Understanding the early pathogenesis of necrotic enteritis in chickens

了解产气荚膜梭菌的早期发病机制和毒力越来越大的消费者压力迫使畜牧业开发替代品，或至少大幅减少抗生素的使用量。我们必须找到有效的工具，以防止坏死性肠炎，一种致命的疾病，从影响无寄生虫鸡。这种疾病是由产气荚膜梭菌的致病菌株引起的，但诱发因素是必要的，以创造一个肠道环境，有利于增加C。产气荚膜杆菌（Cp）。疾病过程的另一部分被认为依赖于NE引起菌株从鸡肠道中取代非致病性大肠杆菌分离株的能力。许多革兰氏阳性菌已被证明产生各种蛋白质类的有毒化合物，称为细菌素，并抑制密切相关的菌株的生长。已在致病性netB阳性NE致病菌株中鉴定出Perfrin，并认为其发挥重要作用。在琼脂斑点试验中，我们观察到并非所有的netB阳性Cp菌株都携带perfrin基因，产生了除perfrin以外的细菌素，并且一些细菌性Cp菌株可以抑制致病性Cp菌株的生长。我们开发了一种体内结扎肠袢模型，在鸡中重现坏死性肠炎，并注意到病变严重程度与肠绒毛内衬Cp杆的数量相关（Parent et al，2016）。因此，Cp附着到肠上皮细胞可能在NE的发病机制中发挥作用，因此我们对IV型菌毛蛋白感兴趣，IV型菌毛蛋白已知在粘附细胞表面中发挥作用。我们最近发现，这些菌毛蛋白也是免疫原性的，当注射到鸟类。所有这些最近的发现，导致了这个建议的研究计划的发展。拟议的工作是创新的，使用一个独特的Cp菌株库，可以访问一个新的动物模型（结扎肠环模型），以培养高素质的人员在快速发展的基因组学，转录组学和生物信息学领域。它将提供独特的答案，并为一个伦理问题带来新的解决方案，这就是使用抗生素养鸡。这个建议代表了我在过去几年中进行的进一步了解坏死性肠炎的研究计划的逻辑连续性。 我的研究计划的目标是更好地了解NE早期感染步骤的致病机制，更准确地说，1）鉴定和表征两种新的Cp细菌素，2）验证三种IV型菌毛蛋白在坏死性肠炎模型中的保护功效，3）使用我们的结扎肠袢模型，使用免疫组织化学，生物标记和细菌转录组学。

Understanding early pathogenesis and virulence of Clostridium perfringens An increasing consumer pressure is forcing animal agriculture to develop alternatives, or at least to substantially reduce the amount of antibiotics used. We must find effective tools to prevent necrotic enteritis, a deadly disease from affecting antibiotic-free chickens. This disease is caused by pathogenic strains of Clostridium perfringens, but predisposing factors are necessary to create an intestinal environment conducive to an increase in the abundance of C. perfringens (Cp). Another part of the disease process is thought to rely on the ability of NE-causing strains to displace non-pathogenic commensal isolates from the gut of chickens. Numerous Gram positive bacteria have been shown to produce various proteinaceious toxic compounds called bacteriocins, and inhibiting the growth of closely related strains. Perfrin has been identified in pathogenic netB positive NE-causing strains and thought to play an important role. When testing various Cp strains on agar spot test, we observed that not all netB positive Cp strains carry the perfrin gene, that bacteriocins other than perfrin were produced, and that some commensal Cp strains can inhibit the growth of pathogenic Cp strains. We have developed an in vivo ligated intestinal loop model reproducing necrotic enteritis in chickens and noted that lesion severity was correlated with numbers of Cp rods lining the intestinal villi (Parent et al, 2016). Thus, Cp attachment to the enterocyte might play a role in the pathogenesis of NE hence our interest in type IV pilins which are known to play a role in adherence to cell surface. We have recently showed that these pilins are also immunogenic when injected in birds. All of these recent findings, have led to the development of this suggested research program. The proposed work is innovative, using a unique Cp strains bank with access to a new animal model (ligated intestinal loop model) to train highly qualified personal in the rapidly developing field of genomics, transcriptomics and bioinformatics. It will provide unique answers and bring new solutions to an ethical problem, which is the use of antibiotics to grow chickens. This proposal represents a logical continuity of the research program on further understanding necrotic enteritis I have been conducting in the past years. The objectives of my research program are to better understand the pathogenic mechanisms underlying the early infection steps of NE, more precisely 1) identify and characterize two new Cp bacteriocins, 2) verify the protective efficacy of three type IV pilins in a necrotic enteritis model, 3) use our ligated intestinal loop model to identify genes involved in early pathogenesis and their effects on the host, using immunohistochemistry, biological markers and bacterial transcriptomics.