lpxL和lpxM基因对O1、O78血清型禽致病性大肠杆菌毒力的影响

Effect of lpxL and lpxM genes on virulence of avian pathogenic Escherichia coli belonging to O1 and O78 serogroups

摘要:

[目的] 为探究脂多糖对O1、O78血清型禽致病性大肠杆菌（Avian pathogenic Escherichia coli，APEC）致病作用的影响。[方法] 选取负责脂质A生物合成相关基因lpxL和lpxM，利用λ噬菌体的Red同源重组系统分别构建APEC E516（O1血清型）和APEC E522（O78血清型）缺失株E516ΔlpxL、E516ΔlpxM、E516ΔlpxLΔlpxM、E522ΔlpxL、E522ΔlpxM和E522ΔlpxLΔlpxM，并通过体内外试验对其生物学特性及致病性进行研究。[结果] 各菌株生长速度基本一致。E516ΔlpxL、E516ΔlpxLΔlpxM、E522ΔlpxM和E522ΔlpxLΔlpxM的抗血清补体杀菌能力和抗鸡巨噬细胞HD-11吞噬能力较野生株显著下降，而缺失株E516ΔlpxM、E522ΔlpxL与野生株相比无明显差异；半数致死剂量测定结果显示，除E516ΔlpxM、E522ΔlpxL外，各缺失株毒力降低1000倍左右；SPF鸡体内动态分布试验结果显示，各缺失株在鸡体内定殖能力较野生株显著下降，但回补株的毒力未能恢复至野生株水平。[结论] lpxL和lpxM基因与O1血清型APEC E516株和O78血清型APEC E522株的毒力有关，但是lpxL和lpxM基因对E516和E522菌株毒力的影响存在差异。

Abstract:

[Objective] To explore the role of lipopolysaccharide in Avian pathogenic Escherichia coli (APEC). [Methods] By using λ-phage Red recombination system, we generated the lipid A biosynthesis associated genes lpxL and lpxM mutants E516ΔlpxL, E516ΔlpxM, E516ΔlpxLΔlpxM, E522ΔlpxL, E522ΔlpxM and E522ΔlpxLΔlpxM. We conducted a series of in vivo and in vitro assays to investigate their biological characteristics and pathogenicity. [Results] The growth rate of the strains was basically the same. The ability of resistance to serum and killing by chicken macrophages were significantly impaired, E516ΔlpxL, E516ΔlpxLΔlpxM, E522ΔlpxM and E522ΔlpxLΔlpxM were significantly attenuated than those of the wild-type strains, and E516ΔlpxM and E522ΔlpxL had no obvious difference in relation to their parent strains. LD₅₀ results show that the pathogenicity of mutants was significantly attenuated than those of the wild-type strains. The colonization and survival model demonstrated that the loads of mutants were significantly decreased compared with those of the wild-type strains in all bloods and organs tested in chickens. The ability of mutants to enter and survive in chicken macrophage HD-11 is significantly reduced compared with those of the parent strains. [Conclusion] These results indicated that lpxL and lpxM genes are critical to the pathogenicity of APEC E516 and E522, lpxL had a more significant effect on virulence of APEC E516, and lpxM gene on virulence of APEC E522.