Effect of co-infections on the pathogenesis of the most important swine viruses

目前正在对猪病毒进行调查，但主要是在单一感染的情况下进行。不幸的是，这种类型的调查并不能代表现实生活中的情况。事实上，几种病原体在一只动物中混合在一起，极大地降低了针对病原体的治疗效率，并使实施预防措施变得更加困难。因此，对混合感染的调查是了解结果和如何治疗传染病的合乎逻辑的步骤。呼吸道中多种感染性病原体的联合构成了已知的猪呼吸道疾病复合体。该复合体包括多种病原体，如猪圆环病毒2型(PCV2)、猪繁殖与呼吸综合征(PRRSV)病毒(PRRSV)和猪流感病毒(SI)，它们分别是断奶后多系统衰竭综合征(PMWS)、PRRS和猪流感传染病的病原体。此外，霉菌毒素脱氧雪腐镰刀菌烯醇(DON)是谷物的主要污染物，猪因饮食原因长期暴露于DON。猪的免疫系统对DON特别敏感。从全球养猪业的经济损失来看，最重要的动物疾病是PRRS(加拿大每年1.5亿美元)。在2005年PMWS暴发期间，魁北克每年有20万头猪死亡。另一种主要疾病是猪流感。该病在猪群中高度流行，在养猪业中造成约2.5%的死亡率。因此，本研究计划的主要目标是进一步确定猪合并感染的发病机制，并在细胞水平上更好地了解为什么在某些情况下存在拮抗作用(这可能导致我们发现新的抗病毒药物)，而在其他情况下，在合并感染过程中猪病原体之间存在协同作用。为了帮助我们研究猪的联合感染，最近对新的病毒允许细胞系进行了基因改造，使其对多种猪病毒具有容许性。因此，本五年研究计划的主要目标是研究PRRSV、PCV2和SI在不同组合中的联合感染以及DON/PRRSV联合感染的发病机制，以更好地了解参与病毒促进和/或抑制的宿主反应。在联合感染实验中，将研究几个参数，例如：病毒复制动力学、细胞周期/细胞死亡(荧光显微镜、酶分析等)、细胞因子(qPCR、ELISA)和细胞通路调节(RNAseq、CRISPR/Cas9)。综上所述，对猪混合感染的研究将使我们在更现实的野外条件下更好地了解猪病毒的致病机理，从而为猪病毒感染性疾病的控制和治疗带来新的发现，具有很高的应用潜力。

Swine viruses are being investigated but predominantly in a context of single infections. Unfortunately, this type of investigation do not represent real life situation. In reality, several pathogens commingle together in a single animal reducing dramatically the efficiency of pathogens-specific treatments and making it more difficult to implement prophylactic measures. Therefore, the investigation of co-infections is a logical step to understand the outcome and how to treat infectious diseases. Associations of multiple infectious agents in the respiratory tract constitute what is known to be the porcine respiratory diseases complex. Several pathogens are included in that complex such as porcine circovirus type 2 (PCV2), porcine reproductive and respiratory syndrome (PRRS) virus (PRRSV), and swine influenza virus (SI), which are the etiological agents involved in postweaning multisystemic wasting syndrome (PMWS), PRRS, and swine flu infectious diseases, respectively. In addition, deoxynivalenol (DON) mycotoxin, a mold product, is a major contaminant of cereal grains and pigs are exposed chronically to DON because of their diet. Pig immune system is particularly sensitive to DON. The most important animal disease, in term of economical loss in swine industry worldwide, is PRRS (150 million $/year in Canada). During the 2005 PMWS outbreak, 200,000 pigs/years died in Quebec herds. Another major disease is swine flu. It is highly prevalent in swine herds and it causes a mortality of around 2.5% in swine productions. Thus, the main goal of the present research program is to further characterize the mechanisms involved in the pathogenesis of co-infections in swine and to better understand at a cellular level why there is in some instance an antagonism (which could lead us to the discovery of new antiviral drugs) and in other instance a synergy between swine pathogens during co-infections. To help us investigate swine co-infections, new virus permissive cell lines were recently genetically modified to render them permissive to more than one swine virus. Thus, the main objective of the present five years research program is to investigate the pathogenesis of PRRSV, PCV2 and SI co-infections in various combinations and DON/PRRSV co-infection, to better understand the host responses that could be involved in virus promotion and/or inhibition. Several parameters will be studied during co-infection experiments such as: virus replication kinetics, cell cycle/cell death (fluorescent microscopy, enzymatic assays, etc.), cytokines (qPCR, ELISA), and cellular pathways modulation (RNAseq, CRISPR/Cas9). In conclusion, the study of swine co-infections will give us a better understanding of the swine viral pathogenesis in a more realistic field condition and subsequently will lead to new discoveries with high applied potential for the control and treatment of swine viral infectious diseases.