| Fusarium wilt of bitter gourd is a devastating disease,counteracted the development of production of bitter gourd seriously,and it occured in the most of bitter gourd grown in China.From 2016 to 2018,we had collected these samples from infccted bitter gourd of nine provinces in China,including Shandong,Henan,Hubei,Hunan,Jiangxi,Guangdong,Guangxi,Fujian and Hainan.Fusarium species were obtained using the tissue isolation method,these isolates were identified by morphology,formae speciales,pathogenicity and variety resistance test.On the basis of the phylogenetic relationships in Fusarium species were studied by the sequence data sets of rDNA-ITS,EF-1α and β-iubulin genes,a special detection technology was established for Fusarium oxysporum f.sp.momodicae based on URP-PCR polymorphic bands,meanwhile,the characteristics of Fusarium oxysporum from nine provinces genetic diversity were studied by ISSR-PCR method.An antagonistic bacteria was isolated with target by Fusarium oxysporum f.sp.momodicae,its biocontrol on bitter gourd,antimicrobial spectrum and greenhouse countrol effect,the morphological character and molecular identification was evaluated.The main results were described as follows:1.One hundred and seventy-three isolates were identified from nine province in China,including 19 isolates from Shandong,22 isolates from Henan,20 isolates from Hubei,17 isolates from Hunan,22 isolates from Jiangxi,21 isolates from Guangdong,20 isolates from Guangxi,20 isolates from Fujian and 12 isolates from Hainan.There was no differentiate virulence between the strains from different provinces,the morphological characteristics of the strains were complex and varied,and no correlation with the geographical distribution.These isolates could infect bitter gourd and luffa,bottle gourd weakly,but not pumpkin,muskmelon,watermelon and cucumber.2.Development of special detection technology for F.oxysporum f.sp.momodicae based on URP-PCR polymorphic bands.The application showed that the specific 294 bp amplicon was generated with the specific pair primers FOMM-SPF/FOMM-SPR in 25 μL of detection volume including 12.5μL of 2× Green Taq Master Mix,1 μL of 10 μmol/L each primers,1 μL of template DNA,and sterile ddH2O to obtain the final volume;The specific pair primers for 294 bp product.The processing methods exhibited the high specificity and sensitivity F.oxysporum f.sp.momodicae that could be detected in the soil and plant samples rapidly and accurately instead of pathogen isolation and pathogenicity test,which would be beneficial in significance for the early diagnosis,early warning,and effective control of the bitter gourd wilt disease.3.The results of genetic diversity showed that the genetic diversity of Fusarium wilt of bitter gourd groups was less differentiation in Shandong,Henan,Hubei,Hunan,Jiangxi,Guangdong,Guangxi Fujian and Hainan.The genetic variation of the isolates in natural populations accounted for 73.45%of the total variation,which was the main source of genetic variation.The UPGMA based on ISSR markers revealed that the genetic similarity coefficient is 0.9620,strains from nine provinces could be divided into group Ⅰ and group Ⅱ,group Ⅰ includeing Shandong and Henan,group Ⅱ includeing Hubei,Hunan,Jiangxi,Guangdong,Guangxi Fujian and Hainan;the result shows obvious north-south distribution characteristics.When genetic similarity coefficient was 0.9676,group Ⅱ is divided into three subgroups,the guangdong,hainan,fujian,jiangxi and hubei for the same subgroup,hunan as a subgroup,guangxi as a subgroup.These results showed that there was a certain correlation between the phylogenetic relationship of the natural populations of Fusarium oxysporum f.sp.momodicae and its geographical origin.4.An antagonistic bacterium strain LWC6 with obvious countrol effect on Fusarium wilt of bitter gourd was screened and identified as pseudomonas aeruginosa by morphological analysis and 16S rRNA gene sequence analysis. |
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