Induction of defence responses and resistance to wheat leaf rust by plant extracts

[摘要] English: Information on the induced disease resistance mechanism in wheat against leaf rust (Pucciniatriticina) by two natural bio-stimulants (ComCat® and the seed suspension of Lupinus albus;SS) and two extracts with antifungal activity (Tulbaghia violacea and Agapanthus africanus)may be of great value both in designing new agrochemicals that stimulate plant resistanceresponses and in developing genetically engineered plants with enhanced disease resistance.The potential of these extracts to control leaf rust in vivo in susceptible (Thatcher) andresistant (Thatcher / Lr15) wheat was investigated. ComCat® and SS had no direct effectwhile the A. africanus extract resulted in the reduction of pustule and necrotic lesionformation in a susceptible and resistant wheat cultivar. T. violacea and A. africanussignificantly inhibited the germination of P. triticina spores and prevented further germ tubedevelopment.Foliar application of the different plant extracts on resistant infected wheat plants activatedβ-1,3-glucanase, chitinase and peroxidase enzyme activities. However, it was only theA. africanus treatment that increased the in vitro activities of these three apoplastic PRproteinssignificantly in both susceptible and resistant wheat cultivars, whether uninfected orinfected. As a result it was decided to concentrate the rest of the study on the A. africanusextract only.The induction pattern of apoplastic proteins from infected susceptible and resistant wheattreated with an A. africanus extract as well as a control treated with distilled water wasfollowed using SDS-PAGE. Clear differences between SDS-PAGE profiles of intercellularproteins from resistant and susceptible as well as untreated and treated plants were observedthroughout the 144 h period after treatment with the extract. In general, resistant plantscontained higher amounts of a 31 kDa protein and the protein was also present at muchhigher detectable levels in plants treated with the A. africanus extract. The molecular masscorresponded to that of β-1,3-glucanase. A Western blot using a polyclonal antibody againstβ-1,3-glucanase from wheat confirmed the identity of the 31 kDa protein to indeed be that ofβ-1,3-glucanase. This overwhelmingly excluded the A. africanus extract from the rest interms of its potent ability to induce a defence response in wheat towards leaf rust. RT-PCR was used in the analysis of the expression of the three defence related genes. Timecourseexperiments confirmed that they were induced in resistant as well as susceptible wheatafter infection. In this study, when resistant and susceptible wheat were treated with anextract of A. africanus 48 h prior to infection, a more pronounced induction of PR2, PR3 andPR9 gene expression occurred. Two different sized fragments were amplified when usingPR9 specific primers and both were induced by infection and by treatment with A. africanusextract in susceptible and resistant wheat. After sequencing, the larger fragment wasconfirmed to be peroxidase, while the smaller fragment shared very high sequence similarityto a retrotransposon gene. It can, therefore, be claimed that A. africanus is responsible for theinduction of PR genes and a retrotransposon gene in wheat.From the results obtained thus far, it was obvious that A. africanus must contain an activecompound(s) that act as an elicitor(s) in the mechanism of the defence reaction of wheatagainst leaf rust infection. Subsequently, activity directed isolation and purification of theactive compound lead to the isolation of a saponin, identified by means of 1H-NMR and 13CNMRspectroscopy as (25R)- 5α spirostane-2α, 3β, 5α-triol 3-O-{O-α-L-rhamnopyranosyl-(1α2)-O-[β-D-galactopyranosyl- (1α3)]- β-D-glucopyranoside}.

[发布机构] University of the Free State