Weinblattmetabolite als Resistenzmarker für eine Plasmopara viticola Widerstandsfähigkeit

Abstract

Downy mildew of grapevine is one of the most important diseases of the European grapevine Vitis vinifera Linné supsp. vinifera. It is caused by the obligate biotrophic oomycete Plasmopara viticola (Berk. & M.A. Curtis) Berl. & De Toni (1888), which was unintentionally introduced into Europe at the end of the 19th century. American grapevines are largely resistant to downy mildew and may contribute in one way or another to the protection of susceptible vines also against P. viticola.

Therefore, this work deals with the metabolite profiling of resistance markers (RM) from volatile secondary metabolites of susceptible and resistant grapevines. Ten genotypes with different resistance to P. viticola were analysed in field and greenhouse trials. Among them were three different Vitis species (V. vinifera, V. riparia, V. labrusca) and some hybrid vines. In a two-year trial, the constitutive markers of three developmental stages (BBCH6, BBCH8, BBCH9) were determined. In addition, induced markers were analysed in a third year. Furthermore, the relationship between leaf position and the occurrence of RM was investigated. The used metabolomic methods were also applied to identify markers for leaf position. For metabolite profiling the grape leaves were analysed using HS-SPME-GC-EI-MS and then were evaluated using non-targeted and targeted analytical methods.

The comparison of the metabolite profiles showed that the developmental stage has the strongest influence on the metabolite profile and the influence of the leaf position is so small that it can be neglected.

A total of 41 constitutive RMs were identified. The metabolites identified came from the substance classes of green leaf volatiles (GLV), norisoprenoids, benzoate derivatives, monoterpenoids, a furan and a sesquiterpene. It was elaborated, that GLV, norisoprenoids, benzoate derivatives and 2-ethylfuran were almost exclusively present in higher concentrations in resistant genotypes. Monoterpenoids and the sesquiterpene α-calacorene were mainly detected in higher concentrations in the susceptible genotypes of V. vinifera. Furthermore, it could be shown, that monoterpenoids were detected in significantly increased concentrations at the developmental stage BBCH6 in the susceptible V. vinifera. In contrast, the sesquiterpene α-calacorene was never detected in BBCH6. It was only found in significantly increased concentrations in the later developmental stages of V. vinifera. GLVs are RM in the resistant genotypes such as V. labrusca and occur here exclusively at the later developmental stages, mainly at BBCH9.

For the constitutive RMs from the classes of norisoprenoids, benzoate derivatives and furan, no dependence of the developmental stages and their occurrence was observed. However, three other metabolites were identified as RMs that showed a strong correlation with the developmental stage. They appeared initially in BBCH6 in significantly increased concentrations in susceptible V. vinifera, but in BBCH9 they showed significantly increased concentrations in the resistant V. labrusca genotypes. These RMs with changing correlation were the two monoterpenoids geranyl acetone and α-terpineol, as well as the GLV (Z)-3-hexenal. To the best of my knowledge, the correlation between occurrence of the constitutive RMs and the developmental stages has not been described before. It is a new finding of this work that monoterpenoids occur as RMs in grapevines mainly at BBCH6 and GLV as well as sesquiterpenes were found as RMs mainly at BBCH9. Norisoprenoids, benzoate derivatives and furan occurred as RMs at all developmental stages tested. Furthermore, monoterpenoids and the sesquiterpene α-calacorene occurred species-specifically more frequently in V. vinifera as constitutive RMs, whereas the furan 2-ethylfuran never appeared as a RM in V. vinifera. Benzoate derivatives and GLVs were most frequently detected as RMs in V. labrusca genotypes. Norisoprenoids appeared most frequently, but not exclusively, as RMs in V. riparia. To the best of my knowledge, the correlation between the occurrence of norisoprenoids as RMs and species specificity to V. riparia is a new finding of this work. Nevertheless, it should be briefly mentioned, that this species specificity refers exclusively to the classification as a constitutive RM. Monoterpenes are also part of the metabolite profile of V. labrusca as well as norisoprenoids are important flavour compounds in quality wines.

24 induced RMs were determined. Contrary to the original expectation, most of the induced RMs (19 out of 24) occurred in the fungus-resistant cultivar Regent and only six induced RMs in the resistant V. labrusca hybrid Blue Isabella and in V. riparia.

For all RMs identified, it was searched for reports on bioactivity in publications. For the RMs 2-ethylfuran, an isomer mix of (Z)- and (E)-ocimene as well as β-cyclocitral, an inhibitory effect on P. viticola has already been published. Still, 20 compounds have been associated here for the first time with a resistance response to P. viticola infection. These are the seven norisoprenoids theaspirane, (E)-β-damascone, (E)-β-damascenone, dihydroedulan I, megastigmatrienone, sulcatone, and carvomenthenal A, the two GLVs nonanol and (Z)-3-hexenyl acetate, and the ten terpenoids p-cymenene, p-cymene, D-limonene, alloocimene, β-myrcene, β-citronellol, hotrienol, (Z)-rose oxide, geranium oxide and α-calacorene.