Effect of phosphite on <i>in vitro</i> growth and zoospore production of <i>P. plurivora</i>.

<p>(a) Inhibition of <i>P. plurivora</i> mycelial growth using different phosphite concentrations. (b) <i>P. plurivora</i> cultures in Petri dishes illustrating the inhibition of mycelial radial growth with increasing phosphite concentrations. The mycelial colonies were six days old. (c) Inhibition of <i>P. plurivora</i> zoospore production at different phosphite concentrations. EC₅₀ shows the concentration that inhibits growth or zoospore production to 50%. Trend-lines were fitted using a logarithmic function. These assays were repeated three times showing similar results. n = 5 for each assay.</p

Effect of phosphite treatment and <i>P. plurivora</i> infection on the expression of defense-related genes.

<p>Absolute gene expression for PR1, PR2, PRP and WRKY (SA signaling pathway) as well as for PR3 and ACO (JA/ET signaling pathway) of beech saplings for all treatments. Purple and yellow bars represent control plants at time zero and after six days. Black and green bars represent phosphite treated plants at time zero and after six days. Blue bars represent infected plants at 6 dpi. Red bars represent phosphite-treated and infected plants at time 6 dpi. Asterisks show level of significance (P≤0.05*; P≤0.01**; P≤0.001***); N.S. Not-significant; n = 4. The experiment was repeated three times showing similar results.</p

Concentrations of phosphite and <i>P. plurivora</i> DNA along time.

<p>(a) Phosphite concentrations of root tissue. (b) qPCR analysis of <i>P. plurivora</i> DNA along time (ng/gDW). Red bars: Plants infected with <i>P. plurivora</i>; blue bars: Plants infected with <i>P. plurivora</i> and treated with phosphite (0.5%). Green bars: Plants treated with Phi. n = 6 plants per treatment. The experiments were repeated 3 times showing similar results. Dpi: days post inoculation. Different letters at the same time points show statistical differences (P≤0.05), N.S. =  Not-significant.</p

Effect of phosphite treatment and <i>P. plurivora</i> infection on plant mortality over time.

<p>Treatments: Con (black square): not phosphite treated and not inoculated control plants; Phi (green square): plants sprayed with phosphite (0.5%) on leaves until run off; Plu (red circle): roots infected with <i>P. plurivora</i>; Phi-Plu (blue triangle): plants sprayed with phosphite (0.5%) on leaves until run off four days prior to infection with <i>P. plurivora</i>. n = 6 plants per treatment. The experiment was repeated three times showing similar results. dpi: days post inoculation.</p

Effect of phosphite treatment on <i>P. plurivora</i> colonization of roots.

<p>Confocal laser scanning microscopy images of cross-sections of beech sapling roots infected with <i>P. plurivora</i> and either treated or not treated with phosphite after 2, 4 and 10 dpi (A, B, C). Beech sapling roots infected with <i>P. plurivora</i> and treated with phosphite (0.5%) after 2, 4 and 10 dpi (D, E, F). (Co) cortex, (CC) central cylinder, White bars represent 50 µm.</p

Effect of phosphite treatment and <i>P. plurivora</i> infection on net CO₂ assimilation rates (µmol CO₂ m⁻²s⁻¹) (a) and water uptake (b) over ten days.

<p>Water uptake was calculated as percent of g/cm² leaf surface of four months-old beech saplings. Treatments: Control: not infected and not phosphite treated plants; Phi: phosphite treatment; Plu: roots inoculated with zoospores of <i>P. plurivora</i>; Phi-Plu: foliar application of 0.5% phosphite on plants prior to inoculation with zoospores of <i>P. plurivora</i>. The experiment was repeated three times showing similar results. N = 6 plants per treatment. dpi- days post inoculation. Different letters at the same time points show statistical differences (P≤0.05), N.S. =  Not-significant.</p

Multiple sequence alignments showing similarity and conserved domains identified by the Interpro resource available at http://www.ebi.ac.uk/interpro/in A) Pep-13 transglutaminase peptide elicitors (according to Brunner et al., 2002 [52]), B) berberin-like proteins (Pfam PF08031), C) carbonic anhydrases (PROSITE PS51144; Pfam PF00194) and D) pectinesterase/pectate lyases (Pfam PF01095/SUPERFAMILY SSF51126).

<p>Signal sequences at N-terminus (SignP) and the consensus sequences are also reported.</p

List of effectors identified in the <i>P. plurivora</i> secretome by high resolution LC MS/MS.

<p>*Sequences containing the RXLR motif.</p><p>**Nucleotide sequence submitted to the EMBL/GenBank/DDBJ databases.</p><p>The secretion prediction according to signal peptide probability of Signal P 4.1 server is reported. Y and N indicate the presence or absence of the signal peptide for secretion, respectively.</p><p>List of effectors identified in the <i>P. plurivora</i> secretome by high resolution LC MS/MS.</p

Schematic representation of proteins with enzymatic activity enriched in <i>P. plurivora</i> secretome.

<p>The most represented categories for the oxidoreductases, transferases and hydrolases are also reported.</p