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

Date Published: December 4, 2014

Publisher: Public Library of Science

Author(s): Lalit Batra, Shailendra K. Verma, Durgesh P. Nagar, Nandita Saxena, Prachi Pathak, Satish C. Pant, Urmil Tuteja, Mary Ann McDowell.

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.

Partial Text: Plague caused by Y. pestis (a Gram negative bacterium) is a zoonotic infectious disease that has profoundly affected the course of history [1], [2] and troubles human populations, leading to millions of deaths. According to the World Health Organization (WHO), plague has been classified as a re-emerging infectious disease [3]. Rodents are the reservoirs for Y. pestis and the fleas transmit the bacteria from rodent to rodent. Infected fleas also transmit bubonic plague, the most common form of the disease from rodents to humans [4]–[6]. Humans are infected accidently after bites from fleas having Y. pestis, by direct contact with blood and tissues of infected animals, or by direct aerosol transmission. The aerosol transmission develops lethal pneumonic plague. The intentional aerosolization of Y. pestis in human population is the main concern of bioterrorism [7]. Plague can be treated with antibiotics at early stage. It has been reported that antibiotic-resistant strains of Y. pestis bacilli have been isolated in Madagascar and Mongolia [8], [9] and showed naturally acquired multi-drug-resistant variants of Y. pestis[10]. These studies suggest that there is an urgent need to develop an effective vaccine that can provide long term protection and to counter the drug resistant variants of Y. pestis.

Y. pestis suppresses the host immune system in susceptible animal species, but the infection survived animals can effectively overcome the re-infection. This hints the possibility of developing effective vaccine that can boost the immune defense mechanisms against plague. Although intensive studies are in progress for several decades on plague [46] there is no safe and efficient vaccine till date. The F1/V based subunit vaccine candidate that evokes mainly humoral immune response, although has shown promising results in animal models, its efficacy in humans is not yet evaluated [47]. Further, the next-generation plague vaccines that are yet to be developed should also evoke cell-mediated immune response [28]. Humoral and cellular immunity potentially contribute to vaccine efficacy [48]. Humoral immunity relies upon production of antibodies by plasma B cells which effectively neutralizes extracellular pathogens while cellular immunity relies upon cytokine-producing capacities of T cells and is particularly effective in eradicating intracellular pathogens [49].



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