Phoebe zhennan (Hemsl.) Yang is an evergreen broad-leaf, tall, grand tree with an open crown, dense foliage, and sweeping branches, commonly used for shade and landscaping in the urban areas of southern China. This tree is also suitable for hilly and mountainous areas, because it provides a good windbreak, anchors the soil, resists fire, and can also be used to build shelterbelts. A pest census by the Chinese Forestry Service in 2015 to 2017 identified a strain of branch wilt in the P. zhennan trees in Sichuan Province, China. This plant disease has since begun to seriously impact the economic value of P. zhennan, with a general incidence of 30 to 40% and a severe incidence of ≥60% in trees found in the Dujiangyan Park, and then spread to the rest of Sichuan Province with tree transplanting. The disease infects weak main branches, nursery stock, saplings, and even adult trees. The cortex of young P. zhennan branches becomes infected early during the progression. The infected region first exhibits a light chestnut brown color with oval-shaped bright spots that eventually, gradually expand. Once the lesions spread and encircle the branch, the branch wilts, its leaves drop, and it becomes bare. There is no obvious boundary between the infected and healthy parts of the tree, and many small black spots, representing pathogenic pycnidia, are scattered or clustered on the twigs. Severely infected P. zhennan trees possess many brittle, bare branches that tend to fall off in severe weather conditions. A number of infected branches were systematically sampled by cutting the epidermis at the interface of infected and healthy areas in 5 × 5-mm sections, surface sterilized with 3% NaClO and 75% alcohol for 30 s, rinsed three times with sterilized distilled water, and cultured on potato dextrose agar (PDA) (25°C for 4 to 7 days). The fungus formed nearly circular colonies with abundant aerial mycelia and changed from white to greyish black initially and black 3 weeks later. This fungus began to produce pycnidia on PDA medium after 3.5 weeks, and production of conidia was doubled after a 10-min UV irradiation and 3 days more of culture in comparison with culturing the fungus under dark conditions. The pycnidia were mostly spherically shaped, measuring 670 to 910 µm in diameter. The conidia were oval, unicellular with thin walls, measuring 15.2 to 17.2 × 4.6 to 6.4 μm in size. PCR was performed using DNA extracted from the fungal colony using universal primers (ITS1/ITS4), and then a fragment of approximately 572 bp was sequenced. Based on the sequence alignment and phylogenetic analyses in ClustalX and Mega 5.0, this fungus (accession no. MG386181) had the highest homology (99%) with Neofusicoccum parvum (accession no. FJ904915), which was previously found to cause branch and trunk cankers in grapevine, walnut, and peach trees in Sweden, Peru, California (U.S.A.), and Greece (Chen et al. 2014; Cheon et al. 2013; Phillips et al. 2013; Thomidis et al. 2011). To complete Koch’s postulates, N. parvum was cultured to produce microconidia on PDA for 7 days in 25°C. The aqueous conidial suspension was diluted to 1 × 10⁷ spores/ml with sterilized water, and 20 3-year-old potted P. zhennan seedlings were inoculated by scratching the stem epidermis with a piece of sandpaper and applying 100 µl of a microconidial suspension (1 × 10⁷ spores/ml) with a brush to each wounded area, and 20 seedlings inoculated with sterilized distilled water served as controls. The inoculated plants showed the same symptoms as the original diseased plants, and the controls remained asymptomatic 20 day after inoculation. The fungus was reisolated from the symptomatic stems and showed the same morphological characteristics as the isolates used for inoculation, thus confirming Koch’s postulates that N. parvum is the causal agent of branch wilt of P. zhennan. This is the first report of N. parvum causing wilt of P. zhennan trees in Sichuan province, China.
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Jun 3, 2024
Christopher J Geden + 3
Open Access
