Study On Molecular Response To Heat Stress And Vibrio Alginolytic Infection In The Immature And Post-spawning Populations Of Pacific Oyster Crassostrea Gigas

Crassostrea gigas,also known as Pacific oyster,belongs to the biological classification of Mollusca,Lanellibranchia,Ostreida,Ostreidae,Crassostrea.It is a wide-temperature and wide-salt mariculture shellfish.As a typical species in this special environment of intertidal zone,it is regarded as a model organism for studying the adaptability of Marine organisms to the intertidal zone environment.C.gigas is one of the most important mariculture species in the world because of its ecological and economic value.But in recent years,mass mortaity of C.gigas are often reported,especially in the summer when sea temperatures rise.To some extent,this has troubled the development of the oyster industry.Preliminary investigation indicates that the phenomenon of mass mortality may be mainly caused by high temperature stress,individual physiological state and pathogenic microorganisms,but there is no overall systematic study.Aiming at this practical problem,this study carried out a systematic environmental and biological investigation on the summer mass mortality of C.gigas,and obtained the gene set of stress response through transcriptome to conduct a preliminary study on its molecular mechanism.The results are as follows:1.In this study,the hydrological conditions(temperature,dissolved oxygen,p H and salinity)of the high-temperature season in the oyster culture area of Rushan city,Shandong province,which is the "hometown of oysters" in China,were examined,The quantity of phytoplankton in the aquaculture water at each time point was counted,the variation of the condition index of oyster at each sampling time points was calculated,and some resistance genes were screened for q PCR of oyster gill tissues in different seasons.The results showed that the HSP70 gene was highly expressed in July,and the immune genes such as PFK,My D88,MPI2,Lys and IL-17 all showed peak expression in August,and the expression level was lower in other months.In addition,in the months when we examined the hydrological data,we found that the highest temperature of the seawater occurred in August,which was consistent with the high expression months we obtained.The analysis of the condition index of oysters shows that this index is closely related to the change of the environment and significantly decreases in the summer months.The statistical results of algae show that the algal abundance is greater near the shore.These results preliminarily revealed the summer culture environment of oyster,and considered that temperature was the main factor of death,which provided effective information for the culture of C.gigas.2.As an internal influence of the organism,spawning plays a role during summer mortality.In order to understand the effects of spawning on the physiological status of oysters,we obtained two populations with different reproductive states(immature and post-spawning oysters)at the same time by method of artificial ripening.We mainly compared the physiological indexes such as glycogen content,condition index and respiration rate of the two populations.The results showed that spawning resulted in a significant decrease in the glycogen content of oysters,and the condition index was also accompanied by a significant decrease in spawning.Glycogen,as an important energy storage material of collective energy,was closely related to the resistance of individuals.For the results of respiration rate,at the same temperature,the respiration rate of post-spawning oysters was significantly higher than that of the immature individuals,which may be a respiratory compensatory mechanism.3.The reproductive activity of C.gigas affects the body’s ability to resist external stress.Among various environmental and biological factors,temperature and pathogen infection are considered to be important factors affecting the survival of oysters.We respectively treated oyster of immature and post-spawning individuals with heat stress treatment at 35℃ and injection treatment of vibrio alginolyticus.After heat stress and vibrio infection,a certain time point was selected to collected individual gill tissues for transcriptome sequencing.Calculating the different gene sets of the population under different reproductive states after stress,furthermore,enrichment analysis was carried out on the genes with significant differences in two populations to systematically understand the molecular mechanism of population response to stress.The results showed that: through the statistics of different genes,it was found that the response degree of oysters in different reproductive states was significantly different when exposed to heat shock,and the population of post-spawning was significantly more susceptible to heat stress.The response degree of the two groups was relatively large after infected with vibrio.GO enrichment analysis found that there was no enrichment to biological function difference in the immature population after heat stress,while the metabolic process was accelerated and apoptosis-related genes were up-regulated in the post-spawning population.Instead,the immune capacity of the population was down-regulated.Compared with the heat stress treatment,vibrio injection had a greater effect on two populations.Among them,the metabolic process of immature population was generally up-regulated,while the immune function was inhibited.The response of post-spawning oysters to stress was weakened and the self-healing process was affected.In summary,mass mortality of C.gigas caused by the interaction of multiple factors.The results of this study showed that the spawning process led to the physiological state of C.gigas was poor,which greatly affected the body’s resistance to temperature and pathogen stress,leading to the occurrence of mass mortality.