Development of an isolate-specific marker for tracking Phaeomoniella chlamydospora infection in grapevines

Abstract

Petri disease causes decline of grapevines worldwide. The grapevine endophyte Phaeomoniella chlamydospora is the most important fungal pathogen associated with this disease. Epidemiological studies of this pathogen have been hampered by its common occurrence in the internal tissue of apparently healthy vines. Development of a molecular marker for a single strain would overcome this limitation and aid experiments designed to answer key questions about the biology of this pathogen. Genetic variation analysis of New Zealand and Italian strains of P. chlamydospora detected a potential molecular marker in New Zealand isolate A21. Characterization of the 1010 bp marker band showed that it had 50% identity to moxY, a gene involved in the aflatoxin biosynthetic pathway of Aspergillus parasiticus. Sequencing of the region flanking the 1010 bp product revealed a single nucleotide polymorphism in the 3' border of the marker band. Primers were designed to amplify a 488 bp fragment encompassing this polymorphic site and cleavage of this product with the restriction enzyme BsrI produced three bands only in isolate A21 and two bands in all other isolates tested. The sensitivity of the PCR-RFLP protocol was increased with a nested PCR approach and the protocol optimized for soil and wood samples. When the nested PCR/RFLP procedure was used to determine the persistence of viable and nonviable spores in soil, the results showed that nonviable spores were undetected after 8 wk whereas viable spores still could be detected at 17 wk.

Qualitative and quantitative molecular analysis indicate the presence of Phaeomoniella chlamydospora in vineyard soils

“…The need for adequate methodologies to detect Pch from soil has been pointed out repeatedly (Damm & Fourie, ). Nevertheless, molecular‐based studies to estimate Pch in soil have only been accomplished in artificially contaminated soils or in nursery soils (Retief et al, ; Ridgway et al, ; Whiteman et al, ). Overall, molecular studies that have attempted to correlate the presence of soil‐borne fungal species with crop decline phenomena, such as replant disease of horticultural and fruity crops are scarce (Franke‐Whittle, Manici, Insam, & Stres, ; Xiong et al, ; Xu, Ravnskov, Larsen, Nilsson, & Nicolaisen, ).…”

Application of <em>Trichoderma</em> to Prevent <em>Phaeomoniella chlamydospora</em> Infections in Organic Nurseries

“…Phaeomoniella chlamydospora tends to be isolated from cuttings more often than Phaeoacremonium spp. and is the most important fungal pathogen associated with Petri disease Fourie and Halleen, 2004;Gaforio et al, 2005;Whiteman et al, 2005;Ridgway et al, 2005;Retief et al, 2006).…”

Studies on the epidemiology of black-foot disease of grapevine in Spain

“…These results could indicate that Ca. luteo-olivacea and Pa. chlamydospora may be surviving in weeds as endophytes or latent pathogens until the moment in which grapevines are stressed (Edwards and Pascoe, 2004;Ridgway et al, 2005;Gramaje et al, 2010b). The potential role of these pathogens as grapevine endophytes and the erratic manifestation of leaf symptoms in infected grapevines have been previously reported by different authors Gramaje et al, 2010b).…”