表达ETEC F41重组干酪乳杆菌口服及滴鼻免疫效果的比较

Comparison of immune response after oral and intranasal immunization with recombinant Lactobacillus casei expressing ETEC F41

摘要:

摘要：【目的】比较小鼠滴鼻与口服接种ETEC F41重组干酪乳杆菌后，机体所产生的抗ETEC F41的黏膜免疫和系统免疫及免疫保护率的差异，为确定ETEC乳酸菌疫苗免疫程序奠定基础。【方法】将构建的重组质粒pLA-F41电转化入干酪乳杆菌，获得阳性重组菌。重组菌在MRS 培养基中进行表达，经Western blot检测目的蛋白的表达，间接免疫荧光及流式细胞术检测外源蛋白展示到菌体表面。SPF级BALB/c小鼠随机分成4组，每组40只，将重组菌以滴鼻和口服途径分别接种2组小鼠，对照组分别接种同剂量的空质粒菌

Abstract:

Abstract:【Objective】In order to represent a promising strategy for mucosal vaccination, oral or intranasal immunization of Specific Pathogen Free (SPF) BALB/c mice were performed. The mucosal immunity, systemic immune and protective immune responses were compared after immunization with the recombinant Lactobacillus casei (L.casei) harboring enterotoxigenic Escherichia coli (ETEC) F41.【Method】The recombinant fusion proteins were detected by Western blot. Surface localization of the fusion protein was verified by immunofluorescence microscopy and flow cytometry. Six-week-old female SPF BALB/c mice (160 heads) were divided into 4 groups for immunization and control. Oral and intranasal immunization of mice was performed with the recombinant strain L. casei harboring pLA-F41or pLA. For oral immunization, the mice were inoculated daily on days 0 to 4, 7 to 11, 21 to 25, and 49 to 53. A lighter schedule was used for nasal immunization (days 0 to 2, 7 to 9, 21and 49). Specific anti-F41 IgG antibody in the serum and specific anti-F41 secret immunoglobulin A (sIgA) antibody in the lung, intestines, vagina fluid and feces of mice were detected by indirect ELISA. The mice orally or intranasally immunized with pLA-F41 /L. casei and pLA/L. casei were challenged with standard-type ETEC F41 (C83919) (2×103LD50).【Result】Mice immunized with pLA-F41/L.casei could produce remarkable anti-F41antibody level. More than 90% survived in oral immunization group whereas more than 85% survived in intranasal immunization group after challenged with C83919, all dead in the control group. Ninety percent of the pups survived in oral immunization group whereas 80% survived in intranasal immunization group after challenged with C83919, but only a 5% survival rate for pups that were either immunized with a control pLA vector or unimmunized.【Conclusion】Oral or intranasal immunization with recombinant L. casei displaying ETEC F41 antigens on the surface induced effective and similar systemic and mucosal immune responses against the ETEC F41.