Table3_Differential responses of selectively bred mussels (Perna canaliculus) to heat stress—survival, immunology, gene expression and microbiome diversity.xlsx

New Zealand’s green-lipped mussel (Perna canaliculus) is an ecologically and economically important species. Marine heatwaves are increasing in frequency around NZ’s coastline, and these events are correlated with increased stress and mortality of some aquaculture species. This study aimed to identify general biomarkers of heat stress in P. canaliculus and to assess whether responses differed between genetically distinct selectively bred mussels. We exposed three families of selectively bred mussels (families A, B and C) to three seawater temperature regimes in the laboratory: 1) a “control” treatment (ambient 12°C), 2) a 26°C heat challenge with a subsequent recovery period, and 3) a sustained 26°C heat challenge with no recovery. We investigated whether the survival, immune response (hemocyte concentration and viability, oxidative stress and total antioxidant capacity), hemocyte gene expression and gill microbiome differed between the families during the temperature challenges. In the sustained heat-stress treatment, family A had the highest survival rate (42% compared with 25% and 5% for families C and B, respectively). Gene expression levels significantly shifted during thermal stress and differed between families, with family A more dissimilar than families B and C. Family C had substantially more genes impacted by temperature treatment and timepoint than the other families, while family B had very little genes/pathways that responded to thermal stress. Genes related to heat shock proteins and immune responses (e.g., AIF1, CTSC, TOLL8, CASP9, FNTA, AHCY, CRYAB, PPIF) were upregulated in all families during heat stress. Microbiome species-richness differed between families before and during heat-stress, with family A having a distinctly different microbiome flora than the other families. Microbial diversity changed similarly in all families exposed to prolonged heat-stress, with species of Vibrio and Campylobacter increasing in these mussels. Our study highlights the use of non-lethal sampling of hemocytes as a diagnostic tool to explore the immune response and gene expression of selectively bred mussels, to predict their response to ocean warming. This approach can identify potential thermotolerant candidates for further selective breeding, which may increase the resilience of the mussel aquaculture industry in a warming ocean.

新西兰绿唇贻贝（Perna canaliculus）是一种兼具重要生态与经济价值的物种。近年来新西兰沿岸海域的海洋热浪发生频率持续升高，这类事件与部分水产养殖物种的应激水平上升及死亡率增加密切相关。本研究旨在鉴定绿唇贻贝热应激的通用生物标志物，并评估不同遗传背景的选育贻贝之间的应激响应是否存在差异。我们在实验室中将3个选育贻贝家系（家系A、B、C）置于3组海水温度处理条件下：1）对照组（环境温度12℃）；2）26℃热胁迫及后续恢复期；3）持续26℃热胁迫且无恢复期。我们探究了在温度胁迫过程中，各贻贝家系的存活率、免疫响应（血淋巴细胞浓度与活性、氧化应激水平及总抗氧化能力）、血淋巴细胞基因表达以及鳃部微生物组是否存在差异。在持续热胁迫处理组中，家系A的存活率最高（42%，家系C和B的存活率分别为25%和5%）。热应激过程中基因表达水平发生显著改变，且不同家系间存在差异，其中家系A的基因表达谱与家系B、C的差异更为显著。相较于其他两个家系，家系C受温度处理及采样时间点影响的基因数量显著更多，而家系B中响应热应激的基因/通路数量极少。热应激期间，所有家系中与热休克蛋白及免疫应答相关的基因（如AIF1、CTSC、TOLL8、CASP9、FNTA、AHCY、CRYAB、PPIF）均出现上调表达。热应激前后，不同家系的微生物组物种丰富度存在差异，其中家系A的微生物群落组成与其他家系显著不同。在持续热胁迫处理的所有贻贝家系中，微生物多样性变化趋势一致，其体内弧菌属（Vibrio）与弯曲杆菌属（Campylobacter）的物种丰度均出现上升。本研究证实，通过非致死采样获取的血淋巴细胞可作为诊断工具，用于探究选育贻贝的免疫响应与基因表达特征，进而预测其对海洋变暖的应激响应。该方法可筛选出潜在的耐热候选个体用于后续选育工作，有助于提升变暖海洋环境中贻贝养殖业的抗逆能力。