First report of the fungus Gnomoniopsis smithogilvyi causing cankers on sweet chestnut (Castanea sativa) in Ireland

Abstract

In autumn 2020 cankers and branch dieback (Figure 1) were found in a five-year-old gene bank plantation of Castanea sativa in Wicklow, Ireland. The planting material had been imported as grafted saplings from the United Kingdom. Isolations were made from the cankers by surface sterilising the outer bark near the cankers with 70% ethanol. Pieces (5 × 5 × 5mm) of woody material were removed and soaked in 2% NaOCl for one minute, followed by two one-minute distilled water washes. After dabbing dry using paper towels the pieces were placed aseptically onto the surface of malt extract agar (MEA; Difco™) and incubated at 21°C in the dark for three weeks. After several days, hyphal tips were taken from the fungal colonies and transferred onto 2% potato dextrose agar (PDA; DifcoTM). DNA was extracted from the mycelium of seven-day old cultures and the ITS region was sequenced using primers ITS5 and ITS4. Gnomoniopsis smithogilvyi (syn. G. castaneae) (Figure 2) was detected. Microscopic observations agreed with the published descriptions of Shuttleworth et al. (2012) and Visentin et al. (2012). The ITS sequences were deposited in GenBank under the accession numbers OM683278 and OM683279. Sixty three-year-old C. sativa saplings were inoculated with either of two G. smithogilvyi isolates, or a control sterile PDA plug (Table 1). After surface sterilising with 70% ethanol, a stem wound was made using a sterile 6 mm cork borer, 10 cm above soil level. A 6 mm mycelial plug of a three-week-old isolate of G. smithogilvyi was placed onto the wound. This was then covered in moist sterilised cotton wool, wrapped in plastic film, and covered by aluminium foil. Control plants were inoculated with sterile PDA plugs. Treatments were randomised and grown outside, between mid-August to mid-November (mean temperature 14.9°C; mean relative humidity 85.5%). After six weeks dark necrotic lesions (Figure 3; Table 1) were observed on all saplings which had been inoculated with G. smithogilvyi isolates, and G. smithogilvyi was re-isolated from all lesions. Gnomoniopsis smithogilvyi caused girdling on numerous saplings (Figure 4). The identity of the reisolated colonies was confirmed as G. smithogilvyi by morphological assessment and ITS sequencing, fulfilling Koch's postulates. All control saplings were found to be asymptomatic, and no G. smithogilvyi was isolated from these (0/8 attempted re-isolations). Gnomoniopsis smithogilvyi causes cankers on C. sativa hybrids in Spain, and on C. sativa in five European countries (Shuttleworth et al., 2016; Lewis et al., 2017; Lione et al., 2019). It also causes chestnut rot in C. sativa in Australia, France, Italy, New Zealand, Portugal and Switzerland (Visentin et al., 2012; Shuttleworth et al., 2016; Coelho & Gouveia, 2021). This is the first report of G. smithogilvyi causing cankers on C. sativa in Ireland. Follow up surveys and tests are planned to fully understand the distribution of G. smithogilvyi in Ireland.