Department of Fisheries Technology Engineering, Faculty of Marine Science, Karadeniz Technical University, Trabzon, 61530, Turkey.
Department of Fisheries Technology Engineering, Faculty of Marine Science, Karadeniz Technical University, Trabzon, 61530, Turkey.
Fish Shellfish Immunol. 2016 Aug;55:21-7. doi: 10.1016/j.fsi.2016.04.018. Epub 2016 Apr 16.
Yersinia ruckeri is a Gram negative bacteria causing yersiniosis in freshwater and marine fish. Lipid A, important for pathogenesis of Gram negative bacteria, biosynthesis pathway requires nine enzyme catalyzed steps. Although there are nine genes encoding lipid A biosynthesis in bacteria, biosynthesis of lipopolysaccharides relies on lpxD gene that encodes the third pathway enzyme. The roles of LpxD in Y. ruckeri virulence have not been studied. In the present study, in-frameshift deletion of lpxD gene and their role in Y. ruckeri virulence in rainbow trout were determined. For this purpose, 92% of the Y. ruckeri lpxD genes were deleted by homologous recombination. After running in SDS-PAGE and staining with silver stain, no LPS was detectable in the Y. ruckeri ΔlpxD mutant. Virulence and immunogenicity of the Y. ruckeri ΔlpxD mutant (YrΔlpxD) were determined in rainbow trout. Rainbow trout immunized with YrΔlpxD with immersion, or intraperitoneal injection method displayed superior protection (relative percentage survival ≥ 84%) after exposure to wild type Y. ruckeri. In conclusion, our results indicated that deletion of the lpxD gene causes significant attenuation of Y. ruckeri in rainbow trout, and LPS deficient YrΔlpxD could be used as a live attenuated vaccine against Y. ruckeri in rainbow trout. This vaccine can protect fish and it can be applied to fish with different methods such as immersion or injection.
鲁克氏耶尔森菌是一种革兰氏阴性细菌，可引起淡水和海水鱼类的耶尔森氏病。脂质 A 对革兰氏阴性细菌的发病机制很重要，其生物合成途径需要九种酶催化步骤。尽管细菌中有九个基因编码脂质 A 生物合成，但脂多糖的生物合成依赖于编码第三途径酶的 lpxD 基因。LpxD 在 Y. ruckeri 毒力中的作用尚未研究。在本研究中，通过同源重组对 lpxD 基因进行框移缺失，并确定其在虹鳟鱼中对 Y. ruckeri 毒力的作用。为此，通过同源重组删除了 92%的 Y. ruckeri lpxD 基因。在 SDS-PAGE 运行并使用银染染色后，在 Y. ruckeri ΔlpxD 突变体中检测不到 LPS。在虹鳟鱼中测定了 Y. ruckeri ΔlpxD 突变体（YrΔlpxD）的毒力和免疫原性。用 YrΔlpxD 免疫虹鳟鱼，通过浸泡或腹腔注射方法，在暴露于野生型 Y. ruckeri 后显示出优异的保护作用（相对存活率≥84%）。总之，我们的结果表明，lpxD 基因的缺失导致 Y. ruckeri 在虹鳟鱼中显著衰减，并且缺乏 LPS 的 YrΔlpxD 可作为虹鳟鱼中 Y. ruckeri 的活减毒疫苗。这种疫苗可以保护鱼类，可以应用于不同方法的鱼类，如浸泡或注射。
