Towards lice-resistant salmon: functional genetics and genome editing to enhance disease resistance in aquaculture

养殖三文鱼是人类健康所必需的优质蛋白质和脂肪酸的主要来源。三文鱼水产养殖业对英国经济的价值约为10亿英镑，并支持许多农村和沿海社区。然而，海虱是全球鲑鱼养殖业的一个主要长期问题。这些寄生虫附着在鲑鱼的皮肤上，以组织、粘液和血液为食。受感染的鱼显示生长受损，继发感染的发生率增加。它们对鲑鱼的健康和福利造成了重大的负面影响，而虱子的预防和治疗成本是鲑鱼养殖业的一大经济负担，每年超过8亿GB。令人鼓舞的是，在鲑鱼物种内部和不同物种之间，对海虱的抵抗力都有很大的遗传差异。虽然通常养殖的大西洋鲑鱼通常容易受到感染，但其他鲑鱼物种，如Coho鲑鱼，则完全具有抵抗力。提高养殖鲑鱼对海虱的先天遗传抵抗力是一种环境友好但未得到充分利用的控制虱子的方法。通过对大西洋鲑鱼的选择性繁殖，已经实现了渐进式的改良，但其漫长的世代间隔减缓了进展。基因组编辑提高了通过对鲑鱼基因组进行精确的有针对性的改变来迅速增加鲑鱼抵抗力的可能性；关键是知道哪些特定的基因是目标。这个项目的重点是了解海虱抗性的遗传机制，并确定基因组编辑的基因目标，以培育抗虱子的大西洋鲑鱼。为了确定目标抗虱基因进行编辑，将采取几种不同的方法，每一种方法都利用最新的基因组技术。首先，将从大量养殖三文鱼中获得全基因组序列，这些三文鱼在挑战后收集了海虱数量。这些基因将被用来定位导致商业大西洋鲑鱼种群抗药性变异的个别基因。其次，已知抵抗海虱的潜在机制是由于成功地在靠近虱子附着部位的局部免疫反应。因此，将使用单细胞测序方法对大西洋鲑鱼和科鲑鱼在虱子攻击后头四天的免疫反应进行详细的基因表达比较，以突出不同细胞群体在虱子附着位置的不同反应。此外，还将对虱子的基因表达进行分析，并确定潜在的免疫调节蛋白及其在宿主中的靶标。第三，基因组编辑方法将被用来评估扰动候选抗性基因对细胞培养和鱼类本身对海虱反应的影响。前者将被用来评估细胞对从海虱分泌的蛋白质的反应，以及敲除每个目标基因对这种反应的后果。这将导致在鲑鱼胚胎中编辑一组最终的目标基因，之后编辑后的鱼将受到海虱的挑战。然后将评估编辑后的鱼与完全同胞控制的鱼相比的抵抗力。该项目的科学计划将通过与工业合作伙伴Benchmark PLC共同制定繁殖和传播编辑后的抗虱子三文鱼的战略来补充。此外，计划举办公众和利益攸关方参与活动，以宣传研究计划和结果，特别侧重于水产养殖中基因组编辑的好处和风险。抗虱子鲑鱼的成功结果将通过预防疫情和消除化学处理的需要，对动物福利和经济产生重大影响。它还将提供一个备受瞩目的例子，说明基因组编辑技术在理解生物学和改善食品安全和动物健康方面的力量。

Farmed salmon is a major source of high quality protein and fatty acids essential for human health. Salmon aquaculture is worth approximately £1Bn to the UK economy, and supports many rural and coastal communities. However, sea lice are a major perennial problem for salmon aquaculture worldwide. These parasites attach to the skin of salmon and feed on tissue, mucus and blood. Infected fish show impaired growth and increased occurrence of secondary infections. They cause significant negative impacts on salmonid health and welfare, while lice prevention and treatment costs are a large economic burden for salmon farming, over £800M per annum. Encouragingly there is substantial genetic variation in resistance to sea lice both within and across salmonid species. While the commonly farmed Atlantic salmon are generally susceptible to infection, other salmonid species such as coho salmon are fully resistant. Improving the innate genetic resistance of the farmed salmon to sea lice is an environmentally friendly, but underexploited approach to lice control. Incremental improvements have been achieved via selective breeding of Atlantic salmon, but their long generation interval slows progress. Genome editing raises the possibility of rapidly increasing the resistance of salmon via precise targeted changes to their genomes; the key is knowing which specific genes to target. This project focusses on understanding the genetic mechanisms underlying resistance to sea lice, and identifying gene targets for genome editing to develop lice-resistant Atlantic salmon.To identify target lice resistance genes for editing, several different approaches will be taken, each exploiting the latest genomic technologies. Firstly, whole genome sequences will be obtained from a large population of farmed salmon on which sea lice counts following challenge have been collected. These will be used to map individual genes that contribute to variation in resistance in the commercial Atlantic salmon population. Secondly, it is known that the mechanisms underlying resistance to sea lice are due to a successful localised immune response close to the attachment site of the louse. Therefore, a detailed gene expression comparison of the immune response of Atlantic and coho salmon in the first four days following a lice challenge will be undertaken, using single cell sequencing approaches to highlight different responses in distinct cell populations at louse attachment sites. This will be complemented by profiling of the gene expression of the lice, and identification of potential immunomodulatory proteins and their targets in the host. Thirdly, genome editing approaches will be used to assess the impact of perturbing candidate resistance genes on response to sea lice both in cell culture and in the fish themselves. The former will be used to assess the cellular response to proteins secreted from the sea lice, and the consequences of knocking out each of the target genes on that response. This will lead to a final set of target genes for editing in salmon embryos, after which the edited fish will be challenged with sea lice. The resistance of the edited fish compared to full sibling control fish will then be assessed.The scientific programme of the project will be complemented by co-development of a strategy for the breeding and dissemination of edited lice resistant salmon, together with industrial partner Benchmark PLC. Furthermore, public and stakeholder engagement events are planned to communicate the research plans and outcomes, with a particular focus on the benefits and risks of genome editing in aquaculture. A successful outcome of lice resistant salmon would have major animal welfare and economic impacts via prevention of outbreaks, and removal of the need for chemical treatments. It would also provide a high profile example of the power of genome editing technology to understand biology and to improve food security and animal health.