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First Report of Volutella buxi Causing Volutella Blight on Buxus bodinieri in Central Plains China

    Authors and Affiliations
    • X. Wang
    • Y. X. Li
    • H. X. Dong
    • X. Z. Jia
    • X. Y. Zhang , Lab. of Forest Pathogen Integrated Biology, Research Institute of Forestry New Technology, Chinese Academy of Forestry, Beijing, P.R. China, 100091; and Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing Jiangsu, China, 210037.

      Boxwood (Buxus spp.) is a commercially important evergreen ornamental plant with worldwide distribution. In autumn of 2014, leaf spots were observed on Buxus bodinieri Levl in a commercial gardening nursery located in Henan Province; the disease continually occurred and spread in the spring of 2015. Initially, black or bronze spots were observed on leaves; the spots gradually expanded and turned white in the center and bronze on the boundary between healthy and blighted tissue. Light rose-colored sporodochia and white mycelium were observed on the abaxial leaf surface. Eventually, the diseased leaves withered and defoliated. Sometimes, twigs displayed black spots and dieback. The plants were still alive but their commercial value had sharply dropped. Diseased twigs and leaves were collected and cut into 1-mm2 pieces, sterilized in 75% ethanol for 30 s, then soaked in 1% sodium hypochlorite for 30 s, washed with sterile distilled water, and placed onto potato dextrose agar (PDA). After 1 day of incubation at 25°C, white aerial mycelia grew from diseased tissues. After 4 to 5 days, pink-orange sporodochia were observed in the middle of the colonies. The conidia were verticillate, unicellular, ovoid to elliptical, and measured 3.78 to 8.56 × 1.86 to 5.22 μm (average of 100 conidia: 5.85 × 3.14 μm). DNA was extracted from one colony containing spores and mycelium. The internal transcribed spacer (ITS) sequence (GenBank accession no. KX870844) of this isolate obtained with forward (ITS1) and reverse (ITS4) primers was 548 bp. When the sequences was compared against the GenBank NR database, the ITS sequence showed 99% identity with ITS sequences annotated as Pseudonectria buxi (KM231776). According to the morphological and molecular features, the pathogen was identified as Volutella buxi (Cda.) Berk (Bezerra 1963). The strain was deposited in the culture collection of Chinese Academy of Forestry and marked as CXY1814. To test the pathogenicity, 3-year-old B. bodinieri were inoculated with spore suspension harvested from a 7-day-old colony. As wounds are necessary for the infection process (Shi and Hsiang 2014), light scratches were made using sterile needles on the adaxial surface of leaves (15/plant). Three plants were sprayed on the wounded leaves with a spore suspension (106 CFU/ml) of V. buxi until runoff, while one plant was sprayed with sterile water as the negative control. The trees were incubated at 25°C in a growth chamber, with a 12-h photoperiod, at 85% humidity. After incubation for 15 days, black or bronze spots were observed on the inoculated leaves; finally, the leaves wilted and dropped. At the same time, we also inoculated three healthy branches of B. bodinieri following the same inoculation method. After incubation on Petri dishes with wet sterile paper for 5 to 6 days at 25°C, black spots were observed on the inoculated leaves of fresh branches, while no symptoms were observed on leaves sprayed only with sterile water. The pathogen was reisolated from the diseased leaves but not from the negative control. V. buxi was determined to be the causal agent of the blight on B. bodinieri. This pathogen was previously reported to cause volutella blight on Korean boxwood (B. sinica var. insularis) in Beijing, China (Shi and Hsiang 2014). Until now, this fungus has spread from Beijing to Henan Province in a short time and infected another important species of boxwood. So the disease has posed a great threat to Chinese important ornamental plant.


      Xuan Wang and Yong Xia Li contributed equally to the work.

      This work was funded by the Fundamental Research Fund for Central Public Welfare Research Institute (CAFYBB2014MB002) and the Special Research Programs for Non-profit of the State Forestry Administration (201204501).