First report of pear trees buds and twigs blight caused by Phoma glomerata in Iran

Due to the observation of a disease symptoms resembling fire blight in pear orchards of Mazandaran province, sampling of the symptoms of blight of buds took place through winter 2018-spring 2019. Samples from infected bud and blighted twigs were cut into 2- to 3-mm pieces, surface of the leaves was sterilized by 75% ethanol for 10 s followed by drowning 3 min in 0.5% sodium hypochlorite, rinsed three times with sterile distilled water, and cultured on water agar (WA) and potato dextrose agar (PDA) with 1.5% streptomycin and tetracycline. Petri dishes were incubated at 25°C. The cultures were initially pale brown and turned dark green with age. Embedded pycnidia were generally formed after 5 days. The pycnidia were agglutinating, globose to subglobose. Hyphal tip from the growing edge of colonies cultured for 3 days at 25°C was transferred to PDA to obtain pure cultures. The colonies were whitish initially and then became olive green to dark brown. Conidia were long, ellipsoid, single-celled, and hyaline or slightly pigmented. The fungus was identified as Phoma glomerata morphologicaly (Boerema et al., 2004). Koch's postulates was done for pathogenicity test. Symptom-free one year old plants and branches were inoculated with spore suspension (1×106 spores/ml). Sterile water was sprayed on another set of plants as non-inoculated control. Each inoculated branch was wrapped in a plastic bag and maintained in a greenhouse at two temperature ranges of 15 ± 2°C and 23± 2°C with 90% rational humidity. After 10 days, symptoms similar to references (Chohan and Chand, 1980) were observed and the same fungus (P. glomerata) was re-isolated. The species of the fungus was confirmed by extraction of genomic DNA from a single spore isolate (Amirdehi et al., 2017 and Doyle and Doyle, 1987). Then, ITS1, 5.8S, ITS2 were amplified with universal primers ITS1 (5'-TCCGTAGGTGAACCTGCGG-3') and ITS4 (5'-TCCTCCGCTTATTGATATGC-3') and maintained at 95°C for 3 min. Thirty eight cycles of PCR were performed by heating at 94°C for 1 min, 55°C for 1 min, and 72°C for 1 min, followed by a final period at 72°C for 10 min (White et al., 1990). The amplicon was sequenced and analyzed using BLAST software, and showed a homology of 99% with a corresponding sequence of Phoma glomerata (Corda) Wollenw & Hochapfel. The fungus was reported by Mathur (1979) on Ficus elastica for the first time from India. The fungus was also reported with symptoms of wilting and burning of the lateral buds of the pear (Pyrus communis) for the first time by Chohan and Chand (1980) from India. In Iran Phoma glomerata was reported from wheat Triticum aestivum L. and cucumber Cucumis sativus L. (Safaee et al., 2000; Hatami et al., 2008), Ficus elastica (Aghapour et al., 2009), Mandarin (Rostami et al., 2011), Rosemary (Moshrefi et al., 2015). Nevertheless, this species has not been observed on P. communis from Iran. This is the first report of bud and twig blight disease of Pear (P. communis) in Iran caused by the fungus Phoma glomerata.