Expression Of The Vp1 Protein Of Fmdv Integrated Chromosomally With Mutant Listeria Monocytogenes Strain

Expression of the VP1 protein of FMDV integrated chromosomally with mutant Listeria Monocytogenes Strain

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Sub-unit vaccines particularly live vector is a new approach to overcome the drawbacks of inactivated FMD vaccine such as improper inactivation during manufacture. Listeria monocytogenes; an intracellular microorganism that implicated with immune-stimulatory properties making it appropriate to utilize as a live bacterial vaccine vector. FMDV VP1 protein has the capability to induce both cellular and humoral immune responses since it considered the most immunogenic part of FMDV capsid and has the most of antigenic sites for viral neutralization. Codon-optimized vp1 gene was cloned into pCW702 vector to construct gene target cassette. That integrated successfully into the LM chromosome via homologous recombination for more stability and prevention of irreversible integrated gene. to generate a candidate vaccinal strain LM△actAplcB-vp1.

Safety evaluation of recombinant LM△actAplcB-vp1 revealed that it could be eliminated from internal organs within 3 days as a safe candidate vaccine. Mice groups were immunized I.V. twice with the recombinant LM△actAplcB-vp1 at intervals of 2 weeks. Antigen-specific IgG antibodies and the level of specific secreted cytokines of CD4+ and CD8+ T cells were estimated to evaluate the immunogenicity of the candidate vaccine. Rapid onset immune response was detected, strong IgG humoral immune response within 14 days post immunization and augmented again after the booster dose. Cellular immunity data after 9 days post the prime dose indicated elevation in the cytokines level with another elevation after the booster dose. This is the first report explain the ability of attenuated mutant LM to be a promising live vector for FMDV.Keywords: FMD, listeria, vector vaccine, recombinant, VP1.

Foot-and-mouth disease (FMD) is a terrible viral infection disease, attacks cloven-hoofed animals, including domestic livestock such as cattle, pigs and sheep. FMD is enzootic in Africa, Asia, and South America. The disease is highly contagious and outbreaks severely affect the economy through the loss of production, trade, and tourism in affected regions and present a constant threat for FMD-free countries(Clavijo et al. 2004; Perry and Rich 2007). Foot-and-mouth disease virus (FMDV), the etiologic agent of FMD, is a non-enveloped, positive-single stranded RNA virus, belonging to the genus of Aphthovirus, The full length of FMDV is about 8.4 kb. It has seven significant serotypes that are not immunologically related(Jamal and Belsham 2013).

FMDV fragment has a four-part structural protein and an eight-part non-structural protein(Du et al. 2007). VP1 as a structural protein is the major immunogenic protein of FMDV as it contains both T and B cell epitopes which trigger the immune system to induce cellular immune response as well as sufficient levels of neutralizing antibodies for protection against infection (Samuel and Knowles 2001). The traditional inactivated FMD vaccine currently is used in most countries to protect and eradicate the disease. Although it plays a great role in providing immune protection but it has several drawbacks; problems in manifestation procedures such as the improper viral inactivation, also the Vaccinal antigen need more purification process in order to prevent undesirable allergic reaction and allow the differentiation be tween vaccinated and infected animals throughout control management, in addition; it couldn’t minimize the shedding of FMD virus since it takes a long time to outset of immunity. For all of this, researchers are trying to develop alternative vaccines to overcome these problems (Cao et al. 2017; Doel 2003).

In previous reports, peptide vaccines based on immunogenic epitopes were developed to overcome the shortcomings of conventional FMD vaccines. However, peptide vaccines couldn’t elicit adequate immune responses alone; it must be used with some specific adjuvants (Li et al. 2007). Live vectored vaccines are the most attractive type among these modern vaccines since it combines the components of a subunit or DNA vaccine with the immunogenic characteristics of a living replicating organism to be more like an adjuvant. A potent attenuated bacterial strain as a vaccine vector has been established for its advantages over conventional vaccines or subunits vaccines. It has the ability to deliver heterologous antigens through its coding gene.

The concept beyond this approach is using the immunological features of the vector to trigger an immune response against both its own and inserted heterologous antigen genes. Typically, this vaccine type has the ability to stimulate the immune system, as it happened in natural infections, to stimulate CD4+ and CD8+ T-cell subsets. the disadvantage of this approach is that the intensity of the immune response is directly related to the pathogenicity and ability of the vector to be replicated and it could be overcome via attenuation of the used vector (Nascimento and Leite 2012). Listeria monocytogenes (LM), an intracellular facultative microorganism, possesses a wide range of mammalian hosts. The capability of LM to invade cytosol of the host cell after phagocytosis and deliver the heterologous antigen to the cytoplasm makes it a potent vaccine vector to conduct a specific immune response (Bierne et al. 2018; Dussurget et al. 2004; Saklani-Jusforgues et al. 2003).

11 February 2020

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