Thierry Barchietto

Dissecting Phosphite-Induced Priming in Arabidopsis Infected with Hyaloperonospora arabidopsidis

Phosphite (Phi), a phloem-mobile oxyanion of phosphorous acid (H3PO3), protects plants against diseases caused by oomycetes. Its mode of action is unclear, as evidence indicates both direct antibiotic effects on pathogens as well as inhibition through enhanced plant defense responses, and its target(s) in the plants is unknown. Here, we demonstrate that the biotrophic oomycete Hyaloperonospora arabidopsidis (Hpa) exhibits an unusual biphasic dose-dependent response to Phi after inoculation of Arabidopsis (Arabidopsis thaliana), with characteristics of indirect activity at low doses (10 mm or less) and direct inhibition at high doses (50 mm or greater). The effect of low doses of Phi on Hpa infection was nullified in salicylic acid (SA)-defective plants (sid2-1, NahG) and in a mutant impaired in SA signaling (npr1-1). Compromised jasmonate (jar1-1) and ethylene (ein2-1) signaling or abscisic acid (aba1-5) biosynthesis, reactive oxygen generation (atrbohD), or accumulation of the phytoalexins camalexin (pad3-1) and scopoletin (f6′h1-1) did not affect Phi activity. Low doses of Phi primed the accumulation of SA and Pathogenesis-Related protein1 transcripts and mobilized two essential components of basal resistance, Enhanced Disease Susceptibility1 and Phytoalexin Deficient4, following pathogen challenge. Compared with inoculated, Phi-untreated plants, the gene expression, accumulation, and phosphorylation of the mitogen-activated protein kinase MPK4, a negative regulator of SA-dependent defenses, were reduced in plants treated with low doses of Phi. We propose that Phi negatively regulates MPK4, thus priming SA-dependent defense responses following Hpa infection.

Characterization of phosphonate uptake in two Phytophthora spp. and its inhibition by phosphate

The uptake of the phosphonate ion, the active breakdown product in plant tissues of the systemic anti-Oomycete compound Fosetyl-Al (aluminium tris-Oethylphosphonate), was investigated in two Phytophthora spp. of differential sensitivity. Uptake was due to the simultaneous operation of two transport systems, one of low affinity (high Km) and one of high affinity (low Km). The relative contribution of each transport system varied with the external concentration of phosphonate, suggesting that phosphonate was a potent regulator of both systems. Phosphate was a partial competitive inhibitor with respect to phosphonate. Phosphate competed with phosphonate for uptake with a Ki of 105 μM for P. cryptogea and 68 μM for P. citrophthora. Uptake was sensitive to pH, showing a maximum at pH 5.0 to 5.5. P. cryptogea was more efficient in phosphonate uptake, although it was less sensitive to inhibition by phosphonate in vitro, than P. citrophthora. This implied that the selective activity of phosphonate was not due to differential rates of uptake of this oxyanion. These results were discussed in relation to the mode of action of phosphonate towards Oomycetes.

Effects of phosphonate on the elicitor activity of culture filtrates of Phytophthora cryptogea in Vigna unguiculata

Abstract When grown in liquid medium of Huang et al., Phytophthora cryptogea produces elicitor active component(s) which causes necrosis, accumulation of the phytoalexins kievitone and phaseollidin and electrolyte leakage from cowpea leaf tissues. Elicitor activity of culture filtrates is enhanced upon treatment of the fungus with 2.44 mM phosphonate, a dose which slightly effects fungal growth in vitro. The necrosis-inducing activity of culture filtrates is diminished after heating, treatment with pronase and protease, whereas active substance(s) is periodate-sensitive. The fact that phosphonate improves fungal cells in vitro to overproduce compound(s) which elicits a resistance response, support the hypothesis that the toxophore may act indirectly in vivo interfering with the processes of pathogenicity.

Modification of the phosphite induced resistance response in leaves of cowpea infected with Phytophthora cryptogea by α-aminooxyacetate

Abstract Treatment of detached cowpea leaves with phosphite, the active breakdown product in plant tissues of fosetyl-Al, leads to the cessation of growth of Phytophthora cryptogea within 24 h of inoculation. Pretreatment of leaves with α-aminooxyacetate (AOA), an inhibitor of the phenylpropanoid pathway, increases the size of lesions in phosphite treated leaves and induces a complete susceptibility at 5 mM. By 24 h after inoculation, phenylalanine ammonia-lyase (PAL) activity is higher in phosphite treated leaves than in untreated leaves. The effects of AOA on PAL activity are paralleled with the effects of the increase of the spread of infection. AOA treatment does not affect phosphite uptake by fungal cells and leaf tissues, while it inhibits kievitone and phaseollidin accumulation in infected treated leaves and causes a reduction in extractable PAL activity. These results suggest that cessation of fungal growth in vivo is not as a result of a direct effect of phosphite on the fungus and strongly support a role of the host defence reactions in the mode of action of phosphite.