Assessment of the acid phosphatase CP01850 from Corynebacterium pseudotuberculosis in DNA and subunit vaccine formulations against caseous lymphadenitis

Abstract

The target cp1002_RS01850 from Corynebacterium pseudotuberculosis was used to construct a DNA and recombinant subunit vaccine against caseous lymphadenitis. Recombinant protein rCP01850 was expressed in Escherichia coli using pAE vector, and DNA vaccine was engineered with pTARGET vector. BALB/c mice were divided in five groups containing eight animals each, inoculated with: pTARGET/cp01850 as DNA vaccine (G1); rCP01850 plus Al (OH)3 as recombinant subunit vaccine (G2); pTARGET/cp01850 and a boost with rCP01850 plus Al (OH)3 (G3); pTARGET (G4); or Al (OH)3 (G5). Mice were inoculated and blood samples were collected on days 0, 21, and 42 for the analysis of total IgG, IgG1 and IgG2a by ELISA. In each group, five animals were challenged with Mic-6 C. pseudotuberculosis strain, and three were used for cytokine quantification by qPCR. Although no group has been protected by vaccines against lethal challenge, G2 showed an increase in the survival rate after challenge. Significantly higher levels of IL-4, IL-12, IFN-γ, total IgG, IgG1 and IgG2a were also detected for G2, evidencing a mixed Th1/Th2 immunological profile. In conclusion, despite no protection level provided by different vaccinal strategies using cp1002_RS01850 from C. pseudotuberculosis, G2 developed a Th1/Th2 immune response with an increase in survival rate.

Vaccines for caseous lymphadenitis: up-to-date and forward-looking strategies

“…bovis BCG expressing pld – I.P 77 Mice (Leal et al 2018 ) M . bovis BCG expressing pld + rPLD boost 88 rCP01850 A.H I.M 0 Mice (Rezende et al 2020 ) pTARGET/ cp01850 – pTARGET/ cp01850 + rCP01850 boost A.H rCP01850 BRPHE S.C 70 Mice (Bezerra et al 2020 ) The table shows the vaccinal formulation, route of administration, protection rate, and animal model used in each experiment CP , Corynebacterium pseudotuberculosis ; S . C , subcutaneous; I .…”

“…This strategy uses only a part of the pathogen for immune recognition; however, the high purity level decreases its immunogenicity, requiring adjuvants to improve the immune response (Christensen 2016 ). Due to this fact, different adjuvants have already been tested in combination to recombinant proteins: Freund’s complete adjuvant (Pinho et al 2009 ; Droppa-almeida et al 2016 ), mineral oil (Selim et al 2010 ), saponin (Silva et al 2014 ; Droppa-almeida et al 2016 ; Silva et al 2018 ), aluminum hydroxide (Brum et al 2017 ; Rezende et al 2020 ), and more recently, propolis (Bezerra et al 2020 ).…”

“…Interesting results were obtained with acid phosphatase rCP01850 from C . pseudotuberculosis , once when associated to aluminum hydroxide, a mixed Th1/Th2 profile was obtained with no significant protection (Rezende et al 2020 ). However, the combination of rCP01850 with hydroalcoholic extract of Brazilian red propolis improved protection to 70% and significant levels of IgG, IFN-gamma, and IL-10 were reported (Bezerra et al 2020 ), confirming that testing different adjuvants with the same recombinant protein might generate completely different results.…”

“…Similarly, the intramuscular injection of pTARGET/ cp01850 was not able to protect mice against C . pseudotuberculosis challenge (Rezende et al 2020 ). Still, pVAX/ hsp60 failed to provide protection against the challenge in mice, despite the IgG-specific humoral immune response induced (Costa et al 2011 ).…”

“…Only in the last few years, modern strategies started to be tested in experimental level aiming to increase efficacy and safety of CLA vaccines. So, subunit recombinant, DNA, and vectored vaccines have been used in an attempt to induce proper immunity and high protection (Brum et al 2017 ; Leal et al 2018 ; Rezende et al 2020 ). Most of the researchers have been using IgG and IFN-γ as immunological markers for good quality CLA vaccines (Bastos et al 2012 ; Thais et al 2018 ).…”