胶孢炭疽菌侵染杨树叶片的组织病理学研究

目的明确胶孢炭疽菌在杨树叶片上的侵染过程，为进一步从分子水平研究该菌的致病机制和杨树抗病分子育种奠定基础。方法用绿色荧光蛋白标记的胶孢炭疽菌菌株BH12-2的分生孢子悬浮液接种健康杨树叶片，采用光学显微镜和电子显微镜观察病原菌的侵染过程和杨树叶片的防卫反应。结果接种4h后，孢子开始萌发产生芽管；8h时芽管顶端形成附着胞；12h时成熟的附着胞中央形成侵染钉；24h时，孢子另一端萌发形成芽管和附着胞；48h后芽管不断分枝异化成菌丝并产生次级分生孢子；接种3d时，附着胞基部的侵染钉穿透寄主角质层和表皮细胞壁膨大形成侵染泡囊，侵染泡囊初始生长在寄主细胞壁和细胞膜之间，不穿透寄主的原生质体，随后产生初生菌丝和次生菌丝的分化；接种4~5d后，次生菌丝在寄主表皮和叶肉组织内大量扩展；第6d时，菌丝聚集在角质层下形成子座组织，并产生分生孢子梗和分生孢子。随着菌丝的扩展，叶片组织发生一系列的病理变化，在侵入点周围的叶肉细胞壁附近产生胼胝质，细胞壁向内凹陷并发生溶解，细胞质消解，叶绿体等细胞器解体以及寄主细胞坏死塌陷，最终在叶表面产生典型的褐色坏死病斑。结论胶孢炭疽菌在侵染过程中，一个分生孢子可萌发形成多个芽管和附着胞，提高其成功侵染的几率；胶孢炭疽菌对杨树叶片的侵染类型为细胞内半活体营养侵染型。 

Histopathology study of poplar leaves infected by Colletotrichum gloeosporioides

[ Objective ] The present experiment was conducted to study the infection process of poplar leaves by Colletotrichum gloeosporioides and the inoculated leaf reaction, providing scientific basis for further research on pathologic mechanism, as well as molecular breeding for disease resistance of poplar. [ Method] The healthy poplar leaves were inoculated with conidia suspension of C. gloeosporioides green-fluorescent-protein-marked strain BH12-2 . The infection process and defense response between pathogen and poplar leaves were observed by optical and electron microscopies. [ Result] The results showed that condium began to germinate and form germ tube at 4 hour post-inoculation ( hpi ) . The germ tube produced appressorium after 8 hpi. The mature appressorium formed infection peg after 12 hpi. A germ tube and appressorium formed at the other top of the germinal conidium after 24 hpi. Germ tubes of abundant branches differentiated into hyphae and produced secondary conidia after 48 hpi. The infection peg formed in the base of appressorium, swelled to form an infection vesicle after penetrating the host cuticle and epidermal cell wall after 3 days post-inoculation ( dpi ) . The infection vesicle initially grew between the host cell wall and cell membrane, without penetrating the protoplast of the host, and subsequently produced primary hyphae and secondary hyphae. The secondary hyphae rapidly expanded in the host epidermal and mesophyll tissues within 4 -5 dpi, gathered in the subcuticular to form stroma tissue in 6 dpi, from which conidiophore and conidia were then developed. With mycelial extension in host tissues, a series of alterations occurred in host tissues, including callose formation around the cell wall of mesophyll cells at the penetration point, downward deformation and dissolution of cell wall, degeneration of cytoplasm, disintegration of organelles such as chloroplasts and collapse of host cells, resulting in appearance of typical brown necrotic spots on the infected leaves. [ Conclusion ] In the infection process of C. gloeosporioides, conidium can germinate and form multiple germs and appressoria to increase the probability of successful infection. C. gloeosporioides adopts the intracellular hemibiotrophic infection strategy for colonization in poplar leaves.