| Chlamys farreri is one of the important commercial shellfish, which naturallydistributes along the coasts of China, Korea and Japan. Recently, the scallop industrysuffered from enormous pecuniary loss caused by the mass summer mortality of C.farreri. Till now, lots of studies have been performed from the environmental ecology,etiology, shellfish immunology, germplasm genetics points of view. However, therestill lacks an effective solution to this mortality issue. In this study, based on theinvestigation of stress resistance related physiology indexes during C. farreri aging,tissue-specific assessment indexes for energy metabolism and antioxidant ability wereestablished. The effects of reproduction and heat stress on scallop individuals wereanalyzed using the assessment indexes in combination with the expression ofregulatory genes. The biochemical indexes for scallop energy metabolism andantioxidant ability will potentially benefit the genetic breeding for sustainabledevelopment of scallop aquaculture, providing useful information for unraveling thereason for summer mortality.1. Construction of assessment indexes for stress resistanceWe analyzed the homeostasis maintenance-related mechanism including thetelomere regulation, DNA methylation, energy metabolism and antioxidant ability, inC. farreri aged1to4years. The telomere investigation results showed that there wasa strong correlation among telomere length, telomerase activity andtelomerase-related gene expression. The4-year old scallops exhibited significantlyshorter telomere as well as lower TERT expression compared with3-year old ones inhepatopancreas. Global methylation analysis showed that the DNA methylation leveldecreased significantly with age in mantle, kidney and hepatopancreas P<0.05. Wealso found that the DNMT1expression between1-year old and4-year old scallops was significantly different in mantle and hepatopancreas P<0.05. Correlationanalysis revealed that the eight biochemical indexes SOD, CAT, GOT, GPT, LDH,ROS, MDA and ATP involved in energy metabolism and antioxidant ability werehighly correlated with age P<0.05. Because of the obvious variation with age aswell as the wide existence in various tissues, we chose the energy metabolism andantioxidant ability related indexes for the subsequent analysis. Based on principalcomponent analysis, the tissue-specific composite assessment index for energymetabolism and antioxidant ability can be calculated. For example, Nkidney=0.30*LDH+0.29*ROS+0.29*ATP+0.272*MDA+0.25*GPT+0.21*CAT+0.18*SOD+0.07*GOT. Correlation analysis revealed that the tissue-specific compositeassessment index for stress resistance had negative correlation with age and positivestudy showed that kidney had the highest correlation with age r=0.82,p-value=9.22e-11.2. Effects of reproduction and heat stress on energy metabolism andantioxidant abilityBy using the composite assessment indexes for stress resistance, the energymetabolism and antioxidant ability of C. Farreri after reproduction and heat stress30°C were estimated. Results showed that the basal biochemical indexes declinedsignificantly in most tissues after reproduction P<0.05. For example, theantioxidant enzyme activity declined90%and the ATP level declined80%in gonadafter reproduction. Besides, the composite indexes of mantle, gill, gonad, kidney,adductor muscle and hepatopancreas were all significantly lower after reproductionP<0.05. Survival analysis under30°C heat stress revealed that the survival rate of1-year old group was significant higher compared with2-4year old groups P<0.05.Statistical analysis showed that the survival rate was89.69%in age1group,37.72%in age2group, and less than15%in3-4year old groups after12.5h heat exposure.The basal biochemical indexes also declined with age under heat stress, and thecomposite index was significantly lower in age1than the other age groups P<0.05. Compared with the control group under normal condition, the scallops aged2to4years had a significantly lower composite index after heat stimulation. Unexpectedly,the composite indexes in kidney and hepatopancreas of age1scallops wassignificantly elevated P<0.05after heat stress. Our results showed that thecomposite index declined after reproduction and heat stress, suggesting a negativeeffect of reproduction and heat stress on energy metabolism as well as antioxidantability in C. farreri.3. Expression analysis of growth, metabolism, aging and reproduction-related signaling pathway genesExpression analysis of PI3K-AKT-TOR IGF-1r-AMPK-FOXO and P53-Sestrinsignaling pathway genes that involved in the regulation of Mn-SOD CAT GPT etc.,showed that expression of genes that positively regulate aging increased with ageunder both normal and heat-stress condition. The elevation was significant P<0.05in some tissues for scallops after reproduction and heat stress. In kidney, theexpression of PI3K, IGF-1r,, FOXO, P53was significantly different amongage groups P<0.05. In gonad, the expression of PI3K, AKT, IGF-1r, wassignificantly increased after reproduction P<0.05, and the expression of TSC2andFOXO was significantly reduced P<0.05. Significant changes in the expression ofthese signaling pathway genes after heat stress was found in gonad, kidney andhepatopancreas. Up-regulation of genes that positively regulate aging suggested thatreproduction and heat stress may accelerate the aging rate of C. farreri, which wasconsistent with the trend reflected by the composite index. |
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