Pseudonectria buxi (also called Volutella buxi) is a fungal pathogen that causes a disease of boxwood (Buxus spp.). This disease has been reported in several countries in Europe and North America, and has been traditionally considered the primary cause of boxwood decline (1), although box blight caused by Cylindrocladium buxicola has claimed notoriety because of its recent introduction to North America. In August 2013, symptoms resembling Volutella blight were observed at a park in Haidian District, Beijing, China, on leaves and stems of Korean boxwood (B. sinica var. insularis). The plants were still alive, but diseased leaves and twigs were yellowed and showed dieback. Symptoms were common on boxwood throughout this park, and the disease was also seen in other nearby areas. Pink sporodochia were observed on some yellowed leaves and stems. Diseased tissues (stems and leaves) were collected, cut into 1 mm2 pieces, surface sterilized in 1% sodium hypochlorite for 1 min, and placed on potato dextrose agar. After 3 days of incubation at room temperature, white fluffy mycelia were seen. The middle of the colonies turned pink by 7 days, and conidia produced on verticillately branched conidiophores in these pink areas were elliptical, 6 to 9 × 2 to 2.5 μm. DNA was extracted from one colony containing spores and mycelium, and a portion of the beta-tubulin gene was amplified using primers designed from highly conserved regions (5′-AACAACTGGGCCAAGGGTC, 5′-GAAGAGTTCTTGTTCTGGA) (3). The 676-bp amplicon was sequenced (GenBank Accession No. KJ755987), and the top matches were two isolates of P. buxi (KC819609 and DQ522522) with identities of 566/567 bp (99.8%) and 551/567 bp (97.2%), respectively. The next best matches were at 92% for Fusarium spp. For inoculation, four pots of 2-year-old healthy B. sinica var. insularis × B. sempervirens cv. Green Velvet were used. As wounds are required for the infection process (2,3), two parallel light scratches were made using needles on adaxial surface of three leaves per plant. One plant was only sprayed with water until runoff, while the other three plants were sprayed with a spore suspension (106 spores/ml) of P. buxi until runoff, covered with plastic bags, and placed at 25°C. After 3 days, pink sporodochia were observed on inoculated wounded leaves, but not on non-wounded leaves. By 10 days, inoculated wounded leaves turned yellow and became covered with sporodochia all over the adaxial surface, and on wound sites on the adaxial surfaces. No signs or symptoms were observed on either non-wounded inoculated leaves or on plants sprayed only with water. P. buxi was re-isolated from the diseased leaves but not the water-treated leaves, to successfully complete Koch's postulates. To the best of our knowledge, this is the first report of P. buxi causing Volutella blight on boxwood in China.
References: (1) J. L. Bezerra. Acta Botanica Neerlandica 12:58, 1963. (2) B. Henricot et al. Plant Pathol. 49:805, 2000. (3) F. Shi and T. Hsiang. Eur. J. Plant Pathol. 138:763, 2014.
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