Genetic Diversity And Pathogenicity Analysis Of Fusarium Pseudograminearum That Is The Dominant Pathogen Causing Crown Rot Of Wheat

Crown rot of wheat is a newly discovered soil-borne disease in China,which is caused by a diverse group of Fusarium species.It occurs generally in Huang-Huai wheat area and shows a tendency of expansion and aggravation,which is a serious threat to wheat production.In this study,we collected samples of wheat plants with symptoms of crown rot from Huang-Huai wheat area in Henan,Hebei,Shanxi,Anhui,Jiangsu and Shandong provinces in May 2014 and 2015.Isolates were obtain through tissue isolation of samples and identified by morphological and molecular approaches that the pathogens and dominant pathogen species of crown rot of wheat in Huang-Huai wheat area were clarified.The genetic diversity of the dominant pathogen of Fusarium pseudograminearum populations collected from different regions in Huang-Huai wheat area was analyzed using inter-simple sequence repeat(ISSR)and URP-PCR marker,and the pathogenicity test of the representative pathogenic isolates were also conducted.The relationships among the genetic diversity of F.pseudograminearum,geographic distribution and pathogenicity were defined in this region.Main results were showed below:1.Wheat plants with symptoms of crown rot were collected from Huang-Huai wheat area.By tissue isolation,morphological and molecular identification,the results showed that the members of the pathogen complex in the Huang-Huai wheat area were a variety of Fusarium species,which included F.pseudograminearum,F.graminearum,F.proliferatum,F.equiseti,F.tricinctum,F.oxysporum,and F.culmorum.Crown rot of wheat in Huanghuai area was primarily caused by F.pseudograminearum and F.graminearum.The frequency of isolation of F.pseudograminearum was highest,accounted for 52.49%,then F.graminearum,accounted for 23.33%.The pathogens in different regions had certain difference.F.pseudograminearum was the dominant pathogen in Northern Henan,Middle of Henan and Hebei,and the frequency of isolation reached 69.89%,58.00% and 53.19%,respectively.In Southern Henan,Western Henan,Eastern Henan,Shandong and Shanxi,the dominant pathogens were F.pseudograminearum and F.graminearum,and the frequency of detection were all above 21%.In contrast,F.graminearum was the dominant pathogen in Jiangsu and Anhui with the frequency of isolation of 100.00% and 33.33%,respectively.2.The genetic diversity of 166 F.pseudograminearum isolates collected from Henan and Hebei winter wheat regions was analyzed with 17 ISSR primers.The amplification results showed that 234 fragments were generated,and 218 fragments were polymorphic which accounted for 93.16% in the total amplified fragments.The average number of bands amplified per primer was 13.76.The Shannon's information index and Nei's gene diversity were highest in Northern Henan and lowest in Southern Henan.By analysis of the genetic similarity coefficient,the relationship of F.pseudograminearum populations of Northern Henan and Eastern Henan were closest and that of Southern Henan and Eastern Henan were the farthest.The coefficient of the population genetic differentiation(Gst)among geographical groups was 0.1571,while it reached 0.8429 within the populations indicating more divergent genetic diversity within the populations.The number of migrants per generation among 6 geographical populations was 2.6819,which indicated that genetic information exchange was frequent among the 6 populations.The dendrogram based on ISSR markers revealed that 6 geographical populations were clustered into two groups at the threshold of a genetically similar coefficient of 0.966.The group ?included five regions,i.e.Northern Henan,Eastern Henan,Middle of Henan,Middle of Hebei,and Western Henan,while Southern Henan belonged to group II.3.The genetic diversity of 261 F.pseudograminearum isolates collected from Huang-Huai wheat area was analyzed with 9 URP primers.The amplification results showed that 132 fragments were amplified,and 128 fragments displayed polymorphism which accounted for 96.97% in the total amplified fragments.The average number of bands amplified per primer was 14.67.The primer URP9 F generated 19 bands,the greatest number of fragments.The number of migrants per generation among 8 geographical populations was 2.8362,which indicated that genetic information exchange was frequent.The relationship of F.pseudograminearum populations of Northern Henan and Middle of Henan were closest and that between Shandong and Southern Henan were the farthest.At the similarity coefficient of 0.962,all isolates were clustered into two groups.The group ?included five regions,i.e.Northern Henan,Middle of Henan,Eastern Henan,Western Henan and Southern Henan.The group II included three regions,i.e.Hebei,Shandong and Shanxi.4.The pathogenicity of the representative of F.pseudograminearum isolates collected from Huang-Huai wheat area in test showed very significant differences.The isolate XX114-9A from Juqiao Town,Xunxian,Hebi,and Northern Henan had the strongest ability to cause the disease with the disease index and incidence rate of 91.11 and 100%,respectively.The isolate FC136-2A from Chengguan Town,Fangcheng,Nanyang,and Southern Henan had the weakest ability to cause the disease with the disease index and incidence rate at 6.67 and 25% respectively.The pathogenicity of different isolates had no significant relationship with geographical origin in which all F.pseudograminearum isolates from different regions of Huang-Huai wheat area was pathogenic even there were some variation the isolates from the same geographic origin.