Abstract
The Pseudomonas fluorescens complex of species includes plant-associated bacteria with potential biotechnological applications in agriculture and environmental protection. Many of these bacteria can promote plant growth by different means, including modification of plant hormonal balance and biocontrol. The P. fluorescens group is currently divided into eight major subgroups in which these properties and many other ecophysiological traits are phylogenetically distributed. Therefore, a rapid phylogroup assignment for a particular isolate could be useful to simplify the screening of putative inoculants. By using comparative genomics on 71 P. fluorescens genomes, we have identified nine markers which allow classification of any isolate into these eight subgroups, by a presence/absence PCR test. Nine primer pairs were developed for the amplification of these markers. The specificity and sensitivity of these primer pairs were assessed on 28 field isolates, environmental samples from soil and rhizosphere and tested by in silico PCR on 421 genomes. Phylogenomic analysis validated the results: the PCR-based system for classification of P. fluorescens isolates has a 98.34% of accuracy and it could be used as a rapid and simple assay to evaluate the potential of any P. fluorescens complex strain.
Highlights
The Pseudomonas fluorescens complex of species is one of the most diverse groups within the Pseudomonas genus, comprising more than fifty validly named species and many unclassified isolates
In order to identify markers that allow the classification of a given strain into one of the eight P. fluorescens phylogroups, all-againstall genome comparisons were performed on 71 P. fluorescens complex genomes (Supplementary Table 1) representing all these groups
The subsequent filtering for highly conserved coding sequences belonging to the core genome of each of the eight phylogroups revealed that the combination of nine markers in a pattern of presence and/or absence are enough for their discrimination (Table 1)
Summary
The Pseudomonas fluorescens complex of species is one of the most diverse groups within the Pseudomonas genus, comprising more than fifty validly named species and many unclassified isolates. More recently multilocus sequence analysis (MLSA) has divided the P. fluorescens complex into a varying number of groups depending on the number and identity of the genomes included in the study (Mulet et al, 2010; Gomila et al, 2015; Garrido-Sanz et al, 2016). These MLSA-based analysis have shown good concordance with phylogenomics and comparative genomics (Garrido-Sanz et al, 2016), in which eight phylogroups have been identified: P. mandelii, P. jessenii, P. koreensis, P. corrugata, P. fluorescens, P. gessardii, P. chlororaphis, and Pseudomonas protegens
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