First Report of Fruit Rot Caused by <i>Mucor inaequisporus</i> on Chinese Plum (<i>Prunus salicina</i>) in China

Plum (Prunus salicina Lindl.) is planted in many places in China. In July 2021, plums were found to be covered with unfamiliar yellow mycelia on the plum trees and on the ground in the campus of Northeast Agricultural University in Harbin. The symptoms on the fruit were initially manifested as white mycelium and water damage, then the mycelium turned yellow and the water immersion area expanded rapidly. Finally, the whole fruit is covered by yellow, dense mycelium, so that the flesh becomes soft rot, and fall off, causing about 3~5% economic loss (Fig. 1A, B). The thirty diseased fruits were collected, and the tissue (5 × 5 mm) was cut from the diseased fruits, and then surface sterilized with 75% alcohol for 30 s, soaked in 3% sodium hypochlorite for 5 min, washed with sterile distilled water for 3 times, and inoculated on potato dextrose agar (PDA) and malt extract agar (MEA) at 28 °C. Strains growing around the tissues from samples were subcultured on PDA, and 3 strains (DNMI01, DNMI02 and DNMI03) were obtained using the single-spore isolation method. On PDA, the hyphae were light yellow in the early stage, and became golden yellow in the later stage. The edge of the colony was fan-shaped, and the growth rate was extremely fast for 3 days to reach a full dish (Fig. 1G). On MEA, the mycelium was light yellow at the initial stage, and became grayish black in the later stage. The colony grew irregularly and grew slowly (Fig. 1H). Sporangiospores have two shapes, yellow, aseptate, globose or subglobose 7.7 to 14.1 × 8.5 to 13.9 μm (n=30), oval 2.5 to 6.4 × 6.2 to 12.0 μm (n=30, Fig. 1C). Columellae frequently pyriform, oval, colorless or yellowish, and 31.5 to 53.7 μm × 50.6 to 83.2 μm (n=30, Fig. 1D, E). Sporangia globose or subglobose, yellow, wall echinulate, and 55.3 to 109.1 × 67.9 to 123.5 μm (n=30, Fig. 1F) (Santiago et al. 2013). Acetyl trimethyl ammonium bromide method was used to extract DNA from 3-day-old. The internal transcribed spacer (ITS) and 28S rRNA partial gene sequences were amplified by primers ITS1 / ITS4 (White et al. 1990) and LR0R / LR5 (O'Donnell 1993), respectively. The sequences obtained by polymerase chain reaction (PCR) were sequenced and the sequences were deposited in GenBank with accession numbers ITS, OR136169-OR136171, and LSU, OR136179-OR136181. BLAST analysis of these sequences showed 96.13%-96.48% similarity to Mucor inaequisporus culture CBS255.36 (JN206177.1), 99.28%-99.58% to culture CBS255.36 (MH867301.1), respectively (Walther et al. 2013). Phylogenetic analysis by maximum likelihood method (MEGA7.0) generated based on the ITS, LSU sequences indicated that the strains formed a supported clade to the related M. inaequisporus type sequences. Strains was found to be most closely related to M. inaequisporus and far from other species. Based on morphological and phylogenetic characteristics, strains DNMI01 to DNMI03 was identified as M. inaequisporus (Lee et al. 2020; Ren et al. 2023). According to Koch 's postulate the pathogenicity test was conducted with healthy Prunus salicina fruits. Ten fruits were surface-disinfected with 75% alcohol for 15 s, rinsed with sterile water 3 times, air-dried, then were inoculated by spraying with a conidial suspension (1 × 106 sporongiospores·mL-1). Sterile water served as a control. The test was repeated 3 times. All fruits were cultured at 27°C. Symptoms typical developed on the fruits after 3 days, yellow hyphae covered the entire fruits, but control fruits remained healthy (Fig. 1I, J). The fungus was reisolated from symptomatic tissues and identified as M. inaequisporus by morphology characteristics and molecular characterization. To our knowledge, this is the first report of Fruit rot caused by M. inaequisporus on Chinese plum (Prunus salicina) in China.