水稻条斑病菌pilT基因在致病性中的功能分析

Roles of pilT gene in pathogenicity of Xanthomonas oryzae pv. oryzicola on rice

【目的】本实验室前期研究发现水稻条斑病菌(Xanthomonas oryzae pv.oryzicola,Xoc)RS105菌株中pil T基因Tn5转座子插入突变体在寄主水稻上致病性明显降低,在非寄主烟草上激发过敏反应(hypersensitive response,HR)的能力也明显减弱。为了揭示pilT基因在Xoc菌株中的功能,本文进行了深入研究。【方法】本实验通过无标记双交换敲除的方法获得Xoc RS105菌株pil T基因缺失突变体RΔpilT,并对该突变体的游动性以及生物膜等表型进行了检测。【结果】与野生型RS105菌株相比,敲除突变体RΔpilT不仅在感病水稻IR24上的致病性显著降低,在非寄主烟草上产生过敏反应的能力减弱,而且突变体游动性降低,生物膜含量增加,互补子能够恢复上述缺陷至野生型水平。qRT-PCR结果显示,在RΔpil T中,hrpG、hrpX、hrcC、clp、rpfG、pilA、pilC基因表达量明显降低。【结论】Xoc中pilT为重要的毒性相关基因,其致病性与游动性和生物膜含量变化相关,并且受clp、rpfG、pilA、pilC等基因调控。pil T基因编码的Pil T蛋白是构成IV型菌毛的亚基之一,为菌体运动提供能量。本文对pilT基因的功能研究,为进一步分析IV型菌毛在Xoc中的功能提供了线索。

[Objective] Our previous study found that a Tn5 transposon inserted mutant in pil T gene of Xanthomonas oryzae pv. oryzicola（Xoc） RS105 strain（the causal agent of bacterial leaf streak） significantly reduced bacterial virulence in susceptible rice plants, and the ability to trigger hypersensitive response（HR） on non-host tobacco. [Methods] To reveal the roles of pil T gene in bacterial pathogenicity in rice, we constructed a deletion mutant of pil T gene in Xoc RS105 named RΔpil T. Then we tested phenotypes of the mutant compared to the wild-type RS105 in bacterial virulence, motility and biofilm formation. [Results] The pil T mutant RΔpil T significantly reduced bacterial virulence in rice and the ability to trigger HR on tobacco. In addition, compared to RS105 strain, RΔpil T mutant displayed severely weakened motility on semisolid medium, and increased the production of biofilm. Full virulence, bacterial motility and biofilm formation was restored to the mutant when Xoc pil T was expressed in trans. Through q RT PCR analysis, we found that pil T gene was regulated by clp, rpf G, pil A and pil C. [Conclusion] These results indicate that pil T gene is essential for full bacterial virulence in rice possibly due to the association with the reduced motility and increased biofilm production by the pil T mutation, implying that the importance of further investigation on the function of Pil T in composition of type IV pili in Xoc.