HSP70 Domain II of Mycobacterium tuberculosis Modulates Immune Response and Protective Potential of F1 and LcrV Antigens of Yersinia pestis in a Mouse Model

Abstract

No ideal vaccine exists to control plague, a deadly dangerous disease caused by Yersinia pestis. In this context, we cloned, expressed and purified recombinant F1, LcrV antigens of Y. pestis and heat shock protein70 (HSP70) domain II of M. tuberculosis in E. coli. To evaluate the protective potential of each purified protein alone or in combination, Balb/C mice were immunized. Humoral and cell mediated immune responses were evaluated. Immunized animals were challenged with 100 LD50 of Y. pestis via intra-peritoneal route. Vaccine candidates i.e., F1 and LcrV generated highly significant titres of anti-F1 and anti-LcrV IgG antibodies. A significant difference was noticed in the expression level of IL-2, IFN-γ and TNF-α in splenocytes of immunized animals. Significantly increased percentages of CD4+ and CD8+ T cells producing IFN-γ in spleen of vaccinated animals were observed in comparison to control group by flow cytometric analysis. We investigated whether the F1, LcrV and HSP70(II) antigens alone or in combination can effectively protect immunized animals from any histopathological changes. Signs of histopathological lesions noticed in lung, liver, kidney and spleen of immunized animals on 3rd day post challenge whereas no lesions in animals that survived to day 20 post-infection were observed. Immunohistochemistry showed bacteria in lung, liver, spleen and kidney on 3rd day post-infection whereas no bacteria was observed on day 20 post-infection in surviving animals in LcrV, LcrV+HSP70(II), F1+LcrV, and F1+LcrV+HSP70(II) vaccinated groups. A significant difference was observed in the expression of IL-2, IFN-γ, TNF-α, and CD4+/CD8+ T cells secreting IFN-γ in the F1+LcrV+HSP70(II) vaccinated group in comparison to the F1+LcrV vaccinated group. Three combinations that included LcrV+HSP70(II), F1+LcrV or F1+LcrV+HSP70(II) provided 100% protection, whereas LcrV alone provided only 75% protection. These findings suggest that HSP70(II) of M. tuberculosis can be a potent immunomodulator for F1 and LcrV containing vaccine candidates against plague.

Plague Vaccine Development: Current Research and Future Trends

“…Immunization with recombinant F1 imparts same degree of protection in mice against subcutaneous or pneumonic plague as does native F1, extracted from Y. pestis (26). In our studies (27), vaccination with recombinant F1 failed to protect mice against bubonic plague. Nevertheless, there exist some virulent strains of F1-negative Y. pestis ; hence, vaccines based exclusively on F1 are not worthwhile against any type of plague (28).…”

“…In a recent study, F1mut-V in formulation with alhydrogel and T4-decorated F1mut-V without any adjuvant imparted 100% protection in mice and rats against pneumonic plague (39). In our recent studies, we also demonstrated that a mixture of recombinant F1 + LcrV antigens in formulation with alum imparts full protection in mice against bubonic plague (27). In conclusion, F1/LcrV vaccine provides strong protective immunity in mice, rats, and rabbits against subcutaneous and pneumonic plague.…”

“…challenge of Y. pestis and Yersinia pseudotuberculosis in Swiss Webster mice (32, 33). We also reported that recombinant LcrV alone provided only 75% protection in mice against bubonic plague (27). The combination of recombinant F1 and LcrV antigens elicited greater protection in comparison to either F1 alone or LcrV alone (34, 35).…”

“…The protection is stimulated by anti-LcrV antibodies, which help by blocking the type 3 secretion system (40, 41). Inclusion of F1 with LcrV enhanced the protection as LcrV does not always provide 100% protection (27). …”

“…HSP70(II) significantly elevated the levels of IL-2, IFN-γ, TNF-α, and IFN-γ secreting CD4 + /CD8 + T cells in F1 + LcrV + HSP70(II) vaccinated group in comparison to the F1 + LcrV group. F1 + LcrV + HSP70(II) combination provided full protection against virulent strain of Y. pestis (S1, Indian clinical isolate) in a mouse model (27). Later, we designed a recombinant trivalent fusion protein F1–LcrV–HSP70(II) and evaluated in a mouse model.…”