First Report of Cobweb Disease Caused by Cladobotryum mycophilum on Cultivated Shiitake Mushroom (Lentinula edodes) in China

From the paper

Abstract

Lentinula edode (shiitake) is one of the highest-yielding edible fungi in China, widely appreciated for its unique rich flavor and medicinal properties (Chang and Miles 2004). During the winter of 2017, the production season of shiitake mushroom, cobweb disease symptoms were observed on shiitake mushroom in Qingyuan, Zhejiang province, China. The incidence of the disease varied between the affected farms, ranging from 60 to 80%. Typical symptoms initially started as the growth of white and thin mycelium over the fruiting bodies of shiitake mushroom. Later, the mycelium of the pathogen grew rapidly and gradually expanded to cover the whole surface of the fruiting bodies. Finally, the fruiting bodies were rotted and may be accompanied by a dark pink spot. Cobweb disease on shiitake has caused serious economic losses to the local mushroom production. Infected tissues of shiitake mushroom were disinfested by soaking in 75% alcohol for 45 s, rinsed three times in sterilized water, and placed onto potato dextrose agar (PDA) plates containing 100 mg/liter of streptomycin sulfate. The growth rate of the fungal colony was reached 22 mm/day on PDA at 25°C. The colony was composed of dense and cottony aerial mycelium. It appears grayish white in the early stages and slowly produces light yellow pigments within 6 days. After 9 to 13 days, the entire medium turns red. The cultures release no camphor odor. Chlamydospore was easier to be observed in the old aged mycelium. Conidiogenous cells were 20 to 33 μm long, 3.5 to 6.0 μm wide basally, with no evident rachis on the phialides. Conidia (n = 300) were transparent, single-celled, cylindrical to narrowly ellipsoidal with diaphragms, with zero to three septa and dimensions ranging from 10.0 to 31.0 × 8.8 to 11.5 μm. The morphological characteristics of the fungus were consistent with previous reports (McKay et al. 1999; Navarro et al. 2012). The total genomic DNA of the isolate (HD05) was extracted using the Nuclear Plant Genomic DNA Kit (CoWin Biosciences, China) following the producer’s protocol. Universal primers ITS4/ITS5 were used to amplify the internal transcribed spacer ITS 4-5.8S-5 DNA region. BLAST analysis showed highest similarity (99%) of the obtained ITS sequence with two sequences of Cladobotryum mycophilum (teleomorph Hypomyces odoratus) (GenBank accession nos. AB527074 and KJ942816) (Back et al. 2010; Zuo et al. 2016). Pathogenicity tests were performed by spraying spore suspensions (1 × 10⁶ conidia/ml) on fruiting bodies of shiitake mushroom. Spore suspensions were obtained from the isolate HD05 of C. mycophilum grown on PDA for 7 days at 25°C. Spores were collected and diluted with sterile distilled water at a concentration of 1 × 10⁶ conidia/ml. Nine bags of fruiting bodies were sprayed with 5 ml of the spore suspension, and another nine bags of fruiting bodies were sprayed with 5 ml of sterile distilled water as a control. A temperature of 20 to 22°C and relative humidity of 82 to 88% were maintained throughout cropping. The initial cobweb symptoms developed 5 days after inoculation. White-gray mycelium can be observed, and on the eighth day after inoculation, cobweb-like mycelium developed on stipes, caps, and primordia. The yield of diseased fruiting bodies was 39.8% of the total yield, and the yield of inoculated bags had a 43.2% loss compared with noninoculated bags, whereas the control treatments stayed healthy. Pathogenicity tests demonstrated that the symptoms of fruiting bodies sprayed with spore suspensions from the isolate were similar to those of the original symptomatic samples. C. mycophilum was consistently reisolated from the inoculated fruiting bodies (100%). Previously, C. dendroides has been reported causing cobweb disease on shiitake mushroom in Spain (Gea et al. 2018). However, to our knowledge, this is the first report of C. mycophilum causing cobweb disease on cultivated shiitake mushroom. This disease needs the attention of the local production department, to prevent further outbreaks of the disease.