Inter- and Intra-Pathogen Interactions Emanating from Coinfection with Different Fungal and Viral strains in Canola Cultivars with Differing Host Resistances.

Few recent investigations examine coinfection interactions between fungal and viral plant pathogens. Here, we investigated coinfections between Leptosphaeria maculans and turnip mosaic virus (TuMV) in canola (Brassica napus). Different combinations of L. maculans isolate P11 and resistance breaking isolates L. maculans UWA192 and TuMV 12.1, were inoculated to three cultivars with differing pathogen resistances/susceptibilities. They were inoculated first to entire or half cotyledons 10-12 days after emergence, and second to opposite entire or half cotyledons on the same day (day 0) or 3 or 7 days afterwards. The parameters measured were L. maculans cotyledon disease index (%CDI), and TuMV systemically infected leaf symptom intensity (SI) and virus concentration (VC). Except when both day 0 inoculations were with isolate UWA192, %CDI values were supressed strongly or only weakly when isolates P11 and/or UWA192 were inoculated to plants with L. maculans single gene resistance (SGR) or polygenic resistance, respectively. However, except when isolate P11 was inoculated first and UWA192 second, these values declined after inoculation day 0 when SGR was absent. TuMV infection suppressed %CDI values, although this decrease was usually smaller following day 0 half cotyledon inoculations. When TuMV temperature sensitive extreme resistance was present and both inoculations were with TuMV, SI and VC values diminished greatly. However, the extent of this decrease was reduced when second inoculations were with L. maculans. SI and VC values were also smaller when SGR was present and second inoculations were with L. maculans. When L. maculans resistance was lacking, SI and VC values were smaller when second inoculations to entire cotyledons were with L. maculans rather than TuMV. This also occurred after second half cotyledon inoculations with isolate P11 but not isolate UWA192. Therefore, diverse inter- or intra-pathogen interactions developed depending upon host resistance, isolate combination, cotyledon inoculation approach and second inoculation timing.

Tomato CYCLOPS/IPD3 is required for mycorrhizal symbiosis but not tolerance to Fusarium wilt in mycorrhiza-deficient tomato mutant rmc.

Mycorrhizal symbiosis requires several common symbiosis genes including CYCLOPS/IPD3. The reduced mycorrhizal colonisation (rmc) tomato mutant has a deletion of five genes including CYCLOPS/IPD3, and rmc is more susceptible to Fusarium wilt than its wild-type parental line. This study investigated the genetic defects leading to both fungal interaction phenotypes and whether these were separable. Complementation was performed in rmc to test the requirement for CYCLOPS/IPD3 in mycorrhiza formation and Fusarium wilt tolerance. Promoter analysis via GFP expression in roots was conducted to determine the role of native regulatory elements in the proper functioning of CYCLOPS/IPD3. CYCLOPS/IPD3 regulated by its native promoter, but not a 2×35S promoter, restores mycorrhizal association in rmc. GFP regulated by the 2×35S promoter is not expressed in epidermal cells of roots, indicating that expression of CYCLOPS/IPD3 in these cells is required for colonisation by the fungi utilised in this research. CYCLOPS/IPD3 did not restore Fusarium wilt tolerance, however, showing that the genetic requirements for mycorrhizal association and Fusarium wilt tolerance are different. Our results confirm the expected role of CYCLOPS/IPD3 in mycorrhizal symbiosis and suggest that Fusarium tolerance is conferred by one of the other four genes affected by the deletion.

Proteome analysis of the Albugo candida-Brassica juncea pathosystem reveals that the timing of the expression of defence-related genes is a crucial determinant of pathogenesis.

White rust, caused by Albugo candida, is a serious pathogen of Brassica juncea (Indian mustard) and poses a potential hazard to the presently developing canola-quality B. juncea industry worldwide. A comparative proteomic study was undertaken to explore the molecular mechanisms that underlie the defence responses of Brassica juncea to white rust disease caused by the biotrophic oomycete Albugo candida. Nineteen proteins showed reproducible differences in abundance between a susceptible (RH 819) and a resistant variety (CBJ 001) of B. juncea following inoculation with A. candida. The identities of all 19 proteins were successfully established through Q-TOF MS/MS. Five of these proteins were only detected in the resistant variety and showed significant differences in their abundance at various times following pathogen inoculation in comparison to mock-inoculated plants. Among these was a thaumatin-like protein (PR-5), a protein not previously associated with the resistance of B. juncea towards A. candida. One protein, peptidyl-prolyl cis/trans isomerase (PPIase) isoform CYP20-3, was only detected in the susceptible variety and increased in abundance in response to the pathogen. PPIases have recently been discovered to play an important role in pathogenesis by suppressing the host cell's immune response. For a subset of seven proteins examined in more detail, an increase in transcript abundance always preceded their induction at the proteome level. These findings are discussed within the context of the A. candida-Brassica juncea pathosystem, especially in relation to host resistance to this pathogen.

Mycotoxins produced by Fusarium species associated with annual legume pastures and 'sheep feed refusal disorders' in Western Australia.

Sheep grazing in Western Australia can partially or completely refuse to consume annual Medicago pods contaminated with a number of different Fusarium species. Many Fusarium species are known to produce trichothecenes as part of their array of toxigenic secondary metabolites, which are known to cause feed refusal in animals. This study reports the identity of Fusarium species using species-specific PCR primers and a characterization of the toxigenic secondary metabolites produced by 24 Fusarium isolates associated with annual legume-based pastures and particularly those associated with sheep feed refusal disorders in Western Australia. Purification of the fungal extracts was facilitated by a bioassay-guided fractionation using brine shrimp. A number of trichothecenes (3-acetyldeoxynivalenol, deoxynivalenol, fusarenon-X, monoacetoxyscirpenols, diacetoxyscirpenol, scirpentriol, HT-2 toxin and T-2 toxin), enniatins (A, A1, B, and B1), chlamydosporol and zearalenone were identified using GC/MS and/or NMR spectroscopy. Some of the crude extracts and fractions showed significant activity against brine shrimp at concentrations as low as 5 µg ml(-1), and are likely to be involved in the sheep feed refusal disorders. This is the first report of chlamydosporol production by confirmed Fusarium spp.; of the incidence of F. brachygibbosum and F. venenatum in Australia and of F. tricinctum in Western Australia; and of mycotoxin production by Fusarium species from Western Australia.