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获得纯度高的青枯雷尔氏菌无致病力菌株,是研发青枯病植物疫苗和防治青枯病害的一种新途径。作者以青枯雷尔氏菌强致病力菌株FJAT-91为出发菌株,通过对hrpB基因敲除,获得无致病力突变菌株FJAT-91⊿hrpB。高效离子交换色谱分离结果表明,FJAT-91和FJAT-91⊿hrpB色谱峰型不同,主要表现在峰的保留时间上,FJAT-91只有单一色谱峰,保留时间为6min,FJAT-91⊿hrpB有P1和P2两个色谱峰,保留时间分别为0.6min和4.5min。利用高效离子交换色谱对FJAT-91⊿hrpB进行纯化,获得只有P1峰的高纯度菌株FJAT-91⊿hrpB-P。FJAT-91⊿hrpB和FJAT-91⊿hrpB-P与其出发菌株FJAT-91的菌落和菌体形态差异明显。致病力测定结果表明,FJAT-91接种4d番茄植株开始发病,10d发病率达100%,FJAT-91⊿hrpB和FJAT-91⊿hrpB-P接种20d均未发病。防效试验结果表明,纯化后的菌株FJAT-91⊿hrpB-P对番茄青枯病的防效(81.64%)比未纯化FJAT-91⊿hrpB防效(61.04%)提高了33.75%。本研究获得一株高纯度的青枯雷尔氏菌无致病力突变菌株FJAT-91⊿hrpB-P具有良好的生防潜力。
Obtaining a virulent strain of Ralstonia solanacearum with high purity is a new way to develop plant wilt disease vaccine and control bacterial wilt. In this paper, the strong pathogenic strain FJAT-91 of Ralstonia solanacearum was used as starting strain, and the mutant strain FJAT-91⊿hrpB was obtained by knocking out hrpB gene. The results of high performance ion exchange chromatography showed that the peaks of FJAT-91 and FJAT-91⊿hrpB were different from each other in peak retention time. FJAT-91 had only a single chromatographic peak with a retention time of 6 min. FJAT-91⊿hrpB had P1 and P2 two peaks, retention time was 0.6min and 4.5min. FJAT-91⊿hrpB was purified by high performance ion exchange chromatography to obtain a high-purity strain FJAT-91⊿hrpB-P with only P1 peak. FJAT-91 ⊿ hrpB and FJAT-91 ⊿ hrpB-P and its starting strain FJAT-91 colonies and cell morphology significantly different. The results of pathogenicity test showed that FJAT-91 inoculated with tomato plants started to develop on the 4th day, and the incidence rate reached 100% after 10 days. No incidence was observed after 20 days of FJAT-91⊿hrpB and FJAT-91⊿hrpB-P inoculation. The results of the anti-efficacy test showed that the control efficacy of the purified strain FJAT-91⊿hrpB-P on tomato bacterial wilt was increased by 33.75% (81.64%) compared with that of the unpurified FJAT-91⊿hrpB (61.04%). In this study, we obtained a strain of non-pathogenic mutant FJAT-91 (hrpB-P) with high purity and high resistance to R. solanacearum.