Manipulation of the infectious bronchitis coronavirus genome for vaccine development and analysis of the accessory proteins.
Manipulation of the infectious bronchitis coronavirus genome for vaccine development and analysis of the accessory proteins.
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DOI：
10.1016/j.vaccine.2007.02.046
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发表时间：
2007-07-26
期刊：
影响因子：
5.5
通讯作者：
Britton P
中科院分区：
文献类型：
--
作者：
Cavanagh D;Casais R;Armesto M;Hodgson T;Izadkhasti S;Davies M;Lin F;Tarpey I;Britton P
传染性支气管炎冠状病毒（IBV）是英国家禽中经济损失最大的单一传染病的原因，在许多家禽业发达的国家可能也是如此。其持续优势的主要原因是其存在许多血清型，由表面刺突蛋白（S）决定，交叉保护性差。尽管在一定程度上受到活疫苗和灭活疫苗的控制，但仍需要新一代IB疫苗。反向遗传或“感染性克隆”系统允许操纵IBV基因组，是这一发展的关键。理想的新疫苗是：遗传上稳定的（即保持稳定的减毒表型）;卵内给药;以及对于刺突蛋白基因的来源是灵活的。IBV的合理减毒需要同时鉴定对复制不必需的基因，并且其缺失将降低致病性。能够修改“核心”疫苗株，使其适用于流行的血清型，需要这样做的程序，并证明“尖峰交换”足以诱导良好的免疫力。我们已经证明，由基因3和5编码的四种小的IBV蛋白对于复制不是必需的;不能产生这些蛋白对产生的病毒滴度几乎没有不利影响。我们目前的分子克隆IBV Beaudette株是非致病性的，因此我们不知道这些蛋白质的缺失会对致病性产生什么影响。也就是说，细胞培养物中各种基因3/5重组IBV的噬斑大小和组成，以及气管器官培养物中减少的输出和纤毛停滞，表明它们比野生型Beaudette具有更低的侵袭性。因此，这些基因仍然是合理减毒的靶标。我们最近获得的证据表明，基因1编码的15种蛋白质中的一种或多种也是致病性的决定因素。因此，基因1也是合理减毒的靶标。用致病性M41株的S蛋白基因替换Beaudette的S蛋白基因，产生了一种重组病毒，该病毒仍然是非致病性的，但确实诱导了对M41攻击的保护。此后，我们还制造了其他的“尖峰交换”重组体，包括带有嵌合体S基因的重组体。独特的是，我们的Beaudette分子克隆在给予18天大的胚胎时是良性的，即使是高剂量，并在这种接种途径后诱导免疫。综上所述，我们的研究结果表明，基于基因组的合理修饰，创造新一代IB疫苗是一个可实现的目标。
Infectious bronchitis coronavirus (IBV) is the cause of the single most economically costly infectious disease of domestic fowl in the UK—and probably so in many countries that have a developed poultry industry. A major reason for its continued dominance is its existence as many serotypes, determined by the surface spike protein (S), cross-protection being poor. Although controlled to some degree by live and inactivated vaccines, a new generation of IB vaccines is called for. Reverse genetic or ‘infectious clone’ systems, which allow the manipulation of the IBV genome, are key to this development. New vaccines would ideally be: genetically stable (i.e. maintain a stable attenuated phenotype); administered in ovo; and be flexible with respect to the source of the spike protein gene. Rational attenuation of IBV requires the identification of genes that are simultaneously not essential for replication and whose absence would reduce pathogenicity. Being able to modify a ‘core’ vaccine strain to make it applicable to a prevailing serotype requires a procedure for doing so, and the demonstration that ‘spike-swapping’ is sufficient to induce good immunity. We have demonstrated that four small IBV proteins, encoded by genes 3 and 5, are not essential for replication; failure to produce these proteins had little detrimental affect on the titre of virus produced. Our current molecularly cloned IBV, strain Beaudette, is non-pathogenic, so we do not know what effect the absence of these proteins would have on pathogenicity. That said, plaque size and composition of various gene 3/5 recombinant IBVs in cell culture, and reduced output and ciliostasis in tracheal organ cultures, shows that they are less aggressive than the wild-type Beaudette. Consequently these genes remain targets for rational attenuation. We have recently obtained evidence that one or more of the 15 proteins encoded by gene 1 are also determinants of pathogenicity. Hence gene 1 is also a target for rational attenuation. Replacing the S protein gene of Beaudette with that from the pathogenic M41 strain resulted in a recombinant virus that was still non-pathogenic but which did induce protection against challenge with M41. We have since made other ‘spike-swapped’ recombinants, including ones with chimaera S genes. Uniquely, our molecular clone of Beaudette is benign when administered to 18-day-old embryos, even at high doses, and induces immunity after this route of vaccination. Taken together, our results point to the creation of a new generation of IB vaccines, based on rational modification of the genome, as being a realisable objective.
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