撒坝猪源大肠杆菌高致病性毒力岛诱导病理损伤的超微结构特征

Ultrastructural Characteristics of Pathological Damage Induced by Escherichia coli High Pathogenicity Island from Saba Pig

为探究撒坝猪源大肠杆菌(E.coli)高致病性毒力岛(HPI)诱导小鼠病理损伤的超微结构特征,本研究将实验室保存的E.coli HPI阳性株(HPI^(+))和E.coli HPI基因缺失株(ΔHPI)进行复苏和培养,分别用E.coli HPI^(+)、E.coli^(Δ)HPI菌株以腹腔接种的方式感染昆明小鼠,检测菌株的半数致死量(50%lethal dose,LD50),通过HE染色和透射电镜观察并分析菌株对小鼠病理损伤的超微结构特征,利用免疫组织化学标记白细胞介素-1β(interleukin-1β,IL-1β)阳性细胞在被感染小鼠肝脏和肾脏组织中的分布,以反映E.coli HPI+及E.coliΔHPI菌株所引起炎症水平的差异。结果显示,E.coli HPI^(+)和E.coli^(Δ)HPI菌株的半数致死量分别为1×10^(7.39)和1×10^(8.62)CFU/mL;HE染色显示,E.coli感染小鼠后,可见肝脏细胞肿胀、变性,肝窦淤血,肾脏间质淤血,肾小管上皮细胞变性脱落等病理变化;超微结构变化显示,肝脏细胞的完整形态消失,胞核呈不规则形态,线粒体畸形,粗面内质网上核糖体脱落,滑面内质网增生;多数肾小管上皮细胞出现胞核固缩,部分细胞核核仁边移、体积增大,足突融合,系膜细胞间隙变宽。此外,E.coli HPI^(+)感染组小鼠于肝脏、肾脏的水肿现象较E.coli^(Δ)HPI菌株感染的小鼠更为明显。免疫组化结果显示,大肠杆菌感染小鼠后,IL-1β蛋白主要表达于肝细胞、中央静脉周围、肾间质细胞和肾小管上皮细胞,且E.coli HPI^(+)感染组的IL-1β表达量高于E.coli^(Δ)HPI感染组。综上所述,撒坝猪源E.coli HPI能够调控E.coli对小鼠的致病性,HPI的调控作用可使E.coli对小鼠肝脏、肾脏造成的病变及超微结构变化更明显,并且能够增加小鼠的炎症反应。

In order to explore the ultrastructural characteristics of pathological damage in mice induced by Escherichia coli(E.coli)highly pathogenic virulence island(HPI)from Saba pig,the E.coli HPI positive strain(HPI^(+))and E.coli HPI gene deletion strain(ΔHPI)preserved in the laboratory were resuscitated and cultured.Then the Kunming mice were infected with E.coli HPI^(+)and E.coli^(Δ)HPI strains by intraperitoneal inoculation,and the 50%lethal dose(LD50)of the strains was detected.The HE staining and transmission electron microscopy were used to observe and analyze the ultrastructural characteristics of pathological damage in mice.Immunohistochemistry was used to mark the distribution of interleukin-1β(IL-1β)positive cells in the liver and kidney tissues of infected mice to reflect the difference in inflammation levels caused by the strain E.coli HPI^(+)and E.coli^(Δ)HPI.The results showed that the LD_(50) of E.coli HPI^(+)and E.coli^(Δ)HPI strains were 1×10^(7.39)and 1×10^(8.62)CFU/mL,respectively.HE staining showed that after E.coli infection in mice,pathological changes,such as swelling and degeneration of liver cells,hepatic sinus congestion,renal interstitial congestion,renal tubular epithelial cells degeneration and shedding,were observed.The ultrastructural changes showed that the complete morphology of liver cells disappeared,the nuclei showed irregular morphology,mitochondrial abnormalities,ribosome shedding on rough endoplasmic reticulum,and smooth endoplasmic reticulum hyperplasia.Most renal tubular epithelial cells showed nucleus pyknosis,some of the nucleoli were shifted and the volume increased,the foot processes were fused,and the mesangial cell gap was widened.In addition,the edema in liver and kidney of E.coli HPI^(+)infection group was more obvious than that of mice infected with E.coli^(Δ)HPI strain.The results of immunohistochemistry showed that IL-1βwas mainly expressed in hepatocytes and pericentral veins,renal interstitial cells and renal tubular epithelial cells after E.coli infection in mice,and IL-1βexpression in E.coli HPI^(+)infection group was higher than that of the E.coli^(Δ)HPI infection group.To sum up,E.coli HPI from Saba pig could regulate the pathogenicity of E.coli to mice.The regulation of HPI could make the pathological changes and ultrastructural changes of liver and kidney of mice more obvious,and increase the inflammatory response of mice.