猪A群轮状病毒SCJY-13株的分离鉴定及致病性分析

【目的】 在从腹泻仔猪粪便中分离鉴定猪A群轮状病毒(Rotavirus group A, RVA)并了解其致病性。【方法】 应用MA-104细胞从仔猪腹泻粪便中分离鉴定RVA, 将其经口感染健康初生仔猪, 观察其临床症状, 并利用实时荧光定量PCR检测排毒、HE染色观察病理变化、免疫组织化学(immunohistochemistry, IHC)试验了解病毒分布。【结果】 分离到1株可引起MA-104细胞明显细胞病变的G9P23] RVA, 命名为RVA/Pig-tc/CHN/SCJY-13/2017/G9P23](简称SCJY-13株), 病毒滴度为10^5.5 TCID₅₀/100 μL。经口感染SCJY-13株的仔猪在感染后11 h出现腹泻, 持续到感染后165 h完全恢复, 其发病率为100%(7/7), 病死率为28.57%(2/7)。感染后8 h可从仔猪肛拭子检测到病毒核酸, 直至感染后192 h, 峰值出现在感染后24 h。感染后54 h各段小肠病毒载量达到最高, 其中回肠中病毒载量最高, 显著高于十二指肠、空肠(P<0.05);HE染色和IHC检测结果显示, SCJY-13在感染仔猪小肠的绒毛及隐窝中大量聚集, 尤其是回肠段; SCJY-13株感染引起十二指肠、空肠和回肠绒毛固有层大量淋巴细胞浸润、黏膜上皮细胞和肠绒毛尖端空泡变性、柱状细胞增多、绒毛断裂脱落等, 以回肠段较严重。【结论】 SCJY-13株能引起新生仔猪100%发病, 其主要靶位区在回肠, 感染后排毒时间长。本试验结果为猪RVA的致病性研究提供了一定的参考依据。

Isolation,Identification and Pathogenicity Analysis of Porcine Rotavirus Group A Strain SCJY-13

【Objective】 This study was aimed to isolate and identify porcine Rotavirus group A (RVA) from feces of piglets suffered from diarrhea, and understand the pathogenicity of the isolate strain.【Method】 RVA was isolated and identified from piglet diarrhea feces by MA-104 cells, and orally infected healthy newborn piglets.The clinical symptoms were observed, the detoxification was detected by Real-time quantitative PCR, the pathological changes were observed by HE staining, and the distribution of virus was tested by immunohistochemistry (IHC).【Result】 A strain of G9P23] RVA, named RVA/Pig-tc/CHN/SCJY-13/2017/G9P23] (SCJY-13 strain for short), which could cause obvious cytopathy of MA-104 cells, was isolated, and the virus titer was 10^5.5 TCID₅₀/100 μL.Diarrhea occurred at 11 h post infection and lasted until 165 h post infection.The incidence rate was 100% (7/7), and the mortality rate was 28.57% (2/7).Viral nucleic acid could be detected from piglet anal swabs at 8 h post infection until 192 h post infection, and the peak appeared at 24 h post infection.At the same time, the viral load in each segment of small intestine reached the highest 54 h post infection, and the viral load in ileum was the highest, which was significantly higher than that in duodenum, jejunum and feces (P < 0.05).HE staining and IHC detection showed that SCJY-13 accumulated in villi and crypts of small intestine of infected piglets, especially in ileum.SCJY-13 strain infection caused a large number of lymphocyte infiltration in duodenum, jejunum and ileum villus lamina propria, mucosal epithelial cells and vacuolar degeneration at the tip of intestinal villus, increase of columnar cells, villus breaking and falling off, especially in ileum.【Conclusion】 The SCJY-13 strain could cause 100% morbidity of neonatal piglets and shed virus for a long time, and the virus was mainly distributed in the ileum of small intestine.These results provided a valuable reference for the pathogenicity study of procine RVA.