Phenotypic and genotypic diversity of fluorescent pseudomonads isolated from field-grown sugar beet

Abstract

A sample of 30 fluorescent pseudomonads isolated from the phyllosphere of sugar beet throughout a single growing season and shown to be closely related on the basis of fatty acid methyl ester (FAME) analysis was subjected to detailed phenotypic and genotypic characterization. Phenotypic traits were assessed on the basis of biochemical properties, assimilation of sole carbon sources, FAME analysis, organic pyrolysate content (MS-pyrolysis), and total cellular protein profiles. With the exception of total cellular protein profiles, numerical analysis of the data revealed two main clusters, each of which was divided into several subclusters. Numerical analysis of total cellular protein data failed to differentiate isolates into two main clusters, but nevertheless grouped isolates into six subclusters. On the basis of biochemical and carbon source assimilation profiles, 19 isolates were identified as Pseudomonas fluorescens biovar v8 eight isolates as P. fluorescens biovar 111 and three isolates as P. syringae pathovar syringae. In general, all methods of phenotypic analysis grouped isolates according to time of sampling and leaf type. Genome analysis was undertaken by pulsed-f ield gel electrophoresis (PFGE) of Pad, Spel, Swal and Xbal macrorestriction fragments and revealed the presence of eight distinct genomic (clonal) groups. These groups correlated closely with the clusters generated by numerical analysis of phenotypic data, but there was no correlation between macrorestriction fragment profile and isolate identification; in fact the variation in macrorestriction fragment patterns within P. fluorescens biovars was as great as the variation detected between biovars, and between P. fluorescens and P. syringae. Statistical evaluation of macrorestriction fragment patterns revealed two examples of recent strain divergence: one was due to the presence of a 400 kbp plasmid within one isolate of a collection of nine otherwise genomically identical isolates, and the other was observed between two phenotypically similar isolates sampled 220 d apart. Genetic variation was expressed in terms of nucleotide diversity (z) and pairwise comparisons yielded values ranging from 0.0029 to 0-1 51 7. The mean intrapopulation genetic variation was high (0*0993), but limited genetic variation was detected among isolates sampled on each occasion. Taken together this suggests a population comprised of a variety of apparently distantly related clones (genomic groups), each adapted to local conditions. Genome sizes were estimated from the sum of Spel restriction fragments and ranged from 4 2 to 5.5 Mbp. Examination of the distribution of Xbal, Spel, Swal and Pacl restriction endonuclease sites showed that the distribution of Spel sites differed significantly from the expected (random) distribution. O'Gara, 1992). In addition, and as a result of the ease wii h which pseudomonads can be genetically manipulated, such strains are potential hosts for foreign genes targeted at the phyllosphere. However, before embarking upcn biocontrol projec...

Molecular and Metabolic Characterization of Cold-Tolerant Alpine Soil Pseudomonas Sensu Stricto

“…However, many of the same and similar reports state that gene exchanges between clusters may also explain some of their results (9,27,29,30), implying less than total congruence between genotypic (rDNA) and phenotypic (nutritional) data. Many studies of Pseudomonas have presented genotypic and phenotypic data and commented on their congruence (15,18,34,39), but few have rigorously tested such congruence. Our results, rigorously showing no congruence, suggest that statements of congruence should be based on careful tests of this issue and that the famous metabolic versatility of the genus might well be founded in large measure on the easy ability to acquire opportunistic metabolic capabilities.…”

The causes of Pseudomonas diversity

“…The diversity of phenotypes is also reflected at the genetic level, and evidence is mounting to suggest that the diversity of genome architecture (of both chromosomes and accessory genetic elements) is of particular significance. Fingerprinting studies performed by many workers have revealed a remarkable degree of restriction fragment polymorphism among strains of a speciesand even among strains that are closely related on phenotypic grounds (see, for example, Ginard et al, 1997 ;Grothues et al, 1988 ;Rainey et al, 1994). Genome sizes vary widely, ranging from 3n7 Mbp for Pseudomonas stutzeri (Ginard et al, 1997) to 7n1 Mbp for Pseudomonas aeruginosa (Schmidt et al, 1996), with the genome sizes of Pseudomonas fluorescens SBW25, Pseudomonas syringae pv.…”

Genotypic and phenotypic analysis of zwittermicin A-producing strains of Bacillus cereus

“…Others have shown that analyses of fatty acid methyl ester (FAME) profiles (Abel e t al., 1962;Kaneda, 1968;Lambert e t al., 1983;DeBoer & Sasser, 1986;Vaisanen e t al., 1991 ;Chase e t al., 1992;Rainey e t al., 1994), DNA patterns in pulsed-field gel electrophoresis (Carlson e t al., 1994;Rainey et al, 1994), RAPDs (Welsh & McClelland, 1990;Mazurier et al, 1992;Wang et al, 1993;Andersen e t al., 1996), 16s rRNA gene sequences (Stackebrandt & Charfreitag, 1990;Ash e t al., 1991a, b;Stackebrandt e t al., 1991 ;Aquino de Muro e t al., 1992), and PCR products with primers based on characterized genes (Gustafson, 1992;Darrasse e t al., 1994;Leite e t al., 1994;Schraft & Griffiths, 1995) have all provided phylogenetic information about members of the B. cereus group or other bacteria. We focused on the methods that we expected to have the greatest power to distinguish bacterial strains within a species.…”