Study On Behavioral And Physiological Mechanism Of Individual Growth Differences Of Apostichopus Japonicus

Apostichopus japonicus is one of the most important mariculture species.In recent years,the cultivation of A.japonicus has developed vigorously in China and has become the pillar industry of mariculture.There was a great growth difference in the culture of A.japonicus,which seriously hindered the improvement of the cultivation efficiency of A.japonicus.In order to achieve efficient culture,the understanding of growth differences needs to be improved.However,what are the differences in behavioral,physiological and molecular regulatory characteristics of A.japonicus at different growth rates?What are the key factors affecting individual growth differences?Aiming at the questions above,this study systematically investigated the behavioral,physiological and molecular regulatory mechanisms of individual growth difference of A.japonicus by using time-lapse photography technology,behavioral quantification software,16S r DNA sequencing technology transcriptomic technology,and metabolomics technology.The effects of personality factors and social interaction factors on individual growth difference of A.japonicus were also investigated.The main study results are as follows:1.Comparison of behavior and physiology of A.japonicus with different growth ratesThe difference in feeding behavior was an important cause of individual growth differences in A.japonicus.Using the same family of A.japonicus（weighed 19.18±0.42g）.A.japonicus showed significant individual growth differences during the 30-day experimental period,with the coefficient of variation of body weight increases rapidly from 2.16%to 27.67%.The specific growth rates were 2.41±0.89%d^-1 and1.10±0.66%d^-1 respectively.Using time-lapse photography and behavioral analysis software,we quantified the locomotor and feeding behavior of A.japonicus and found that there were no significant differences in the locomotor behavior characteristics（total distance and movement time）of A.japonicus with different growth rates.However,the feeding time（133.5±150.91 min）and fecal discharge（0.53±0.44 g）of slow-growing A.japonicus were significantly lower than those of fast-growing individuals（feeding time:283±168.98 min;fecal discharge:1.22±0.66 g）,indicating that its feeding behavior was significantly reduced,which might be related to their lower growth rate.There were significant differences in physiological structure,energy metabolism,digestive capacity,and intestinal microorganisms among different growth rates of A.japonicus.At the level of the physiological structure,fast-growing A.japonicus had a lower body wall percentage,which meant that it had a higher percentage of body luminal fluid and intestinal tract,which facilitated the intake,absorption,and transformation of food,and thus promoted growth.At the level of energy metabolism and digestive capacity,the resting metabolic rate(21.60±4.33μg O₂g^-1h^-1)and trypsin activity（65.62±12.42 U/mg）of slow-growing A.japonicus were significantly lower,indicating a significant decrease in its energy metabolism level and digestive capacity,which may be a physiological adaptation to the reduced feeding behavior.In addition,analysis of gut microorganisms in A.japonicus with different growth rates using 16S r DNA sequencing revealed a decrease in gut microbial diversity and an increase in the abundance of potentially pathogenic bacteria（Vibrionaceae and Burkholderiaceae）in slow-growing A.japonicus,while the relatively high abundance of bacteria in fast-growing A.japonicus（Rhodobacteraceae,Rubritaleaceae,and Pirellulaceae）were mostly associated with the degradation of algal polysaccharides.Decreased energy metabolism and digestibility as well as imbalance of gut microbes may be associated with lower growth rates.2.Molecular regulation characteristics of A.japonicus with different growth ratesAnalysis of differentially expressed genes and differential metabolites in A.japonicus with different growth rates revealed that slow-growing A.japonicus was inhibited in growth and suffered from oxidative stress.RNA sequencing and Ultra-Performance Liquid Chromatography-Mass Spectrometry（UPLC-MS）were used to identify and correlate differentially expressed genes and differential metabolites in A.japonicus with different growth rates.The RNA sequencing results showed that 216and 169 significantly up-regulated and significantly down-regulated genes were identified in slow-growing individuals compared to fast-growing individuals,respectively.KEGG enrichment analysis of all differentially expressed genes revealed that the enriched pathways included"ribosome genesis in eukaryotes","spliceosome","apoptosis"and"Hippo signaling pathway",etc.The results of differential metabolite identification showed that 29 metabolites were significantly up-regulated,and 28metabolites were significantly down-regulated in slow-growing A.japonicus compared with fast-growing A.japonicus in positive ionization mode,and 8 metabolites were significantly up-regulated,and 17 metabolites were significantly down-regulated in slow-growing A.japonicus in negative ionization mode.The KEGG enrichment analysis revealed that the key metabolic pathways were"taurine and subtaurine metabolism","arginine biosynthesis","ascorbic acid and alanuric acid metabolism"and"oxidative phosphorylation"."and"oxidative phosphorylation",etc.The correlation analysis of transcriptome and metabolome showed that collagen-encoding genes were significantly down-regulated,some negative growth regulators and apoptosis-inducing genes were significantly up-regulated,and gluconeogenesis and oxidative stress metabolic pathways were intentified in slow-growing A.japonicus,indicating that its growth was severely inhibited and it suffered from severe oxidative stress and needed to consume its own energy to maintain the normal functioning of the organism.3.Effects of personality factors on individual growth differences of A.japonicusA.japonicus possessed personality（consistency of behavioral differences）,and some personality behavior characteristics（movement distance）and resting metabolic rate had good repeatability.Differences in feeding behavior and energy conversion between individuals may be important reasons for differences in growth.Through the isolation culture experiment,it was found that there were great differences in food intake,food conversion rate and specific growth rate among individuals,with the differences being 3.47,6.42 and 5.64 times.The growth rate was positively correlated with food intake and food conversion.Through repeated measurement of resting metabolic rate and personality behavior characteristics,it was found that the resting metabolic rate has good repeatability,while only the movement distance has good repeatability in personality behavior characteristics,and the movement time ratio,latent time,feeding time all have no repeatability,which may be related to the adaptation to the test environment.In addition,the growth rate of A.japonicus was positively correlated with energy metabolism and negatively correlated with stress response.However,the growth rate,resting metabolic rate and cortisol of A.japonicus were not correlated with their personality behavior characteristics.4.Effects of social interaction factors on individual growth differences of A.japonicusThe main reason affecting individual growth differences of A.japonicus was competition for food and adopting appropriate food feeding methods could effectively reduce individual growth differences.Under the condition of sufficient and evenly distributed food,there was no significant difference in the growth rate between isolated and group-farmed individuals.In addition,different from the results of previous studies,the group-farmed A.japonicus in this study did not show greater growth differences.By comparing the behavioral indicators and energy metabolism indicators of isolated cultured and group cultured A.japonicus,it was found that their locomotor behavior,feeding behavior and energy metabolism were not significantly different.Analysis of the gut microbes of isolated and group-cultured A.japonicus using 16S r DNA sequencing revealed that their gut microbial composition was largely similar at the phylum and family levels,and bacteria associated with algal polysaccharide degradation were found in both,but an increased abundance of potentially pathogenic bacteria（Shewanella）was found in isolated cultured A.japonicus,suggesting that social contact and interaction may have an effect on maintaining gut microbial homeostasis and health.In conclusion,this study systematically investigated the behavioral,physiological and molecular regulatory mechanisms of individual growth differences in A.japonicus.The relationship between personality factors and the growth and physiology of A.japonicus was clarified.The key factor affecting the individual growth difference of A.japonicus cultured in group was identified.And a theoretical model was proposed to explain the individual growth differences of A.japonicus in the culture environment.The results provide new insights into the individual growth differences of A.japonicus,and provide a theoretical basis for improving the efficiency of A.japonicus culture,which is conducive to the sustainable and healthy development of A.japonicus culture industry.