利用大蜡螟幼虫和小鼠感染模型筛选猪链球菌血清2、3和9型三价灭活疫苗候选菌株  

Screening of Trivalent Inactivated Vaccine Candidate Strains of Streptococcus suis

作　　者：朋璐 张衡 庞思琪 乔竹林 张小芬 谭臣[1] 宋云峰[1] 周锐[1] 黎璐 PENG Lu;ZHANG Heng;PANG Siqi;QIAO Zhulin;ZHANG Xiaofen;TAN Chen;SONG Yunfeng;ZHOU Rui;LI Lu(National Key Laboratory of Agricultural Microbiology,The Cooperative Innovation Center for Sustainable Pig Production Co-sponsored by Province and Ministry,College of Veterinary Medicine,Huazhong Agricultural University,Wuhan 430070,China)

机构地区：[1]华中农业大学动物医学院,农业微生物资源发掘与利用全国重点实验室,生猪健康养殖省部共建协同创新中心,武汉430070

出　　处：《畜牧兽医学报》2024年第9期4077-4090,共14页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家重点研发计划(2021YFD1800404)。

摘　　要：旨在评估临床分离的猪链球菌血清2、3、9型强毒株制备的三价灭活疫苗对小鼠的免疫保护效果。使用大蜡螟幼虫感染模型和小鼠感染模型,从临床分离的猪链球菌中筛选出血清2、3和9型强毒株各1株(SS1803、SS1803024、SS1696),检测其生长曲线和毒力因子。将菌株分别灭活及混合灭活后与佐剂混合,用4×10^(7)CFU、8×10^(7)CFU SS1803、2×10^(8)CFU、5×10^(8)CFU SS1803024、1×10^(8)CFU、5×10^(8)CFU SS1696及三价灭活苗(2×10^(7)CFU SS1803^(+)1.5×10^(8)CFU SS1803024^(+)3×10^(7)CFU SS1696)分别对BALB/c小鼠进行两轮免疫,采集血清检测特异性抗体水平,分别用3种菌株致死剂量攻毒后观察小鼠免疫保护率,同时用亚致死剂量攻毒三价苗免疫组,测定小鼠血、脑、肺、脾的组织载菌量和细胞因子IL-6、IL-12水平,并观察脑、肺、脾、肾中的组织病理变化。结果显示:3株疫苗候选菌株的毒力基因鉴定分别为SS1803:gapdh^(+)/sly^(+)/fbps^(+)/orf2^(+)/mrp^(-)/89K^(-)/gdh^(+)/epf^(+),SS1803024:gapdh^(+)/sly^(+)/fbps^(+)/orf2^(+)/mrp^(-)/89K^(-)/gdh^(+)/epf^(-),SS1696:gapdh^(+)/sly-/fbps^(+)/orf2^(+)/mrp^(-)/89K^(-)/gdh^(+)/epf^(-)。单价和三价疫苗免疫组均能产生对应免疫血清型的IgG抗体,以产生IgG1型抗体为主。三价疫苗免疫组能对2、3、9型3个菌株攻毒分别提供83.3%、66.7%和66.7%的保护率,能显著降低感染后小鼠组织中的载菌量,降低血清炎性因子水平和组织病变程度。本研究筛选临床流行率高的2、3、9型强毒株,联合制备三价灭活疫苗,能够对小鼠提供良好的免疫保护力,为猪链球菌多价疫苗的研发提供新的材料。The aim was to evaluate the immune protective effect of a trivalent inactivated vaccine prepared from virulent strains of clinically isolated Streptococcus suis serotype 2,3 and 9 on mice.One virulent strain each of S.suis serotype 2,3 and 9 were screened from clinical isolates using Galleria mellonella and mice infection models(SS1803,SS1803024,SS1696),and the growth curves and virulence genes were tested.The strains were separately inactivated and mixed inactivated and mixed with adjuvants.4×10^(7)CFU,8×10^(7)CFU SS1803,2×10^(8)CFU,5×10^(8)CFU SS1803024,1×10^(8)CFU,5×10^(8)CFU SS1696,and trivalent inactivated vaccine(2×10^(7)CFU SS1803^(+)1.5×10^(8)CFU SS1803024^(+)3×10^(7)CFU SS1696)were used to immune the BALB/c mice in two rounds,serum was collected to detect the levels of specific antibodies and the immune protection rates of the mice were observed after challenge with lethal doses of the three strains respectively.Then,a sub-lethal dose were used to challenge the trivalent immunized group,the bacterial loads in blood,brain,lung and spleen and the levels of cytokines IL-6 and IL-12 of mice were measured,and the histopathological changes in brain,lung,spleen and kidney were also observed.The virulence genes of the three vaccine candidate strains were identified as:SS1803:gapdh^(+)/sly^(+)/fbps^(+)/orf2^(+)/mrp^(-)/89K^(-)/gdh^(+)/epf^(+),SS1803024:gapdh^(+)/sly^(+)/fbps^(+)/orf2^(+)/mrp^(-)/89K^(-)/gdh^(+)/epf^(-),SS1696:gapdh^(+)/sly-/fbps^(+)/orf2^(+)/mrp^(-)/89K^(-)/gdh^(+)/epf^(-).Both monovalent and trivalent vaccine immune groups induced IgG antibodies corresponding to the immunized serotypes,with the production of IgG1 antibodies predominating.The trivalent group provided 83.3%,66.7%and 66.7%protection against serotype 2,3 and 9 strains,respectively.It also effectively reduced the bacterial loads in the tissues,and reduced the levels of serum inflammatory factors and the degree of tissue lesions of mice after infection.In this study,highly virulent strains of serotype 2,3 and 9 with high clin

关 键 词：猪链球菌 疫苗候选菌株 三价灭活疫苗 免疫保护效果 

分 类 号：S852.611[农业科学—基础兽医学]