猪胸膜肺炎放线杆菌脂多糖的制备及对小鼠的免疫保护作用

Preparation and application of lipopolysaccharide on Actinobacillus pleuropneumoniae

采用酚-水法制备血清3型、4型、5型、7型和8型猪胸膜肺炎放线杆菌(APP)脂多糖(LPS),以该多糖免疫小鼠,进行同源攻毒保护试验,结果LPS在20μg/只的免疫剂量下可对小鼠产生较强的保护作用,用同血清型菌株对免疫后的小鼠攻毒,仅表现肺脏轻微出血,无死亡,而对照组未经免疫直接攻毒的小白鼠全部死亡,肺脏严重出血;小鼠免疫后第2d就可以检测到抗体,并且抗体水平上升较快,到第6d抗体达到最高水平,之后,抗体水平开始下降,但下降幅度不大,可持续2个月左右;交叉保护试验结果表明血清3型LPS对血清5型和7型APP,血清4型LPS对血清5型和7型APP没有保护作用,免疫后的小鼠攻毒仍表现多数死亡;血清3型LPS对血清4型和8型APP有交叉保护作用,血清4型LPS对血清3型和8型APP有交叉保护作用,血清5型、7型、8型LPS对5个血清型的APP都有交叉保护作用,免疫后的小鼠攻毒无死亡,仅表现肺脏有不同程度的出血。上述结果表明LPS是APP的主要免疫保护性抗原之一,该研究为APP亚单位疫苗的研制及应用提供了理论依据。

The lipopolysaccharide of serotype 3, 4, 5, 7 and 8 on Actinobacillus pleuropneumoniae were collected and roughly purified by hydroxybenzene - water, then the extracted lipopolysaccharide were used as antigens to challenge the mouse with the same serotype bacteria, the results showed that the mouse were well protected immunized with 20 μg lipopolysaccharide. The mouse challenged with the same serotype bacteria were not dead at this dose , while the mouse of controled group were all dead. Two days after immunization, IgG was examined in the mouse. At the 6th day, the mouse exhibited significantly high IgG against lipopolysaccharide. Then the IgG slowly began to decline. In this research we found that the LPS of serotype 3 had no cross-reactions with APP of serotype 5 and 7, LPS of serotype 4 had no cross-reactions with APP of serotype 5 and 7, and most of the mice challenged with the different serotype bacteria were dead. While LPS of serotype 3 had cross-reactions with APP of serotype 4 and 8, LPS of serotype 4 had cross-reactions with APP of serotype 3 and 8, LPS of serotype 5,7 and 8 had cross-reactions with APP of all 5 serotypes, no mouse were dead after challenging with the different serotype bacteria. These results revealed that lipopolysaccharide would be one of the main protective antigens of Actinobacillus pleuropneumoniae. This research provided the foundation for preparation and application of the subunit vaccine on Actinobacillus pleuropneumoniae