烤烟霉变病原菌鉴定及其生防菌筛选

为明确云南省大理州烟叶烘烤变黄期叶柄和部分叶片霉变的病原菌，依据柯赫氏法则，采用组织分离法分离病原菌，经室内和烤房内验证致病性，结合形态学和多基因序列分析鉴定病原物种类，运用平板拮抗法和活体生测法筛选生防菌。结果表明，分离物的形态学与根霉属（Rhizopus sp.）真菌相似，其ITS序列与少根根霉菌（Rhizopus arrhizus.，异名Rhizopus oryzae）模式菌株CBS 112.07^T序列一致性大于99%；基于内转录间隔区（ITS）、肌动蛋白（ACT）和翻译延伸因子1-α（TEF）的多基因序列系统发育分析也表明，病原菌与少根根霉菌亲缘关系最近。综合形态学和多基因序列分析结果，确定烤烟霉变病病原菌为少根根霉菌。通过平板对峙法筛选获得5株抑菌能力较强的拮抗菌，其中芽孢杆菌（Bacillus spp.）Z002和05-1205在烤房内的防治效果均在70%以上。

Causal Pathogen Identification and Antagonistic Bacteria Screening of Tobacco Leaf Mold

The objective of this study was to identify the pathogen and screen the biological control agents of tobacco leaf mold, which causes damage losses during the yellowing stage of leaf curing each year in several counties of Dali Bai Autonomous Prefecture of Yunnan Province, China. Infected tissues were used for isolating the causal pathogen, which were classified by combining results from the morphology and multiple gene sequences analysis. Antagonistic activities of bacteria against the pathogen were evaluated in vitro and in vivo. Fungi obtained from infected tissues were grouped into the genus Rhizopus based on morphological characteristics. Furthermore, internal transcribed spacer region (ITS) sequences of isolates had the highest similarity with R. arrhizus subsp. arrhizus CBS 112.07^T. Similar results were obtained from multiple sequence alignment and phylogenetic analyses using the partial sequences of ITS, actin (ACT), translation elongation factor 1-α (TEF), and the largest subunit of RNA polymerase II (RPB1). Above all, the pathogen causing tobacco leaf mold in Dali is R. arrhizus. Besides, five antagonistic bacteria had robust inhibition activities against hyphal growth of R. arrhizus in vitro. Among these bacteria, the Bacillus strains Z002 and 05-1205 were selected for biological control efficiency tests in the barn and showed strong evidence to support them as potential biological agents with relative control efficiency >70%.