文蛤弧菌耐受性状的量化及其遗传解析

Genetic variation for anti-disease trait has been widely reported in various aquaculture species, and the moderate heritability of anti-disease trait was estimated in clam Meretrix petechialis. It indicated that there is a large potential for genetic improvement in anti-disease trait. However, there are a few reports about systematic breeding for improvement of anti-disease traits compared with the selection for growth traits in animal breeding. Main reason is that the study of anti-disease trait and its incorporation into breeding programs have been hindered by the barrier to determining and measuring accurate and appropriate phenotypes. Up to now, traits like anti-disease ability have been measured directly using challenge trials, dead or survivor status were used as phenotypic scores. The dead or survivor phenotype reflect both resistance and tolerance trait as the total overall ability to maintain fitness or life in the face of disease infection in host. The present study in aquaculture breeding was seldom designed to distinguish the difference in disease resistance and tolerance. Disease resistance (the host’s ability to limit parasite burden) and tolerance (the host’s ability to limit the damage caused by a given parasite burden) actually represent different aspects of disease resistance and defense. Resistance is typically measured as the inverse of parasite burden, and tolerance is usually defined as the slope of host fitness against infection intensity. So, one aim of our project is to distinguish and measure resistance and tolerance trait by establishment the equation of pathogen load and survival status in the clam Meretrix petechialis. .Tolerance trait is still expensive and difficult to measure as above mentioned, so marker-assisted selection (MAS) or indirect selection on the selection candidate is necessary for tolerance selection in clam breeding programs. Analyses of gene expression profiles as an indirect test for disease resistance have been proposed as alternative technologies in selective breeding. Another aim of our project is to discover likely indicator gene expression data affecting tolerance trait and estimate theirs genetic parameters the by using statistical models. In a word, the project is valuable for better understanding of disease resistance and tolerance and provided the basis for optimal breeding strategies in genetic improvement of clam M. petechialis.

抗病性状具有广泛的遗传变异基础，但以提高抗病力为目标的育种与其他如针对体长、体重等生长性状的选育相比相对滞后，造成这种状况的一个重要原因是对抗病力性状的界定不清以及抗病性状测定评价相对困难。在本项目中我们将通过建立病原菌载量-存活率关系曲线，实现文蛤弧菌抗性和耐受性的区分和评价，使抗病选育的目标性状更为清晰和明确；通过耐受机制相关通路基因表达数据和耐受表型数据的相关分析，筛选与耐受性密切相关的基因mRNA标记性状，估计该标记性状的遗传参数，对弧菌耐受性进行间接选择，提高抗病育种的准确性和预见性。本研究立足文蛤抗弧菌病这一重要经济性状，通过明确抗病类型、筛选耐受性相关标记性状，评估遗传参数，预期研究结果可以加深对贝类抗弧菌病这一复杂性状的理解，同时对于以文蛤为代表的滩涂贝类抗病选育方案的制定和育种实践具有重要的应用前景。

在建立了稳定的弧菌人工感染体系的基础上，确定了能够使不同家系死亡率呈现出最佳分辨率的人工感染条件，明确了感染时间和文蛤肝胰腺弧菌载量之间的关系曲线。由此我们提出了通过分析第1天到第3天天肝胰腺组织载菌量的斜率变化来评价家系相对抗性（清除病原菌的能力）的方法。通过测定同一家系在不同的感染强度下，载菌量和家系死亡/存活率对应数据，建立病原菌载量存活率关系曲线，对家系进行耐受性的评价。通过以上的工作我们实现了对抗性和耐受性两种性质迥异甚至相互拮抗的性状的区分和评价，建立了除死亡率之外的文蛤抗性评价指标。在此基础上我们进一步分析了文蛤不同发病阶段的肝胰腺转录水平对感染的动态响应，发现代谢和免疫系统在感染过程中相互协调，代谢过程通过调节代谢资源的分配，支持文蛤在感染过程中的防御和生存，表明代谢系统可以作为抗性和耐量性状之间的纽带和桥梁。在对文蛤抗病性状进行分解的基础上，开发了一批与抗性和耐受性密切相关的基因mRNA标记。进一步对这些基因表达性状的遗传参数和遗传力进行了估计，发现这些基因表达性状的遗传力分布在 0.113-0.530之间。在选育过程中通过标记育种值对亲本进行评价和选择构建了 F2 代家系，弧菌感染实验表明，F2 代的弧菌抗性显著提高，感染后的存活率较 F1 代提高了10%-15%，获得了良好的抗性遗传进展。证实了利用标记进行选择的可行性。培育的“科浙2号”抗弧菌新品种通过了农业部的良种审定（GS-02-001-2021）。以上研究结果帮助我们加深了对文蛤抗弧菌病这一复杂性状的理解，同时开发了大量抗性、耐受性相关的标记，对于以文蛤为代表的滩涂贝类抗病选育实践具有重要的应用前景。