高致病性维氏气单胞菌胞外产物对斑点鲖的致病性
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四川农业大学动物医学院,四川农业大学动物医学院 成都 温江;四川农业大学动物疾病与人类健康四川省重点实验室 成都 温江;四川农业大学动物科技学院水产养殖系 成都 温江,四川农业大学动物医学院,四川农业大学动物科技学院水产养殖系 成都 温江,四川农业大学动物医学院 成都 温江,四川农业大学动物医学院 成都 温江,四川农业大学动物医学院 成都 温江,四川农业大学动物医学院 成都 温江,四川农业大学动物医学院 成都 温江,四川农业大学动物医学院 成都 温江,四川农业大学动物医学院 成都 温江

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教育部“长江学者和创新团队发展计划”创新团队项目(IRT0848)


Investigation on the pathogenicity of extracellular products of a high virulence Aeromonas veronii strain isolated from Ictalurus punctatus
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Sichuan agricultural university,,Sichuan agricultural university,,,,,,,,

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    摘要:

    为了研究维氏气单胞菌及其胞外产物的致病机制对预防和治疗斑点鲖维氏气单胞菌病的作用。通过对一株高致病性斑点鲖源维氏气单胞菌胞外产物(extracellular product,ECP)的提取并结合体内外实验研究其酶活性与致病性,以期为维氏气单胞菌的致病机制研究提供新思路。采用饱和硫酸铵沉淀法,对一株高致病性的斑点鲖源维氏气单胞菌的ECP进行了提取,将其进行浓度检测、SDS-PAGE分析和酶活测定;并将提取的ECP攻毒斑点鲖,通过病理学分析研究其致病机制。提取的ECP经考马斯亮蓝试剂盒确定其浓度为0.743 mg/mL; SDS-PAGE分析发现该ECP类型丰富,分子大小主要集中在20.0~33.0 ku。运用琼脂平板扩散法对ECP中主要毒力和代谢相关酶活性进行体外测定,发现ECP具有脂酶、蛋白酶、卵磷脂酶、DNA酶和溶血活性,不具有淀粉酶、明胶酶、脲酶活性;并对与毒力密切相关的溶血活性进行了进一步的溶血谱绘制,发现ECP对其他多种动物红细胞都有溶血活性,对鱼类红细胞溶血性较强,但对鸡、鸭红细胞无溶血活性。 ECP攻毒健康斑点鲖后,发现其具有明显致病性,死亡率高达100%。病鱼大体病理变化为体表黏液增多,出现褪色斑以及不同程度的出血斑;眼球充血;肛门红肿外凸;剖检病变表现:鳃丝充血肿胀、肝脏肿大、散在少量出血点,脾脏肿大暗红,肠道扩张、肠壁变薄、肠腔内有大量黄色黏液。组织学病变主要表现为肝细胞变性、坏死;脾脏淋巴细胞减少,大量结缔组织增生;胃、肠道黏膜上皮坏死脱落。该株维氏气单胞菌的胞外产物具有多种酶活性,且对斑点鲖具有明显的致病性,推测该胞外产物在维氏气单胞菌入侵、感染甚至致死斑点鲖过程中都发挥了重要作用。

    Abstract:

    Aeromonas veronii is an acute epidemic pathogen of channel catfish (Ictalurus punctatus), causing obvious symptoms to hosts, such as distension of abdomen, enteritis and hemorrhage in multiple organs. The goal of this research was to reveal the pathogenicity of A. veronii extracellular products (ECP), and lay a foundation for vaccine and treatment. The pathogenicity of the extracellular products extracted from a high A. veronii isolated from I. punctatus was investigated in vitro and in vivo. In vitro, A. veronii extracellular products were used to detect the biologic activities and the hemolytic spectrum to a variety of erythrocyte. Then, an in vivo research on challenging the channel catfish with ECP was applied to confirm the pathogenicity. Biologic activities showed that the ECP possess lipase, protease, lecithase, DNAzyme, hemolysin; and the hemolytic effect of ECP showed a high hemolytic activity to fish erythrocyte, while no hemolytic activity in chicken and duck erythrocyte. The in vivo research on channel catfish showed that the ECP from the high A. veronii strain had an obvious pathogenicity to channel catfish, with a death rate of 100%. Diseased fish showed increasing mucus and fade spots on the skin, with eye congestion, red anus, liver hemorrhage, oedema of gill, rough surface of the spleen and intestinal expansion with thinning intestinal wall. Histopathological examination showed that edema, vacuolar degeneration, even necrosis in liver; reticuloendotheliosis spleen with few lymphocytes; and hemorrhage and necrosis of the lamella epithelia. The results of in vitro and in vivo showed that ECP of the A. veronii has a variety of extracellular enzymes, which may contribute to pathogenicity of the channel catfish.

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贺扬,华丽,汪开毓,陈德芳,王均,王二龙,王兴丽,杨倩,吴学婧,何洁,赖为民.高致病性维氏气单胞菌胞外产物对斑点鲖的致病性[J].水产学报,2016,40(3):457~467

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  • 收稿日期:2015-09-05
  • 最后修改日期:2015-12-08
  • 录用日期:2016-03-22
  • 在线发布日期: 2016-03-29
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