RT Journal Article SR Electronic(1) A1 Ma, Jinzhu A1 Yu, Liquan A1 Song, Baifen A1 Yu, Yongzhong A1 Zhang, Shaoduo A1 Wei, Yuhua A1 Wu, Zhijun A1 Yao, Di A1 Yu, Wei A1 Zhu, Zhanbo A1 Cui, YudongYR 2018 T1 The double adjuvants LTB and CpG significantly enhanced the immuno-protective effects of recombinant GIT derived from Staphylococcus aureus and Streptococcus in mice JF Journal of Medical Microbiology, VO 67 IS 3 SP 432 OP 440 DO https://doi.org/10.1099/jmm.0.000666 PB Microbiology Society, SN 1473-5644, AB Purpose. In this study, we prepared GapC1-150-IsdB126-361-TRAP (GIT) proteins plus heat-labile enterotoxin B (LTB) as an intra-molecular adjuvant, together with CpG to further enhance its immunogenicity. Methodology. Initially, the target genes were acquired and inserted into pET-32a (+) vectors to express LTB–GIT protein. LTB–GIT expression was confirmed by Western blotting and its immunocompetence was estimated through ELISA. Further, we immunized BALB/c mice with the LTB–GIT plus CpG adjuvant. After the second immunization, the antigen-specific CD4+ cell responses for IFN-γ, IL-2, IL-4 and IL-10 were monitored by intracellular cytokine staining (ICS) assay. After the third immunization, the level of IgG antibodies in the serum from immunized groups was assessed by ELISA, and the protective immune response was appraised by Staphylococcus aureus and Streptococcus dysgalactiae challenge. Results. The ELISA results showed that the OD450nm value of the LTB–GIT group was significantly higher than that of the BSA group. The group immunized with LTB–GIT plus CpG exhibited significantly stronger CD4+ T cell responses for IFN-γ, IL-2, IL-4 and IL-10 compared to the group immunized with LTB–GIT, GIT alone orLTB–GIT plus CpG. In addition, the group immunized with LTB–GIT plus CpG generated the highest level of IgG antibodies against GIT among all of the groups, and our results also showed that LTB–GIT plus CpG markedly improved the survival percentage of mice compared to other groups. Conclusion. We confirmed that the novel double adjuvants, LTB and CpG, are able to significantly improve GIT-induced immune responses. This formula could be a promising strategy for enhancing the immune efficacy of multi-subunit vaccines against Staphylococcus aureus and streptococcal infection., UL https://www.microbiologyresearch.org/content/journal/jmm/10.1099/jmm.0.000666