain 18bAG…”

“…RiboPrinting (Bruce, 1996;Bruce et al, 1995) was performed using the Qualicon TM RiboPrinter system (Qualicon, Wilmington, Del., USA) and following the manufacturer's recommended procedures. The digestion with EcoRI restriction endonuclease resulted in resolved patterns consisting of at least 6 bands for both strains.…”