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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.

Phytophthora capsici, a highly dynamic and destructive oomycete pathogen, causes devastating diseases on a wide range of plants worldwide. However, the detailed molecular mechanisms of pathogenicity is still largely unclear. In this study, three different mRNA pool libraries were constructed from its developmental stage, early or late infection stage of the model plant Arabidopsis thaliana, and then were investigated by the RNA-Seq approach. The results demonstrated that 1456 novel transcripts that had not linked to any annotated gene were identified, and 296 genes were found to undergo alternative splicing. Comparative analysis of three different libraries further showed that distinct transcriptional changes of pathogenicity genes were found. A large number of genes containing cell wall degrading enzymes, major facilitator superfamily genes and cytochrome P450 genes were highly induced during infection. In addition, several types of well-known effectors including RxLR, CRN, Elicitin and NLP proteins also showed high transcript abundances during infection. The transcriptional levels of six effector genes during the infection process were further validated by qRT-PCR. Collectively, this study provides a basic understanding of pathogenic mechanisms of P. capsici during the interaction with plants.

Phytophthora ramorum populations are clonal and consist of three lineages. Recent studies have shown that the clonal lineages may have varying degrees of aggressiveness on some host species, such as Quercus rubra. In this study, we examined virulence, sporulation and elicitin production of five P. ramorum isolates from each of the three clonal lineages. Virulence (lesion size) and sporulation (sporangia production) were determined on wound-inoculated detached leaves of Rhododendron catawbiense ‘Nova Zembla’. Lesion area differed between the clonal lineages (p

The differential expression of four Phytophthora cinnamomi elicitin genes was analysed by Real Time RT- PCR. In in vitro cultures, the a-cinnamomin gene showed the highest level of expression, the b-cinna- momin gene (b-cin) was the most inducible, and the HAE transcripts were in low abundance. Tran- scription of all the elicitins was active during the active growth of the pathogen when infecting cork oak (Quercus suber) roots, and as host colonization progressed, the level of b-cin expression fell, while that of a-cin rose. In an antisense transgenic strain, the silencing of b-cin also negatively affected the expression of other elicitin genes in the cluster. The reduction in planta growth of the b-cin knock-out is related to the altered pattern of elicitin gene expression, supporting the idea that one of the functions of elicitins is related, directly or indirectly, with pathogenesis. 2009 Published by Elsevier Ltd.

Elicitin and a new protein 75 kDa elicitor were purified from the culture filtrate of Phytophthora palmivora, a pathogen of Hevea brasiliensis (rubber plant). Elicitin was obtained by using a one step of DEAE cellulose chromatography and the new elicitor was obtained by two steps of chromatography: a DEAE cellulose column followed by a hydrophobic column. Both elicitors were stable to heat and a wide range of pH values, but were sensitive to ProteaseK. Both elicitors induced scopoletin, peroxidase isozymes (with substrate o-dianisidine and scopoletin) and total phenolic compounds in cell suspension of H. brasiliensis with similar kinetics. In addition, both elicitors induced peroxidase enzyme (o-dianisidine), total phenolic compounds and enhanced local resistance against P. palmivora on young rubber tree seedlings. However, the increase of peroxidase enzyme and total phenolic compounds in rubber tree seedlings was different from those in cell suspension. Furthermore, during the expression of local resistance the zoospore of P. palmivora induced the peroxidase enzyme (o-dianisidine) more rapidly and with higher level than the control plants. H. brasiliensis is more responsive to the new elicitor than elicitin in triggering defense responses. That is the new elicitor was active at a concentration lower than those required for elicitin, about a 30-fold decrease for activation defense responses in cell suspension. For induction of peroxidase enzyme (o-dianisidine), phenolic compounds and local resistance of rubber plants against P. palmivora, the 75 kDa protein was active at about a 2-fold lower concentration when compared to elicitin.

We report on the growth pattern of the root rot pathogen Phytophthora quercina in roots of oak seedlings and on the localization of its elicitin quercinin. Some hours after infection, the pathogen was mainly found in the intercellular spaces of the cortical parenchyma. Some parenchyma cells were also invaded by P. quercina and the plasmalemma of those cells as well as of neighbouring cells separated from the cell wall. Transmission electron microscopy studies proved that P. quercina penetrated the cell walls forming haustorial-like structures in invaded cells. These structures and cell wall appositions were covered with electron dense material. Immunofluorescence investigations in combination with a specific anti cryptogein antibody clearly showed that the P. quercina elicitin quercinin was located within the hyphal cell wall and seems to be released into invaded cells. In addition, immunogold labelling showed that quercinin was found in the intercellular spaces as well as in penetrated cells. Finally, we demonstrated with ELISA that quercinin was produced by the pathogen in infected tissue during the whole growth phase.

The Phytophthora spp. tested produced several elicitin isoforms with different toxicities to tobacco. A general chromatographic procedure is described which can be used to isolate and purify isoforms in large amounts. α Elicitins are acidic molecules with a hydrophobic domain around position 13, while β elicitins are basic with a hydrophilic domain at this position. The molecular weights, isoelectric points, amino acid compositions and N-terminal sequences of these molecules are reported. Their molecular properties, including their unique peculiar chromatographic behaviour, correlated with their level of toxicity to tobacco. Elicitins were shown to have nonspecific toxicity to other solanaceous plant species. The toxicity classification established with tobacco was different for the other plant species.