Occurrence of fruit stalk necrosis on <i>Pyrus bretschneideri</i> caused by <i>Alternaria alternata</i> in China

Pyrus bretschneideri Rehd. is an important pear species widely cultivated in northern China. Since June 2025, a previously unreported fruit stalk necrosis has been observed in commercial orchards of P. bretschneideri cv. Hongxiangsu in Linyi County, Shanxi Province, China (Danzizhen, Liucun village; 35.17°N, 110.59°E). The disease was characterized by brown to dark-brown necrosis restricted to the fruit stalk, with lesions extending internally along the peduncle, resulting in premature fruit drop. No visible symptoms were observed on fruit flesh or leaves (Fig. 1A). Disease incidence varied among trees within the orchard, and severely affected trees exhibited up to approximately 20% fruit drop. The affected trees were approximately 15 years old and had been top-grafted with ‘Hongxiangsu’. Notably, on the same tree, fruit stalk necrosis was observed only on ‘Hongxiangsu’, whereas the original cultivar ‘Suli’ remained symptomless under field conditions. To isolate the causal agent, symptomatic fruit stalk tissues were surface-disinfested in 75% ethanol for 30 s, rinsed twice with sterile distilled water, air-dried, and placed on potato dextrose agar (PDA). Plates were incubated at 28°C in the dark. A total of 18 fungal isolates with similar colony morphology were consistently obtained from five symptomatic fruit stalk tissues collected from two diseased trees. Colonies on PDA were gray to dark gray and produced abundant conidia. Conidia were formed in chains, obclavate to ellipsoid, with 1–6 transverse septa and 0–3 longitudinal septa, and measured approximately 18–45 × 7–15 μm (n = 30), consistent with the morphological characteristics of Alternaria alternata (Fig. 2A–G). Based on ITS sequencing and morphological characteristics, all 18 isolates were identified as members of the genus Alternaria, and no other fungal genera were recovered from the symptomatic tissues. For molecular identification, genomic DNA was extracted from representative isolates TG-1, TG-3, and TG-5. The internal transcribed spacer (ITS), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), translation elongation factor 1-alpha (TEF1-α), and Alt a1 genes were amplified and sequenced using primers and protocols described by Woudenberg et al. (2015). The obtained sequences were deposited in GenBank under accession numbers PX909743–PX909745 (ITS) and PX921101–PX921109 (GAPDH, TEF1-α, and Alt a1). Phylogenetic analysis based on concatenated sequences of the four loci was performed using MrBayes. The representative isolates clustered with reference strains of A. alternata with a posterior probability of 1.0, confirming their taxonomic placement (Fig. 2H). Pathogenicity tests were conducted on fruit stalks of both detached and attached pear fruits to fulfill Koch’s postulates. In three independent experiments, wounded fruit stalks were inoculated with mycelial plugs from actively growing cultures, whereas control fruit stalks were mock-inoculated with sterile PDA plugs. All inoculated fruit stalks developed necrotic symptoms identical to those observed under field conditions, whereas mock-inoculated controls remained asymptomatic, indicating that wounding alone did not induce the symptoms (Fig. 1B–F). The fungus was re-isolated from symptomatic tissues in each experiment, and the recovered isolates were identified as A. alternata based on morphological characteristics and sequence analysis, thereby fulfilling Koch’s postulates. Species of Alternaria, including A. alternata, have been reported to cause various diseases on pear, such as black spot, leaf spot, dead flower bud, and fruit diseases on Pyrus communis in several countries, including Italy, the Netherlands, and India (Wenneker et al. 2019; Prencipe et al. 2023; Kaur et al. 2025). However, fruit stalk necrosis associated with A. alternata on P. bretschneideri has not been previously documented. Under field conditions, fruit stalk necrosis was observed only on ‘Hongxiangsu’ scions, whereas the original cultivar ‘Suli’ on the same trees remained symptomless. This cultivar-specific occurrence warrants further investigation. To our knowledge, this is the first report of Alternaria alternata causing fruit stalk necrosis on Pyrus bretschneideri in China. This study provides a basis for accurate diagnosis and effective management of this newly observed disease in pear orchards.