Evaluation of the immunogenicity of the P97R1 adhesin of Mycoplasma hyopneumoniae as a mucosal vaccine in mice

Department of Biological and Physical Sciences, University of Southern Queensland, Toowoomba, Queensland, Australia.
Journal of Medical Microbiology (Impact Factor: 2.25). 08/2006; 55:923-9.


The immunogenicity of P97 adhesin repeat region R1 (P97R1) of Mycoplasma hyopneumoniae, an important pathogenesis-associated region of P97, was evaluated in mice as a mucosal vaccine. Mice were immunized orally with attenuated Salmonella typhimurium aroA strain CS332 harbouring a eukaryotic or prokaryotic expression vector encoding P97R1. Local and systemic immune responses were analysed by ELISA on mouse sera, lung washes and splenocyte supernatants following splenocyte stimulation with specific antigens in vitro. Although no P97R1-specific antibody responses were detected in serum and lung washes, significant gamma interferon was produced by P97R1-stimulated splenocytes from mice immunized orally with S. typhimurium aroA harbouring either expression system, indicating induction of a cell-mediated immune response. These results suggested that live bacterial vectors carrying DNA vaccines or expressing heterologous antigens preferentially induce a Th1 response. Surprisingly, however, mice immunized with the vaccine carrier S. typhimurium aroA CS332 induced serum IgG, but not mucosal IgA, against P97R1 or S. typhimurium aroA CS332 whole-cell lysate, emphasizing the importance of assessing the suitability of attenuated S. typhimurium antigen-carrier delivery vectors in the mouse model prior to their evaluation as potential vaccines in the target species, which in this instance was pigs.

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Available from: Trilochan Mukkur, Oct 03, 2015
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    • "Live bacteria harboring vectors encoding either R1, or R1 and R2, have also been evaluated for their potential as vaccines against M. hyopneumoniae infection. Mice that orally ingested Salmonella typhimurium harboring eukaryotic or prokaryotic expression vector encoding the R1 region showed R1-specific Th1 responses [28]. Intranasal and oral immunization of Erysipelothrix rhusiopathiae strains expressing the C-terminal portion of P97, including the R1 and R2 regions, to pigs reduced the severity of pneumonic lung lesions caused by M. hyopneumoniae infection [29,30]. "
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    ABSTRACT: Infection by Mycoplasma hyopneumoniae and Actinobacillus pleuropneumoniae, either alone or together, causes serious respiratory diseases in pigs. To develop an efficient multi-disease subunit vaccine against these pathogens, we produced a chimeric protein called Ap97, which comprises a deletion derivative of the N-terminal region of the A. pleuropneumoniae ApxIII toxin (ApxN) and the R1 and R2 repeats of M. hyopneumoniae P97 adhesin (P97C), using an E. coli expression system.The levels of both IgG1 and IgG2a isotypes specific for ApxN and P97C in the sera of Ap97-immunized mice increased, and Ap97 induced the secretion of IL-4 and IFN-gamma by mouse splenocytes. Antisera from mice and pigs immunized with Ap97 readily reacted with both native ApxIII and P97 proteins. In addition, immunization with the Ap97 vaccine effectively protected pigs against challenge with both pathogens. These findings suggest that Ap97 confers immunogenicity, and is an effective vaccine that protects pigs against infection by M. hyopneumoniae and A. pleuropneumoniae.
    BMC Veterinary Research 02/2014; 10(1):43. DOI:10.1186/1746-6148-10-43 · 1.78 Impact Factor
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    • "The C-terminal portion of P97 contains two repeat regions (R1 and R2), and the cilium binding site is located in the R1 region (AAKPV/E) (Minion et al., 2000). In the last two decades, different strategies have been used to develop vaccines using P97 (King et al., 1997; Shimoji et al., 2003; Chen et al., 2006, 2008; Conceicao et al., 2006; Ogawa et al., 2009; Okamba et al., 2010). Nevertheless , subunit vaccine production using Escherichia coli has several advantages over other methods. "
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    ABSTRACT: The surface adhesin P97 mediates the adherence of Mycoplasma hyopneumoniae to swine cilia. Two reiterated repeats R1 and R2 are located at the C-terminus of P97. The purpose of this study was to evaluate the immunogenicity of Montanide adjuvant IMS 1113 plus soluble subunit proteins rR1, rR1R2 and their chimeric forms coupled with B subunit of the heat-labile enterotoxin of Escherichia coli (LTB). Each recombinant protein in this study was capable of eliciting anti-R1 specific humoral antibodies (IgG), mucosal antibodies (IgG and IgA) and INF-γ production. The chimeric protein rLTBR1R2 elicited the quickest humoral antibody response among the recombinant proteins. Serum and bronchoalveolar lavage analysis revealed that each recombinant protein was capable of inducing both Th1 and Th2 responses. Importantly, all of the proteins induced an anti-R1-specific Th2-biased response in both humoral and mucosal compartments, similar to the response observed in a natural infection or vaccination process. These observations indicate that rR1, rR1R2, rLTBR1 and rLTBR1R2 with IMS 1113 might represent a promising subunit vaccine strategy against porcine enzootic pneumonia in pigs. This article is protected by copyright. All rights reserved.
    FEMS Microbiology Letters 01/2014; 352(2). DOI:10.1111/1574-6968.12389 · 2.12 Impact Factor
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    • "Against swine enzootic pneumonia (SEP), caused by Mycoplasma hyopneumoniae, plasmid DNA coding the heat shock protein gene (P42) should be a suitable candidate as it is capable of inducing both Th1 and Th2 immune responses. Against SEP, the capability of P97 adhesin repeat region of M. hyopneumoniae to produce immunogenicity in mice in DNA vaccine formulation has also been described (Chen et al. 2006). However, considerable research has been directed towards developing DNA vaccines to prevent swine fever, FMD and psuedorabies infection in pigs. "
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    ABSTRACT: Inoculation of plasmid DNA, encoding an immunogenic protein gene of an infectious agent, stands out as a novel approach for developing new generation vaccines for prevention of infectious diseases of animals. The potential of DNA vaccines to act in presence of maternal antibodies, its stability and cost effectiveness and the non-requirement of cold chain have heightened the prospects. Even though great strides have been made in nucleic acid vaccination, still there are many areas that need further research for its wholesome practical implementation. Major areas of concern are vaccine delivery, designing of suitable vectors and cytotoxic T cell responses. Also, the induction of immune responses by DNA vaccines is inconclusive due to the lack of knowledge regarding the concentration of the protein expressed in vivo. Alternative delivery systems having higher transfection efficiency and the use of cytokines, as immunomodulators, needs to be further explored. Recently, efforts are being made to modulate and prolong the active life of dendritic cells, in order to make antigen presentation a more efficacious one. For combating diseases like acquired immunodeficiency syndrome (AIDS), influenza, malaria and tuberculosis in humans; and foot and mouth disease, Aujesky's disease, swine fever, rabies, canine distemper and brucellosis in animals, DNA vaccine clinical trials are underway. This review highlights the salient features of DNA vaccines, and measures to enhance their efficacy so as to devise an effective and novel vaccination strategy against animal diseases.
    Veterinary Research Communications 07/2008; 32(5):341-56. DOI:10.1007/s11259-008-9040-3 · 1.24 Impact Factor
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