组氨酸氨解酶HutH对副溶血弧菌生物学特性及致病性的影响

Effects of histidine aminonia-lyase on the biological characteristics and pathogenicity of Vibrio parahaemolyticus

摘要:

组氨酸氨解酶(HutH)作为组氨酸代谢通路上的第一个代谢酶，控制细菌内部组氨酸的代谢。HutH在多数细菌中高度保守，参与细菌的能量代谢平衡。【目的】选取HutH作为研究对象，探究其对副溶血弧菌生物学特性以及致病性的影响。【方法】利用同源重组的方法构建缺失株ΔhutH和回补株CΔhutH。研究HutH对副溶血弧菌生长特性、组氨酸利用能力、组氨酸代谢相关基因表达水平、运动性、生物被膜、环境耐受、细胞毒性以及对小鼠毒力的影响。【结果】与野生型菌株相比，hutH基因缺失不影响副溶血弧菌的生长特性、耐酸耐碱能力、耐盐能力和群集运动。但ΔhutH在组氨酸作为唯一碳源的M9极限培养基中生长受到显著抑制。另外，我们证实，hutH基因缺失使组氨酸代谢操纵子内的相关基因转录水平显著下降，基因VP0889的表达水平提高。hutH基因缺失导致副溶血弧菌生物被膜形成能力下降、泳动能力下降、对HeLa细胞的毒性降低、对ICR小鼠的致死率显著降低。【结论】本研究表明，hutH基因缺失影响副溶血弧菌代谢组氨酸的能力，而且首次证实，HutH在副溶血弧菌的生物被膜形成、运动性和对小鼠毒力等方面具有重要作用，为从调控组氨酸代谢途径来进行细菌防控提供思路。

Abstract:

As the first metabolic enzyme in the histidine metabolic pathway, histidine aminonia-lyase (HutH) controls the metabolism of histidine in bacteria. HutH is highly conserved in most bacteria and participates in the balance of bacterial energy metabolism.[Objective] To investigate the effects of HutH on the biological characteristics and pathogenicity of Vibrio parahaemolyticus. [Methods] The hutH gene-deleted mutant ΔhutH and the complementary strain CΔhutH were constructed by homologous recombination. The effects of HutH on the growth characteristics, histidine utilization ability, histidine metabolism related gene expression level, motility, biofilm, environmental tolerance, cytotoxicity and mouse toxicity of V.parahaemolyticus were researched. [Results] Compared with wild strains, the deletion of hutH gene did not affect the growth characteristics, acid and alkali tolerance, salt tolerance and swarming ability of V.parahaemolyticus. But the growth of ΔhutH was significantly inhibited in M9-limited medium with histidine as the only carbon source. In addition, we confirmed that the deletion of hutH significantly decreased the transcription level of histidine metabolism related genes in hut operon and increased the expression level of the VP0889 gene. The deletion of hutH resulted in the decrease in biofilm formation ability, swimming ability, toxicity to HeLa cells and mortality to institution of cancer research female mice. [Conclusion] This study shows that the deletion of hutH affects the ability of V.parahaemolyticus to metabolize histidine, and it is the first time to confirm that HutH plays an important role in the biofilm formation, motility and toxicity toward mice of V.parahaemolyticus, which provides ideas for bacterial prevention and control by regulating histidine metabolism.