Presence/absence of putative homolog proteins in nine pseudomonad strains infecting <i>Corylus avellana</i> trees.

<p>The indication concerns the presence of copper resistance genes, bacteriocins, and the flagellin cluster (A), and ice nucleation activity, pectic enzymes, sucrose metabolism, and siderophores (B). Strain legend is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0131112#pone.0131112.t001" target="_blank">Table 1</a>.</p

Divergence dendrogram based on the concatenated dataset of the four housekeeping genes.

<p>The dendrogram was built assessing <i>gapA</i>, <i>gltA</i>, <i>gyrB</i>, <i>rpoD</i> genes and by applying the <i>E</i>. <i>coli-Salmonella</i> model to nine pseudomonad strains infecting <i>Corylus avellana</i>. MYA: millions of years before present. Strain legend is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0131112#pone.0131112.t001" target="_blank">Table 1</a>. <i>Pseudomonas cannabina</i> pv. <i>alisalensis</i> Pcal BS91 was included in the analysis as outgroup.</p

Recombination networks regarding four housekeeping genes of the nine pseudomonad strains infecting <i>Corylus avellana</i> trees.

<p>The networks were built using <i>gapA</i>, <i>gltA</i>, <i>gyrB</i>, <i>rpoD</i> genes. Strain legend is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0131112#pone.0131112.t001" target="_blank">Table 1</a>. The scale bar indicates the number of substitutions per nucleotide position. Strong signal of recombination are present solely between the <i>Pseudomonas syringae</i> pathovars. <i>P</i>. <i>cannabina</i> pv. <i>alisalensis</i> Pcal BS91 was included in the analysis as outgroup.</p

Split consensus network obtained from 2.812 trees for the nine pseudomonad strains infecting <i>Corylus avellana</i>.

<p><b>A.</b> The tree has a cut-off value of 0.19. The scale bar indicates the number of substitutions per nucleotide position. Strain legend is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0131112#pone.0131112.t001" target="_blank">Table 1</a>. No reticulation was observed between the <i>Pseudomonas avellanae</i> and <i>P</i>. <i>syringae</i> pathovar strains. <b>B</b>. Neighbor-net tree obtained from 2.812 core gene sequence alignments. The dendrogram (i.e., a total of 1.977.984 nucleotides) regards the nine pseudomonad strains infecting <i>Corylus avellana</i> trees. The scale bar indicates the number of substitutions per nucleotide position. Bootstrap values are shown at the main nodes. No reticulation was observed between the <i>Pseudomonas avellanae</i> and <i>P</i>. <i>syringae</i> pathovar strains. In both trees, <i>P</i>. <i>cannabina</i> pv. <i>alisalensis</i> Pcal BS91 was included in the analysis as outgroup. Strain legend is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0131112#pone.0131112.t001" target="_blank">Table 1</a>.</p

Comparative Genomic Analyses of Multiple <i>Pseudomonas</i> Strains Infecting <i>Corylus avellana</i> Trees Reveal the Occurrence of Two Genetic Clusters with Both Common and Distinctive Virulence and Fitness Traits

<div><p>The European hazelnut (<i>Corylus avellana</i>) is threatened in Europe by several pseudomonads which cause symptoms ranging from twig dieback to tree death. A comparison of the draft genomes of nine <i>Pseudomonas</i> strains isolated from symptomatic <i>C</i>. <i>avellana</i> trees was performed to identify common and distinctive genomic traits. The thorough assessment of genetic relationships among the strains revealed two clearly distinct clusters: <i>P</i>. <i>avellanae</i> and <i>P</i>. <i>syringae</i>. The latter including the pathovars <i>avellanae</i>, <i>coryli</i> and <i>syringae</i>. Between these two clusters, no recombination event was found. A genomic island of approximately 20 kb, containing the <i>hrp/hrc</i> type III secretion system gene cluster, was found to be present without any genomic difference in all nine pseudomonads. The type III secretion system effector repertoires were remarkably different in the two groups, with <i>P</i>. <i>avellanae</i> showing a higher number of effectors. Homologue genes of the antimetabolite mangotoxin and ice nucleation activity clusters were found solely in all <i>P</i>. <i>syringae</i> pathovar strains, whereas the siderophore yersiniabactin was only present in <i>P</i>. <i>avellanae</i>. All nine strains have genes coding for pectic enzymes and sucrose metabolism. By contrast, they do not have genes coding for indolacetic acid and anti-insect toxin. Collectively, this study reveals that genomically different <i>Pseudomonas</i> can converge on the same host plant by suppressing the host defence mechanisms with the use of different virulence weapons. The integration into their genomes of a horizontally acquired genomic island could play a fundamental role in their evolution, perhaps giving them the ability to exploit new ecological niches.</p></div

Average nucleotide identity (ANIm) values calculated between genomes of nine <i>Pseudomonas</i> strains causing disease to <i>Corylus avellana</i> trees (see also Table 3).

<p>Average nucleotide identity (ANIm) values calculated between genomes of nine <i>Pseudomonas</i> strains causing disease to <i>Corylus avellana</i> trees (see also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0131112#pone.0131112.t003" target="_blank">Table 3</a>).</p

Venn diagram of the type III secretion system effector genes complement of nine pseudomonad strains infecting <i>Corylus avellana</i> trees.

<p>Pave: <i>Pseudomonas avellanae</i>; Psave: <i>P</i>. <i>syringae</i> pv. <i>avellanae</i>; Psc: <i>P</i>. <i>s</i>. pv. <i>coryli</i>; Pss: <i>P</i>. <i>s</i>. pv. <i>syringae</i>. *: not present in <i>P</i>. <i>avellanae</i> BPIC 631.</p

Maximum-likelihood tree based on the concatenated sequences of four housekeeping genes.

<p>Dendrogram based on MLSA analysis of <i>gapA</i>, <i>gltA</i>, <i>gyrB</i> and <i>rpoD</i>, for a total of 6.812 nucleotides regarding <i>Pseudomonas avellanae</i> and <i>P</i>. <i>syringae</i> pathovars <i>avellanae</i>, <i>coryli</i> and <i>syringae</i> strains infecting <i>Corylus avellana</i> trees. The horizontal lines show the genetic distance. The numbers at the node are support value estimated with 100 bootstrap replicates. Only bootstrap values > 75 are indicated. The scale bar indicates the number of substitutions per nucleotide position. <i>P</i>. <i>cannabina</i> pv. <i>alisalensis</i> Pcal BS91 was included in the analysis as outgroup. Strain legend is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0131112#pone.0131112.t001" target="_blank">Table 1</a>.</p

Structure of the genomic island Pcor GI1 of about 20 kb found in all nine pseudomonad strains infecting <i>Corylus avellana</i> trees.

<p>It was obtained using the SEED viewer comparative tool. The genomic structure of the island resulted conserved among the strains. Two representative structure regarding <i>Pseudomonas syringae</i> pv. <i>syringae</i> CRAFRU 11 and <i>P</i>. <i>avellanae</i> BPIC 631 are shown. Gene legend is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0131112#pone.0131112.s010" target="_blank">S2 Table</a>.</p

General features of draft genomes for the phytopathogenic <i>Pseudomonas</i> strains causing infection to <i>Corylus avellana</i> trees assessed in this study.

<p>Psave CRAPAV 013 and Psave CRAPAV 037 are the same strains previously named as ISPaVe 013 and ISPaVe 037, respectively.</p><p>^T: pathotype strain.</p><p>General features of draft genomes for the phytopathogenic <i>Pseudomonas</i> strains causing infection to <i>Corylus avellana</i> trees assessed in this study.</p