Sapstain of Japanese Black Pine Caused by Ophiostoma piceae in Korea

Abstract

Sapstain fungi that cause blue or black discolorations of sapwood indiverse trees are one of the important fungal groups in forest healthas tree pathogens, and in forest products industry as wood-stainers(Seifert, 1993). Sapstain, caused by fungi, is the cosmetic damage ofwood that reduces wood quality and the values of commercial woodproducts. In July 2006, bluish or brown colored signs of sapstainwere found on cut sapwood of Japanese black pine (Pinusthunbergii) in Taean, Korea (Fig. 1A). The signs of sapstain werealso found on cut sapwood of Japanese black pine infected withpinewood nematode in Jinju in June 2007. Voucher specimens weredeposited at Dankook University Culture Collection, Cheonan,Korea (DKM 0510, 0410, 0412, 0413, 0601).The fungal colony was brown and dark on MEA, light brown onPDA and black on oat meal agar (Fig. 1B). Optimal growthtemperature and pH for the fungus was 22 and pH 6. The fungusshowed tolerance to cycloheximide (250 μg/ml). The optimal pHfor growth was on PDA and MEA. Observation of morphologicalfeatures on a phase-contrast light microscope (Karl Zeiss Axioskop40) and a scanning electron microscope (Hitachi S-4200) revealedthat the fungus has Sporothrix and Pesotum anamorph of O. piceae-complex. Conidial mass on apex of synnema was milky color.Synnema were dark brown and 500-1300 ( −1600)μm. Synemmat-ousconidia were hyaline and 5-15×1-3.5 μ m (Fig. 1C-E). Peritheciaand ascospores were not found in our cultures. To confirm the identity of this fungus, partial β-tubulin gene wasamplified with primers T10 and BT12 (Kim et al., 2003) and directlysequenced. The determined nucleotide sequence was deposited inGenBank (EU012505). The determined sequence shared 100%sequence identity with those of O. piceae. Based on β-tubulin genesequence, a cladogram was constructed using PAUP by the neigh-bor-joining method with the heuristic search option (Swofford,2002). The fungus grouped with O. piceae (Fig. 2). This moleculardata confirmed the fungus is O. piceae. An inoculation test on woodchips of Japanese pine displayed that the fungus colonized anddeveloped dark and/or brown stain in the chips (Fig. 3B). On theinoculated chip, the fungus formed its typical brown pigmentedsynnema having a droplet of conidial mass (Fig. 3C). So far m, ore than 150 species of Ophiostoma have been descrbed,ibut their occurrence has rarely been studied in Korea. This is the firstreport of O. piceae in Korea. Since we isolated O. piceae from a pinediseased by pinewood nematode that recently has devastated pinetrees in Korea, further study on Ophiostoma fungi should be done toexplore their impact on forest health and wood discoloring damage.