First Report of Blight Disease on Buxus Caused by Cylindrocladium buxicola in France.

Cylindrocladium buxicola Henricot causes twig blight on Buxus spp., severe defoliation, and eventually death of plants, especially in young seedlings (1). The disease was first observed in the United Kingdom and New Zealand in the 1990s and recently, the fungus was detected in other European countries (1). In November 2006, box blight symptoms were observed in a Buxus sempervirens nursery located in the South West of France (La Reole). Since then, more diseased samples from other French sites, including forestry areas and ornamental garden nurseries, have been received, indicating that this disease is spreading. Symptomatic twig samples were sent for lab analysis and dark brown spots were observed on the leaves, sometimes coalescing to cover the entire leaf surface, with black streaks on the stems. Fungal fruiting structures were observed directly on the leaf surface and were examined with a stereomicroscope. Microscopic slides were then prepared by gently pressing a clear adhesive tape onto the surface covered by mycelium and spores, which was further stained with lactic acid and methyl blue. Cylindrical, straight, biseptate, hyaline conidia, 53.8 to 75.3 (64.4) × 4.4 to 5.2 (4.6) μm, were observed, sterile hyphae with terminal vesicles ended with a pointed apex, and conidiophores were penicilliate; all of those characters were consistent with C. buxicola (2). To support the diagnosis, fruiting structures were plated on malt agar media supplemented with 100 ppm of chloramphenicol. The pure culture obtained showed a whitish mycelium with a tan brown center that was in line with the original description of C. buxicola (2). DNA was extracted from the pure culture and the internal transcribed spacer (ITS) region was amplified by PCR using the ITS1-ITS4 primer pair (4). Nucleotide sequence was determined and deposited on GenBank (Accession No. JQ743502). BLAST analysis of the sequence showed 100% identity with all currently available C. buxicola ITS sequences, which confirmed our morphological diagnosis. To our knowledge, this is the first report of C. buxicola (teleomorph Calonectria pseudonaviculata) in France. The occurrence of this disease in France worries the nursery industry since losses can sometimes be dramatic as seen in United Kingdom, where the disease is widespread (3). References: (1) B. Henricot. The Plantsman 9:153, 2006. (2) B. Henricot and A. Culham. Mycologia 94:980, 2002. (3) B. Henricot et al. Plant Pathol. 49:805, 2000. (4) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications, 1990.