Gnomoniopsis associated with necrosis of leaves and chestnut galls induced by Dryocosmus kuriphilus

Chestnut (Castanea spp.) is a very important crop in the Monti Cimini area (Viterbo, Italy) which is seriously infested with the key pest, chestnut gall wasp Dryocosmus kuriphilus Yasumatsu (Hymenoptera: Cynipidae) (Speranza et al., 2009). In May 2009 such seriously infested chestnut plants inthe Monti Cimini area were found with necrotic leaves and galls. Lesions on leaves were irregular and variable in size, lemon green to amber in colour with green margins (Fig. 1). Initially, galls were olive green then becamedark brown (Fig. 2). Inside the galls larvae of chestnut gall wasp were dead. Tissue from the edge of lesions was surface-sterilized using 1% sodium hypochlorite, rinsed in sterile distilled water and aseptically cut in 5-10 pieces not exceeding 5 x 5 mm, plated on potato dextrose agar (PDA) in Petri dishes containing an antibiotic solution (0.2% streptomycin sulphate) and incubated at 20 ± 3° C. Light brown fungal colonies were observed after one week close to the tissue (Fig. 2) and numerous orange slimy masses of conidia were observed on the culture surface. The colonies were regular in outline with a clear and thinner margin (Fig. 3). Conidia were oval to oblong, sometimes slightly obovoid, straight or curved and measured 5.5 - 8.0 µm x 2.0 - 3.0 µm (mean = 6.8 x 2.5 µm, n=210) (Fig. 4).

The fungus was identified at Centraalbureau voor Schimmelcultures (CBS, Utrecht, The Netherlands) as an undescribed species from the genus Gnomoniopsis Berl. Identification was based on both DNA sequencing (ribosomal ITS) and morphology (GenBank Accession Nos. EU254851, EU254854, EU254855). The pathogenicity of the fungus was tested in the Tuscia University experimental farm using freshly isolated cultures. Ten chestnut galls were inoculated, after slight pinprick injury, with an aqueous suspension containing 2 x 10⁶ spores/ml and incubated in clean cheesecloth covered with Parafilm. Control-injured galls were sprayed with sterile distilled water and incubated in the same manner. Control plants did not show any disease symptoms 60 days after inoculation. In contrast,the fungus produced characteristic blight symptoms on 90% of the inoculated galls. Similar blight symptoms were observed on chestnut twigs inoculated with 4 mm discs of seven-day-old PDA cultures. Isolations from these lesions confirmed the presence of a fungus identical to Gnomoniopsis. Although chestnut leaf blight has been reported to be caused by some species of the genus Gnomonia (Sogonov et al., 2008), this appears to be the first record of a fungus in the genus Gnomoniopsis associated with the necrosis of chestnut galls caused by Dryocosmuskuriphilus. This Gnomoniopsis sp. has not been reported in Italy on non gall-infested leaves of chestnut trees (Cristinzio, 1986). The relationship between the necrotrophic propensity of Gnomoniopsis and chestnut gall wasp infestation is under investigation.