Molecular Detection And Screening Bactericides For Control Of Kiwifruit Bacterial Canker In The West Of Hunan

Kiwifruit bacterial canker disease, a devastating and serious disease threating to the kiwifruit production, had been listed as quarantine agent of Chinese forest plant. This dis-ease had been reported in Hunan, Shananxi, Sichuan, Fujian and Anhui provinces, causing enormouse economic losses of kiwifruit production. In this study, we had collected dis-eased samples of kiwifruit bacterial canker from the major producing areas of kiwifruit in Jishou, Luxi, Mayang, Fenghuang and Yongshun County of Hunan Province. The isolated and purified strains of canker disease were detected and identified using molecular tech-nology. We have tested 7 kinds of commonly bactericides for the disease control in green-house and field test using the plate count and field spray+lesion spots smear methods. The main results read as follows:1. The 14 isolates, coded as YSHY, YSML, YSJK, MYHY, MYCH, FHML, FHHY, JSCH, JSHY, LXHY, LXML, JSML, TDCY, TDHY, were isolated and purified from the samples infected by kiwifruit bacterial canker in the west of Hunan.2. By use of PCR, the 16S rDNA16S rDNA and 16S-23S rDNA spacer sequences (ITS) of isolates were amplified separately with primer pairs 27F/1492R and PsaF1/PsaR2. We obtained 16S rDNA gene of 1.4 kb and 16S-23S rDNA-ITS of 1.2 kb from the diseased samples. The fragments were purified and sequenced. We obtained 16S rDNA gene and 16S-23S rDNA-ITS in length of 1383 bp and 280 bp respectively. The 14 strains of 16S rDNA and 16S-23S rDNA-ITS sequences were identical. It was concluded that the 14 iso-lates were the same strain of kiwifruit canker disease.3. The results of the Blast searching of 16S rDNA gene indicated that:the 16S rDNAs of the 14 isolates were identical to that of the foreign reports in France 181, New Zealand ABAC9, Italy IKB4, etc. It shared 99.78% similarity with the Japan KW11, NCPPB3740 strains, Italy PsaH108 strain and Korea KBE9 strain with three bases differences. It shared 99.71% similarity with the strain 11-830-1 which was isolated in China with four bases differences. The results of sequencing and homologous comparison with other 16S rDNA gene showed that the isolates belonged to Pseudomonas. syringae pv. actinidiae (Psa).4. The result of the Blast searching of 16S-23S rDNA-ITS sequences indicated that: the isolates were identical with the strains reported SXHY11-1 in China, EFA131.1 in Spain and PA838 in Portugal. There were two bases differences with strain LMG5075 of kiwifruit bacterial canker disease reported in South Korea, which was reported to be P. sy-ringae pv. morsprunorum. The sequences similarity and phylogenetic tree showed that the isolates were in the same clade and belonged to P. syringae pv. actinidiae. The above re-sults showed that the 14 isolates from the kiwifruit with bacteria canker disease in the west of Hunan belonged to P. syringae pv. actinidiae.5. We had tested seven kinds of bactericides to control the disease in greenhouse and in the field. The results showed that 20% Saisentong SC had the best antibacterial effect with the EC50 of 64.01μg/mL, followed by 72% streptomycin with the EC50 of 80.28μg/mL. The EC50 of 50% Zn thiazole-Zhongshengmycin,85% Solanacearum este-Hymexazol, Ko-cide 3000(46% copper(Ⅱ) hydroxide),3% Zhongshengmycin and 80% mancozeb were 129.42μg/mL,242.75μg/mL,143.50μg/mL,256.66μg/mL,167.82μg/mL respectively. The 7 bactericides showed a good control of kiwifruit canker in the field test. It was showed that 600 times liquid of 20% Saisentong had the best control effect of 87.05%, fol-lowed by 800 times liquid of 50% Zn thiazole-Zhongshengmycin with the control effect of 85.33% and 800 times liquid of 72% streptomycin with the control effect of 85.27%. The control effects of 700 times liquid of 80% mancozeb,1300 times liquid of Kocide 3000(46% copper(Ⅱ) hydroxide),1300 times liquid of 85% Solanacearum este-Hymexazol and 1000 times liquid of 3% Zhongshengmycin were 77.30%,74.24%,72.03%,62.76% respectively. Based on the above results,600 times liquid of 20% Saisentong and 800 times liquid of 72% streptomycin can be promoted to apply in the field. We also found that it was better of the striking off scraps, comparing with spraying method.