Isolation,Identification And Virulence Determination Of Sorghum Anthracnose Pathogen In Sichuan

The history of sorghum liquor brewing in China has been written for more than 700 years.A series of famous liquor brands,such as Maotai in Guizhou,Fenjiu in Shanxi,Jiannanchun in Sichuan,Laojiao in Luzhou and Wuliangye,are brewed with sorghum as the main ingredient or condiment.Therefore,sorghum has become an indispensable part of liquor industry.However,the anthrax disease in China is serious and can reduce sorghum production by more than 30%.Early studies suggested that the pathogen of sorghum anthracnose was Colletotrichum graminicola(Cesati)Wilson.However,recently,foreign scholars confirmed that the pathogen of sorghum anthracnose was Colletotrichum sublineola,not C.graminicola,through genome sequencing and genomic analysis.In the mainland of China,the identification of sorghum anthracnose is rarely reported.C.graminicola is used as the pathogen of sorghum Anthracnose in some literatures.This conclusion is inconsistent with foreign literatures.Therefore,in order to clarify the pathogen of sorghum anthracnose,follow-up studies have been carried out.Sixty-three samples of sorghum anthracnose were collected from five districts in Sichuan Province,namely,Yibin Nanxi District,Yibin Jiangan County,Yibin Changning County,Zigong Fushun County and Zigong Da'an District.Fifty-four samples were collected from six districts,including Yongchuan,Zhongxian,Lincheng,Hebei Nangong,Weifang,Shandong Province and Yuanjiang,Yunnan Province.A total of 117 samples of sorghum anthracnose were collected.149 single spore strains were purified by single spore isolation.According to the morphological characteristics,all strains can be divided into three types:crescent-shaped,cylindrical and sickle-shaped.There are 120 strains in crescent-shaped,such as strain ZG-FS-1.The morphology and morphology of colonies,mycelial growth rate,conidia size and morphology,and appressorium size and morphology are analyzed.C.sublineola is similar to the fungus strain of the Tao;10 cylindrical strains,such as strain ZG-DA-2,are similar in morphology to the reported strain C.cliviicola,and 19 other strains are similar to Fusarium and Pythium.Combined with molecular biology,149 strains were clustered by ITS sequence analysis.The results were consistent with morphological classification.The 120-month conidia strains were clustered into one group,belonging to C.sublineola;10 cylindrical conidia strains were clustered into one group,belonging to C.cliviicola;the other 19 strains were divided into five branches according to the strains.Originally,one strain was selected from C.sublineola in 11 regions,and two strains were selected from C.cliviicola.The other four genes(GAPDH,CHS-1,ACT and TUB2)were analyzed.Through multi-gene tree building,11 C.sublineola strains were divided into the same branch as the standard strain C.sublineola YN1728,which not only verified the results of ITS analysis,but also further tabulated them.The 11 strains belong to C.sublineola.In addition,the remaining two strains are similar to C.cliviicola CBS 125375,belonging to C.cliviicola.We analyzed the pathogenicity of 13 strains after polygenic sequencing on Sorghum Hui 1 material.The results showed that 11 strains of C.sublineola could be pathogenic.After isolation and purification by single spore,we obtained strains consistent with the original strain in morphology and gene sequence.It showed that these 11 strains were pathogenic bacteria of sorghum anthracnose,but two strains of C.cliviicola,ZG-DA-2 and CQ-ZX-2,were pathogenic bacteria of sorghum anthracnose.Sorghum is not pathogenic,so the pathogen of sorghum Anthracnose in Sichuan is C.sublineola,while C.cliviicola is not the pathogen of sorghum anthracnose.ZG-FS-1 was also found to be highly pathogenic and HB-NG-1 was weakly pathogenic among 11 tested strains.The two strains and CQ-ZX-2 were inoculated with different materials,such as Hui 1,Zao 21 R,0953R and 402 B,as well as 11 Sichuan sorghum cultivars such as Jinnuo Liang 1.402 B,Chuannuo Liang2 and Jinuo Liang 1 were found to be highly resistant to dominant pathogenic bacteria and to anthrax in sorghum.There is a good application potential in disease breeding,but Jinnuo sorghum No.1 and Jinnuo sorghum 234 are more susceptible.We should pay attention to the prevention and control of anthrax in the planting process of these two varieties.