First Report of Postharvest Fruit Rot in Solanum muricatum Aiton Caused by Alternaria alternata in southwest China.

Abstract

Solanum muricatum is native to South America and well known for its sweet, attractive, nutritious fruits. S. muricatum has been cultivated in China since the 1980s and increasingly popular (Li et al. 2015). In November 2021, an unknown fruit rot was observed in Shilin County of Yunnan Province (24.77 °N, 103.28 °E). The incidence of this disease was about 16% of 500 postharvest S. muricatum fruits after 7 d in storage room (25°C, 90% relative humidity). The initial symptoms were small brown spots on the fruit surface, which gradually expanded into irregular brown or black lesions, and gray-white mold developed in the center of the lesions, eventually the fruit turned rot. To isolate the pathogen, ten fruits with typical symptoms were collected and surface-sterilized with 75% ethanol for 45 s. Small fragments (5 × 5 mm) from the margin of lesions on fruit were disinfected with 1% sodium hypochlorite for 60 s, washed three times with sterile water then transferred to potato dextrose agar (PDA), and incubated at 28 ± 1℃ for 3 days (Li et al. 2022). Two fungal isolates with the same morphology were obtained and purified by single-spore isolation method. The colony was covered with thick fluffy aerial mycelia and the center was dark brown or black with white margins. Conidia were brown, pyriform or ellipsoid, with 1 to 3 longitudinal and 2 to 6 transverse septa, 15.12 to 34.01 × 6.90 to 12.73 μm (21.22 × 9.69 μm on average, n=50) in size. These morphological characteristics were consistent with Alternaria alternata (Li et al. 2015; Xiang et al. 2021; Alberto. 1992). For molecular identification, genomic DNA was extracted from a representative isolate, and primers ITS1/ITS4 (Gardes et al. 1993), TEF-F/TEF-R (Lawrence et al. 2013), Alt-F/Alt-R (Hong et al. 2005), GPD-F/GPD-R (Berbee et al. 1999) and EPG-F/EPG-R (Peever et al. 2004) were used to amplify the internal transcribed spacer (ITS), translation elongation factor 1-alpha (TEF), Alternaria major allergen (Alt a1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and endo-polygalacturonase (endoPG), respectively. The obtained DNA sequences (ITS: OM049821; TEF: OM069656; Alt a1: OM069655; GAPDH: OM069654 and endoPG: OM069653) showed over 99% homology with that of A. alternata (GenBank Accession No. MN856355.1 (565/573 bp); MN258023.1 (267/267 bp); KY923227.1 (491/501 bp); LC131645.1 (608/609 bp) and MN698284.1 (452/454 bp)). A phylogenetic tree based on the combined ITS, TEF, Alt a1, GAPDH, and endoPG sequences using the maximum likelihood methods with Kimura 2-parameter model, bootstrap nodal support for 1000 replicates in MEGA7.0 (Li et al. 2019) revealed that the isolate was assigned to A. alternata. To confirm pathogenicity, 10 μL spore suspension (1.0 × 106 conidia/ml) obtained from 7-day-old PDA cultures of each isolate were inoculated on 15 needle-wounded and 15 non-wounded surface-disinfected fruits, respectively. Healthy fruits were inoculated with sterile water as controls and the experiment was repeated 3 times. All fruit were incubated at 25 ± 1℃, 90% relative humidity. After 7 days, all the wounded and non-wounded fruit inoculated with A. alternata showed similar symptoms to those observed on the previously fruits, while the control fruits remained healthy. The same pathogen was again isolated from the inoculated fruits, thus Koch's postulates were fulfilled. A. alternata causing fruit rot of Prunus avium and Mangifera indica in China were reported in previous studies (Ahmad et al. 2020; Liu et al. 2019). As far as we know, this is the first report of postharvest fruit rot on S. muricatum caused by A. alternata in southwest China. This work provides a basis for the development of control strategies of the disease in the future.