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First Report of Trichoderma crassum Causing Leaf Spot on Tomato (Solanum lycopersicum) in Ohio

    Affiliations
    Authors and Affiliations
    • Lijing Zhao1
    • Piao Yang1
    • Wenhui Li2
    • Zhenzhen Zhao1
    • Ye Xia1
    1. 1Plant Pathology Department, College of Food, Agricultural, and Environmental Sciences, Ohio State University, Columbus, OH 43210, U.S.A.
    2. 2College of Horticulture and Landscape Architecture, Northeast Agriculture University, Harbin 150030, China

    Trichoderma is a genus of wood-decaying fungi generally found in soil (Druzhinina and Kubicek 2005). Trichoderma crassum was confirmed to be a sister species to T. virens according to molecular sequencing results (Chaverri et al. 2003). A foliar disease with ∼70% incidence on Solanum lycopersicum was observed in a greenhouse at The Ohio State University (40°0′8″ N, 83°1′36″ W), Columbus, U.S.A., in December 2021. On average up to 60% of the leaves on 2-month-old tomato plants were infected. Initially, dark-gray, irregular spots appeared at the leaf tips. As the disease progressed, yellow necrotic lesions were observed surrounding the preformed disease spots. Finally, the infected leaves appeared curled and wilted as a whole. Leaf fragments from three tomato plants 40 inches apart were cut from the diseased lesions, surface sterilized with 75% ethanol (30 s) and 1% NaOCl (60 s), and rinsed with sterilized deionized water three times. Nine pieces of the sterilized leaf tissues were placed on PDA plates and incubated at 28°C in the dark in an incubator for 4 days. Pure cultures of five isolates were acquired and examined with a light microscope. The fungus from all the isolates changed from white to dark green with a radial pattern and profuse sporulation on PDA. Round conidia were observed under a light microscope. DNA was extracted from two representative isolates, which showed the same morphology. The internal transcribed spacer (ITS) region and a conserved fungal rRNA region were amplified using the primers ITS1/ITS4 (5′-TCCGTAGGTGAACCTGCGG-3′ and 5′-TCCTCCGCTTATTGATATGC-3′) (White et al. 1990) and SR6f/SR7r (5′-TGTTACGACTTTTACTT-3′ and 5′-AGTTAAAAAGCTCGTAGTTG-3′) (Hirose et al. 2012), respectively. The PCR products were further sequenced by Sanger sequencing. Based on the BLAST results through NCBI, the ITS sequences of the two isolates were 99% (566/572) and 98% (558/572) identical to T. crassum DAOM 164916 (EU280067). Their SR sequences both showed 99% (290/293; 289/293) identity to the same strain. A phylogenetic tree was also created with the sequences of the ITS region by MEGA software (version 11). Therefore, the fungus was identified as T. crassum based on its morphological characteristics (green conidia), Sanger sequencing results, and a phylogenetic tree. To complete Koch’s postulates, 5-mm-diameter fungal agar discs of 7-day-old pure cultures were used for the inoculation of 18 healthy leaves of six tomato cv. M82 plants that were 2 months old. Sterile pure PDA discs of equal size were used for the mock inoculation as a comparison. The fungal plug method was chosen in this study because it had been widely applied for characterization of the fungal pathogens causing leaf spot disease (Pornsuriya et al. 2020; Yang et al. 2021). Five days later, the same symptoms as those that occurred on the previously naturally infected tomato plants were observed on all the inoculated leaves. However, there were no symptoms on leaves from the mock inoculation. The fungus reisolated from the symptomatic leaves showed consistent morphology (dark green with radial sporulation) to the original isolates. Thus, T. crassum was verified as the causal agent of the foliar disease on S. lycopersicum cv. M82 in our greenhouse. To our knowledge, this is the first report of T. crassum leading to leaf spot and wilt on tomato in Ohio. The identification of the causal agent lays the groundwork for the development of necessary disease management techniques.

    The author(s) declare no conflict of interest.

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    Funding: We acknowledge the funding support from CFAES Internal Grants Program 2021009.

    The author(s) declare no conflict of interest.