First Report of Apple Bull’s Eye Rot Caused by Neofabraea malicorticis in Italy

Bull’s eye rot is an important postharvest disease affecting apples (Malus domestica) worldwide. Fruit infection occurs in the field, but rot lesions develop only after a period in cold storage (about 90 days). The disease is caused by fungi belonging to genus Neofabraea (N. vagabunda, N. malicorticis, N. perennans, and N. kienholzii) (Spotts et al. 2009). However, no data are available on the presence of species other than N. vagabunda in Italy. To verify the current causal agents of apple bull’s eye rot in the Emilia-Romagna region, a pathogen survey of apples stored in several local packinghouses and showing bull’s eye rot symptoms was carried out in 2014 and the causative agents identified. Small sections from the edges of rotten tissue of 100 fruits were placed on tomato agar (TA) and kept at 15°C. Sporulation was observed after 14 days of incubation. Mycelia originated from the single spore of 20 isolates were used for DNA extraction. PCR was used to amplify the β-tubulin (tub2) gene using primers F 5′-CTTTCTCCGTTGTCCCATCC-3′ and R 5′-GAACATTGCGCATCTGGTCC-3′ (Cao et al. 2013). After sequencing and sequence assembly, BLAST analysis showed that one isolate (M1) had 99% identity to N. malicorticis, while other isolates belonged to N. vagabunda. The sequence of N. malicorticis isolate was submitted to GenBank (accession no. KP263368). Morphological characteristics of N. malicorticis isolate were examined after 14 days of incubation on TA at 15°C. Colonies were circular, centrally umbonate, with abundant white floccose mycelium. Average growth rate was 22 ± 2 mm in 14 days. Conidia were one-celled, cylindrical, rounded at apex, aseptate, hyaline, and measured 6 to 16 μm (mean 10 ± 2.4 μm) × 2 to 5 μm (mean 3 ± 0.8 μm), similar to micro and intermediate conidia formed on oatmeal (Chen et al. 2015). To prove pathogenicity, Koch’s postulates were conducted on 10 ‘Cripps Pink’ apples, surface-sterilized, wounded, and inoculated with 20 μl conidial suspension (104 conidia ml–1). After 90 days of storage at 0°C, artificially infected apples showed lesions of 21 to 29 ± 1 mm diameter without conidiomata on peel surface (symptoms identical to those observed on apples that were originally obtained from cold storage). Water-only controls were symptomless. The pathogen was reisolated from decayed tissue. To our knowledge, this is the first report of N. malicorticis causing bull’s eye rot of apples in Italy. This finding suggests that, beside N. vagabunda, other species of Neofabraea may occur in Italy and this could represent an emerging problem on apples. Epidemiological studies performed on a wider scale could improve the knowledge of current Neofabraea species occurring in Italian orchards, leading to the adoption of specific control strategies to reduce fruit postharvest losses.