Evaluation of a yeast-based BVDV vaccine approach

牛病毒性腹泻病毒(BVDV)已被认为是养牛业的主要经济疾病风险，导致了一系列相关问题。BVDV感染的控制对养牛业具有重要的经济意义，因为该病毒会引起一系列临床疾病和生殖障碍，以及潜在的继发性感染，如乳房炎，对生产周期的所有阶段都有不利影响，并且根据国家的不同，制定了各种BVDV根除计划(RidPath，2013)。生产损失主要源于繁殖失败和急性BVDV感染期间的免疫抑制，这使犊牛容易患呼吸系统或肠道疾病。事实上，BVDV感染对先天性和获得性免疫系统的影响是独一无二的，并导致严重的免疫功能障碍。仅接种疫苗不足以预防BVDV对胎儿的感染和损失。这是因为没有一种BVDV疫苗被证明能提供完全的胎儿保护。尽管人们对生物安全措施和PI根除计划的认识有所提高，但在许多国家，疫苗接种仍然是控制BVDV感染的最重要的控制策略(RidPath，2013)。由于BVDV毒株之间存在异质性，提供针对BVDV感染的获得性免疫保护具有挑战性。改良的活疫苗和灭活疫苗都被证明是有效的。体液和细胞免疫反应都是保护性的。在自然感染或改良活疫苗免疫后，大多数血清抗体是针对病毒蛋白E2和NS2/3的，对Erns和E1蛋白的反应很少。目前的BVDV疫苗需要注射和冷链，到目前为止，所有商业上可用的疫苗都是在细胞培养中生产的。在这里，新的口服疫苗可能提供一种更方便的给药途径，因此如果证明与目前的可注射疫苗一样具有保护作用，将具有极大的优势。酿酒酵母(S.cerevisiae，S.cerevisiae)是一种非侵袭性、非致病的生物，通常被认为是“通常被认为是安全的”(GRAS)物种，因此，它是一种有吸引力的运送抗原和治疗分子的工具(Chen等人，2005；Garrait等人，2007)。事实上，仅有5个表达卵白蛋白的重组酿酒酵母颗粒被证明能够诱导树突状细胞为基础的T细胞增殖，其程度与饱和数量的卵白蛋白多肽相同。伴随而来的是CD80、CD86、CD40、CD54和MHCII的上调以及树突状细胞产生IL-12的增加(Stubbset等人，2001年)。与在小鼠中产生的这些数据类似，Lu Etal和Franzusoff等人的工作已经证明，与表达肿瘤抗原的灭活酿酒酵母免疫相比，与单独使用抗原相比，免疫可引起更强的反应，并且与使用其活的对应物进行免疫所产生的抗肿瘤反应相同(Ardiani等人，2010年；Franzusoff等人，2005年；Lu等人，2004年)。事实上，利用酿酒葡萄球菌作为疫苗载体，可以通过给药到粘膜部位，包括口腔、呼吸道和生殖道，有效地诱导粘膜免疫和系统免疫(Allnutt等人，2007年；Sasagawa等人，2005年；Shin等人，2005年)。特别是在哺乳动物中，酿酒酵母已被证明具有更好的安全性和有效性，由于其细胞壁中存在TLR配体(Ogra等人，2001)，以及由于其大小的抗原性，它被证明能激发先天和获得性免疫反应。因此，我们建议评估以非分泌形式在其表面表达BVDV E2和/或NS2/3蛋白的重组酿酒酵母作为口服接种疫苗的价值。

Bovine Viral Diarrhoea Virus (BVDV) has been implicated as a majoreconomic disease risk for the cattle industry, resulting in a variety ofassociated problems. Control of BVDV infection is economicallyimportant to the cattle industry because the virus causes a varietyof clinical diseases and reproductive disorders, as well as potentialsecondary infections such as mastitis that adversely affectessentially all stages of the production cycle, and depending on thecountry, various BVDV eradication programs are in place (Ridpath,2013). Production losses primarily stem from reproductive failureand from immunosuppression during acute BVDV infection, whichpredisposes calves to respiratory or enteric diseases. Indeed, theeffects of a BVDV infection on the innate and acquired immunesystem are unique and result in dramatic immune dysfunction.Vaccination alone is not sufficient to protect against foetal infectionand losses due to BVDV. This is because no single BVDV vaccine hasbeen shown to give complete foetal protection. Despite increasedawareness regarding bio-security measurements and PI eradicationprograms, vaccination is still the most important control strategy forcontrolling BVDV infections in many countries (Ridpath, 2013).Providing acquired immune protection against infection with BVDVis challenging due to the heterogeneity that exists among BVDVstrains. Both modified live and killed vaccines have been shown tobe efficacious. Both humoral and cellular immune responses areprotective. Following natural infection or vaccination with amodified live vaccine, the majority of the serum antibodies aredirected against the viral proteins E2 and NS2/3, with minorresponses against the Erns and E1 proteins. Current BVDV vaccinesrequire injection and a 'cold chain', and to date all commerciallyavailable vaccines are produced in cell culture.Here, new oral vaccines may provide a more convenient route ofadministration, and thus being of great advantage, if shown to be asprotective as the current injectable ones. Saccharomyces cerevisiae(S. cerevisiae), a non-invasive, non-pathogenic organism, is generallyconsidered to be a "generally regarded as safe" (GRAS) species andfor this reason, it is an attractive tool for delivering antigens andtherapeutic molecules (Chen et al., 2005; Garrait et al., 2007).Indeed, as little as 5 recombinant yeast particles of S. cerevisiaeexpressing ovalbumin have been shown to induce dendriticcell-based T cell proliferation to the same degree as those primedwith saturating amounts of ovalbumin peptide. This wasaccompanied by up-regulation of CD80, CD86, CD40, CD54 and MHCII as well as an increase in IL-12 production by dendritic cells (Stubbset al., 2001). Similar to these data generated in mice, work by Lu etal and by Franzusoff et al in humans has demonstrated thatimmunisation with inactivated S. cerevisiae expressing tumourantigens elicits a far stronger response compared to antigen alone,and was equal to the anti-tumour response generated byimmunisation using their live counter-parts (Ardiani et al., 2010;Franzusoff et al., 2005; Lu et al., 2004). Indeed, S. cerevisiae used asa vaccine vehicle can effectively elicit mucosal and systemicimmunization by administration to mucosal sites, including the oral,respiratory and genital tracts (Allnutt et al., 2007; Sasagawa et al.,2005; Shin et al., 2005). In mammals in particular, S. cerevisiae hasbeen demonstrated to have superior safety and efficacy, where ithas been shown to elicit both innate and adaptative immuneresponses due to the presence of TLR ligands in its cell wall (Ogra etal., 2001) as well as its antigenic feature due to its size.Thus, we propose to assess the value of using recombinant S.cerevisiae yeast expressing BVDV E2 and/or NS2/3 proteins on itssurface in a non-secreted form as a vaccine candidate for oraladministration.