Construction and biological characterization of a dual sagA/aroA gene deletion mutant strain of donkey-derived Streptococcus equi subsp. equi
10.16303/j.cnki.1005-4545.2025.11.09
- VernacularTitle:驴源马链球菌马亚种sagA和aroA双基因缺失菌株的构建及其生物学特性分析
- Author:
Bing LIU
1
;
Guangyuan LIU
;
Nannan GAO
;
Zhaoliang DING
;
Jie YU
;
Chuanlu WEI
;
Haijing LI
;
Hua WANG
;
Shishan DONG
;
Jianbao DONG
Author Information
1. 河北农业大学动物医学院,河北保定 071001;东阿阿胶股份有限公司国家胶类中药工程技术研究中心,山
- Publication Type:Journal Article
- Keywords:
strangles;
Streptococcus equi subsp. equi;
gene knockout;
attenuated strain;
vaccine
- From:
Chinese Journal of Veterinary Science
2025;45(11):2365-2371
- CountryChina
- Language:Chinese
-
Abstract:
Strangles,caused by Streptococcus equi subsp.equi,remains one of the most prevalent and high-incidence infectious diseases in intensive donkey farms,posing a significant threat to the healthy development of the donkey industry.Vaccination serves as an effective measure for the pre-vention and control of the disease,however,there is currently no attenuated vaccine against this disease in China.To provide a candidate strain for the development of a live attenuated strangles vaccine,this study focused on a wild-type S.equi subsp.equi strain isolated from donkeys.Using homologous recombination gene knockout technology,the aroA gene(encoding 5-enolpyru-vylshikimate-3-phosphate synthase)and the sag A gene(encoding the precursor of streptolysin S toxin)were sequentially deleted to construct a double-gene-deletion strain(ΔsagA/aroA).The virulence and key biological characteristics of the mutant strain were systematically evaluated.TheΔsagA/aroA strain was successfully generated,exhibiting complete loss of hemolytic activity and maintaining stable genetic inheritance over 60 consecutive passages.Electron microscopy revealed that the mutant retained morphological characteristics compared to the wild-type strain,and its growth rate was significantly slower(P＜0.000 1).Virulence assessment using a challenge dose of 1× 105 CFU/0.2 mL(the minimum fully lethal dose of the wild-type strain)demonstrated markedly attenuated virulence in the mutant.Immunization trials with 1 ×104 CFU/0.2 mL of theΔsagA/aroA strain revealed a increase in ELISA antibody titers by day 7 post-vaccination,and higher levels at days 14 and 21.Notably,antibody levels in the experimental group were significant-ly higher than those in the control group(P＜0.000 1).These findings confirm that the double-gene-deletion strain S.equi subsp.equi ΔsagA/aroA exhibits reduced virulence while retaining im-munogenicity,which suggested it can be used as a promising candidate strain for further develop-ment of a live attenuated strangles vaccine.
