Identification, Overwintering And Infection Process Of The Pathogen Causing Adzuki Bean Rust

Adzuki bean rust is one of the major diseases of adzuki bean and epidemic in the growingregions, which has influenced seriously the yield loss and quality of adzuki bean. However, thereare confusions about the overwintering of teliospores and infection processes of urediospores,especially the agent of adzuki bean rust. Thus, the identification of the agent of adzuki bean rust,overwintering of teliospores and infection processes of urediospores, can lay foundations for theresearch, including the life cycle and molecular analysis. Thus, the objectives of the currentstudy were⑴to identify the pathogen causing adzuki bean (Vigna angularis) rust in Daqing,Heilongjiang province, observing the number of germ pores of urediniospores, germ porelocation and the wall thickness of teliopores, and sequencing ribosomal DNA internal transcribedspacer (rDNA-ITS).⑵to clarify the teliospores of adzuki bean rust overwintering inHeilongjiang Province, investigating the teliospore germination rate, which subjected in differentconditions.⑶To elucidate the infection process including urediospores germination, penetrationand spread in host tissue of Vigna angularis using light and fluorescence microscope.1. A pure culture of rust isolate ZXL01was obtained by the method of single pustuleisolation. Its taxonomic status was determined by observing the number of germ pores ofurediniospores, germ pore location and the wall thickness of teliopores, and sequencingribosomal DNA internal transcribed spacer (rDNA-ITS). Morphological investigation showedthat most of the urediospores of ZXL01had two germ pores which were far from spores’ equatorarea. The wall thickness of teliopores ranged from2.9to3.3μm. The rDNA-ITS sequence ofZXL01was clustered in one clade with two reference isolates of Uromyces vignae (GenBankaccession numbers AB115718and AB115731) at99%bootstrap levels in the phylogenetic tree.A500bp amplified product was obtained by the specific primers UV-ITSF/R, which wasspecific for U. Vignae, the host range test showed that the pathogen could only infect Vignaangularis.2. In order to clarify the teliospore of adzuki bean rust overwintering in HeilongjiangProvince, the teliospores germination rate, which subjected in different conditions, wasinvestigated. The teliospore could germinate after treated in5conditions, including buriedunderground10cm, hanging outdoor, room temperature,4℃and-20℃. However, the germination rate was significantly different. The germination rate of4℃and-20℃conditonwas up to35%.27℃with darkness was suitable for teliospores germination. Our resultsindicated that teliospore could overwinter safely in Heilongjiang Province.3. The infection process of urediospores, the causal agent of adzuki bean rust, in adzukibean (Vigna angularis) tissue was investigated by light and fluorescence microscope. Theurediospre penetrated leaves by formation of appressoria on the tip of the germ tube6h post-inoculation (hpi). The typical haustorial mother cell was observed24hpi. The typical haustoriumin the epidermal cells and intercellular hypae were observed48hpi. Colony of the pathogen wasobserved in the mesophyll tissue120hpi. The lesion occurred on the leaves. The freshurediospores was produced8dpi.