Functional analysis of elicitins and identification of cell wall proteins in Phytophthora cinnamomi

Phytophthora cinnamomi Rands, a soil-borne oomycete plant pathogen, is a major threat to natural vegetation in many countries, especially Australia. The disease that it causes also has economic impacts on several agricultural and forestry systems worldwide. To investigate the virulence mechanisms of P. cinnamomi the roles of elicitins and elicitin-like proteins were examined using a susceptible model species, Lupinus angustifolius. Elicitins are 10 kDa proteins secreted by many Phytophthora species. Here, elicitins were isolated from P. cinnamomi liquid culture and separated according to their isoelectric point. A β-cinnamomin elicitin was then purified and used to develop an immunoaffinity purified antibody as a tool to examine virulence. The specificity of the β-cinnamomin antiserum was confirmed through Western-blot analysis. β-cinnamomin was found to be produced at different P. cinnamomi life stages and in inoculated plant roots by using immuno detection via confocal laser scanning microscopy. Pre-treatment of P. cinnamomi zoospores with β-cinnamomin antiserum and then inoculation of susceptible L. angustifolius roots revealed a partial loss of virulence of the pathogen. In addition, the expression of the β-cinnamomin gene at an early infection stage in susceptible L. angustifolius suggested an intrinsic role of the elicitin. Furthermore, the identification of covalently bound cell wall proteins in P. cinnamomi mycelia in this study has opened up the opportunity to investigate their role in virulence.