密度胁迫对珍珠龙胆石斑鱼生长和生理的影响

Effects of density stress on growth and physiology of Epinephelus fuscoguttatus ♀×Epinephelus lanceolatus ♂

关键词:  珍珠龙胆石斑鱼  养殖密度  氧化应激  消化代谢  HPI轴

Key words:  Epinephelus fuscoguttatus♀×Epinephelus lanceolatus♂  Stocking density  Oxidative stress  Digestion and metabolism  HPI axis

摘要:

养殖密度是影响工厂化循环水养殖的重要因素之一，为探究密度胁迫对珍珠龙胆石斑鱼(Epinephelus fuscoguttatus ♀×Epinephelus lanceolatus ♂)幼鱼生长、消化代谢酶活性、抗氧化酶活性和下丘脑-垂体-肾间组织(HPI)轴相关激素水平及基因表达的影响，实验选取1 800尾规格一致、体格健康的珍珠龙胆石斑鱼，随机分为3个密度梯度：低密度100尾[(3.14±0.13) kg/m3, LD]、中密度200尾[(6.31±0.13) kg/m3, MD]和高密度300尾[(9.56±0.24) kg/m3, HD]进行实验。研究表明，HD组特定增长率、肥满度及存活率显著低于其他各组(P<0.05)；饵料系数和变异系数随密度的升高显著上升(P<0.05)；HD组抗氧化酶和代谢酶活性显著高于其他各组(P<0.05)，而消化酶活性显著低于其他各组(P<0.05)；HD组HPI轴相关激素水平显著高于其他各组(P<0.05)，HPI轴相关基因(crhr1、nr3c1及nr3c2)的表达量随密度的升高显著上调(P<0.05)，crh-bp表达量则相反(P<0.05)。本研究结果可为深入认识密度胁迫对珍珠龙胆石斑鱼生长、消化代谢、氧化应激及内分泌的影响提供科学依据，为实际生产过程中珍珠龙胆石斑鱼幼鱼养殖密度的设定提供理论参考。

Abstract:

The pearl gentian grouper (Epinephelus fuscoguttatus ♀×Epinephelus lanceolatus ♂) is the main cultured species of marine fish in factory recirculating aquaculture system (RAS). High density presents the main characteristic of intensive farming model represented by RAS, and density stress is a fundamental factors affecting fish welfare. Density stress causes a series of changes in fish growth performance, digestive and metabolic capacity, oxidative stress states, and endocrine homeostasis. However, research on how the pearl gentian grouper copes with density stress, and the effects of density stress on the growth, digestion, metabolism, oxidative stress, and endocrine production requires further research. Seeking a suitable stocking density can improve the culture efficiency of the pearl gentian grouper and avoid culture risks. Selecting an appropriate stocking density is an important problem to be solved in the factory recirculating aquaculture model. Therefore, this study investigated the changes of growth performance, digestive and metabolic capacity, oxidative stress, and Hypothalamus - Pituitary - Interrenal (HPI) axis related parameters of pearl gentian grouper under different density conditions. Three density gradient groups were set: low-density (LD), medium-density (MD), and high-density (HD). In the LD, MD, and HD groups, 100, 200, and 300 pearl gentian groupers were added per barrel with a density of (3.14±0.13), (6.31±0.13), and (9.56±0.24) kg/m3, respectively. The three experiments were performed in parallel for 60 days. Fish were fed a formula feed of 2.5% of their body weight in the morning and evening (08:00, 17:00) daily. The density stress test was performed for 60 days in a factory recirculating aquaculture system [temperature (27±2) ℃, dissolved oxygen (8.0±1.0) mg/L, pH 7.8 and salinity 24±1]. The body weight, body length, and total length of all the fish were measured every 20 days. A total of 18 juvenile pearl gentian groupers were randomly collected from each density group, for a total of 54. Anesthesia was administered (MS-222, 80 mg/L), tail vein blood was drawn with a 2 mL disposable syringe, and centrifuged (4 000 r/min, 4 ℃, 10 min) to obtain a supernatant that was stored at –20 ℃ for testing. Before testing the corresponding indicators, the brain, liver, intestine, stomach, and kidney were frozen in liquid nitrogen. The results showed that the optimum stocking density of pearl gentian grouper was (15.48–29.67) kg/m3 under the conditions of water [temperature (27±2) ℃, dissolved oxygen (8.0±1.0) mg/L, pH 7.8, and salinity 24±1]. In terms of growth, the specific growth rate and condition factor in the HD group were significantly lower than those in the other groups (P<0.05), and the feed coefficient and coefficient of variation were significantly increased with the increase of density (P<0.05). In addition, the survival rate of the HD group was significantly lower than that of the other groups (P<0.05). Density stress results indicated that the growth retarded, weight difference increased, and mortality increased with increasing density. In terms of digestion and metabolism, the digestive enzyme activities (pepsin, trypsin, lipase, and amylase) in the HD group were significantly lower than those in the other groups (P<0.05). Density stress results in a significant decrease in the digestive performance of the fish. The metabolic enzyme activities (PK, SDH, HK and LDH) in the HD group were significantly higher than those in the other groups (P<0.05). In terms of oxidative stress, the activities of antioxidant enzymes (MDA, SOD, CAT and GSH-Px) in the HD group were significantly higher than those in other groups (P<0.05). In terms of HPI axis, the levels of HPI axis related hormones (CRH, ACTH and CORT) in the HD group were significantly higher than those in other groups (P<0.05). qRT-PCR was used to determine the expression levels of HPI axis-related genes, and the density stress upregulated crhr 1, nr3c 1, and nr3c 2 (P<0.05), while crh-bp was downregulated (P<0.05). In summary, the study revealed that the effects of density stress on growth performance, digestive and metabolic capacity, stress, and HPI axis of the pearl gentian grouper. Density was negatively correlated with the growth performance and caused a significant decrease in digestive enzyme activity, significantly increased metabolic enzyme activity, intensified oxidative stress, and dysregulated HPI axis hormones and related genes. The results of this experiment can provide theoretical reference for the establishment of stocking density for juvenile pearl gentian grouper in the production process. Our findings provide scientific evidence for further understanding the effects of density stress on the growth, digestive metabolism, oxidative stress, and endocrine production of the pearl gentian grouper.