First Report of Ceratocystis changhui Causing Postharvest Fruit Rot of Peach in Kunming, China.

Abstract

The peach (Prunus persica (L) Batsch) is a predominant commercially grown stone fruit in China (Lee et al. 1990). Ceratocystis changhui is an aggressive pathogen causing typical black rot symptoms on corms of taro (Colocasia esculenta) (Liu et al. 2018), it has not been reported on other hosts. During the summer and autumn of 2013, a postharvest fruit rot disease was observed on several peaches at a farmer's market (N 25°02'; E 102°42') in Kunming City, Yunnan Province, China. The incidence of the disease varied from 5 to 20%. Necrotic spots were first observed on the infected peach fruit (Prunus persica cv. shuimitao). The spots enlarged gradually and developed into a brown, water-soaked and rotted lesion. Eventually, the whole fruit became soft, rotted and covered with a gray-brown mycelium (Fig. 1 A, B). The isolates were obtained from the symptomatic tissues incubated on slices of fresh carrot root (Moller et al. 1968). After 5 to 10 days of incubation, perithecia and mycelium were observed growing on carrot slices. Spore masses were removed from the apices of perithecia, transferred to potato dextrose agar medium (PDA) and incubated at 25°C for 5 to 10 days, followed by single-spore isolation. All eight single-spore isolates from peach fruits obtained in this study were deposited in the State Key Laboratory for Conservation and Utilization of Bio-Resources in Yunnan, Yunnan Agricultural University, China. In culture, mycelium was initially white, gradually turned to greyish-green or brown (Fig. 1E, F). Measurements were made 7 days after the formation of perithecia. Perithecia (Fig. 1G) were black, globose, 185.71 to 305.56 μm × 142.86 to 264.29 µm and showed a long black neck, 600 to 957.14 µm (Fig. 1H). Ascospores (Fig. 1I) were helmet-hat shaped and 2.86 to 6.67 µm ×3.81 to 4.76 µm. Cylindrical conidia (Fig. 1J) 6.67 to 38. 95 µm × 2.86 to7.62 µm were observed. Chlamydospore (Fig. 1K), 8.57 to 13.33 μm × 5.71 to 9.52 μm, were ovoid or obpyriform, smooth. To further verify pathogen identity the internal transcribed spacer (ITS) region of rDNA was amplified using primers ITS1F and ITS4 (Thorpe et al. 2005), and the total genomic DNA from the mycelia of five isolates was extracted using a CTAB method (Lee &Taylor 1990). The nucleotide sequences have been blasted and deposited in the GenBank database. Analysis of the ITS sequences from the isolates T1-1yp, T1-2yp, T2-1yp (GenBank accession no. KY580895-KY580897) showed 99% to 100% similarity with isolates C. changhui CMW43272 (KY643886), CMW43281 (KY643884), CMW46112 (KY643891) and CMW46113 (KY643892) from taro in China. Phylogenetic trees based on the maximum-likelihood (ML) method were constructed using MEGA 7. ITS sequences of other Ceratocystis spp. were attained from NCBI for comparative analysis (Liu et al. 2018), and Davidsoniella virescens (CMW11164) served as outgroup. The robustness of ML tree was evaluated with 1,000 bootstrap (BS) values. The pathogen was identified as C. changhui based on the phylogenetic analysis (Fig. 2). Three isolates (T1-1yp, T1-2yp, T2-1yp) were used for pathogenicity. Nine Prunus persica cv. yingzuitao fruits at early maturity (8 points out of 10) were wound inoculated with 200μL conidia suspension of the fungus (approximately 2.0 × 106 conidia / mL). Degreasing cotton dipped in sterile water was used to raise the humidity in preservation boxes. Boxes were incubated for 10 days at 25°C. Three peaches as controls were treated only with sterile distilled water in the same way. Symptoms of sunken lesions and fruit rot were observed two days after inoculation, and measured at 1.8 to 3.2 cm from the inoculation point within 5 days (Fig. 1C: right, D). The same pathogen was re-isolated from them confirming Koch's postulates. Control peaches remained symptomless. This fungus was morphologically and phylogenetically identified as C. changhui. To our knowledge, this is the first report of C. changhui on postharvest peach in Yunnan, China. The disease will affect quality and taste of peach, so it is critical to deploy appropriate management strategies to limit the fungus spread.