Effects of fasting on metabolic and immunological responses of common carp (Cyprinus carpio) to a further acute stress

Common carp were fasted for 0, 3 and 14 days prior to exposure to a 2‐hr crowding stress. Serum cortisol, glucose, lactate, _{T4, T3}, serum alternative complement (ACH50), lysozyme, total immunoglobulin (Ig) and blood WBC were determined before and 0, 6 and 24 hr after the crowding stress. Three days of fasting led to a significant decrease in serum levels of lactate and thyroid hormones. Fourteen days of fasting led to initiation of physiological process to maintain serum glucose by increase in serum cortisol level and lactate utilization. These fish had lower levels of thyroid hormones suggesting suppression of basal metabolism. The fish also had lower WBC and serum lysozyme and total Ig levels indicating immunosuppression. After the crowding stress, the fish fasted for 3 days had significantly lower lactate, thyroid hormones and total Ig levels. The fish fasted for 14 days had higher cortisol but lower glucose and thyroid hormones levels suggesting impairment in metabolic responses to the crowding stress. Immunological and health indicators including WBC, and ACH50, lysozyme and total Ig of these fish were significantly lower than the other groups after the crowding stress. Overall, the results show that a 14‐day fasting leads to stress responses, reduction in metabolism and immunosuppression. Moreover, 14 days of fasting lead to greater cortisol response to a further crowding stress, which induces marked immunosuppression and fallen welfare.     

Individual variations in behavior and free cortisol responses to acute stress in tiger pufferfish Takifugu rubripes

Abstract:   The aim of the present study was for individual variation in behavior to be characterized and related to differences in physiological stress responses in juvenile tiger pufferfish (fugu) Takifugu rubripes . A set of siblings ( n  = 330) from wild parents was subjected to behavior tests, and some were classified into active and inactive groups ( n  = 42 each group). The active animals consisted of those individuals that kept active swimming during 6 min after transfer from one tank to another, and the inactive animals were those that stopped swimming within 6 min. Time-course acute stress responses, which were elicited by anesthesia and subsequent removal of a pectoral fin, were compared between the active and inactive fish. The stress indicators used were free and total plasma cortisol, and plasma glucose. Although no significant difference was observed at each sampling point between groups, the inactive fish required shorter recovery time from acute stress in all three indicators than the active fish. The percentage of free to total plasma cortisol in the pre-stress condition was significantly lower in the inactive group than in the active group. Our findings indicate that fugu has behavioral variation that is associated with stress responsiveness. The inactive fish are expected to be more tolerant of stress and suitable for aquaculture.     

The physiological response to capture and handling stress in the Atlantic sharpnose shark, Rhizoprionodon terraenovae

Secondary effects of capture and handling stress in the Atlantic sharpnose shark, Rhizoprionodon terraenovae, were investigated in this study. Twenty-four sharks were serially bled and changes in several hematological parameters were monitored over a 1-h time period, following capture by hook-and-line. Blood samples were obtained from each shark at 0, 15, 30, 45 and 60 min. All 0-min (initial) blood samples were obtained within 3 min of capture. Blood glucose (9.2–13.1 mmol l^–1), lactate (1.5–28.9 mmol l^–1), and plasma osmolality (871–929 mOsm kg^–1) all increased after capture, whereas blood pH (6.86–6.78) declined. Hematocrit values (initial = 25.1%) remained unchanged throughout the 1-h stress period. Due to the short amount of time it took to obtain the initial sample and the lack of a significant relationship between the initial time and the initial parameter levels, all initial samples are considered the best approximation of the predisturbance resting levels. The use of repeated measures in this study enables us to describe the dynamics of the secondary stress response in the Atlantic sharpnose shark.     

计算机视觉技术在鱼类应激状态监测中应用研究

从行为参数的量化方法、应激状态下的行为变化两方面介绍了计算机视觉技术在鱼类应激状态监测方面的研究进展,并提出了进一步研究的方向是实际应用过程中量化鱼行为的新方法研究,以及根据鱼在不同条件下行为参数的变化特点,结合统计方法和人工智能算法建立自动判断应激的方法。     

Consistency of stress response to repeated handling in the gilthead sea bream Sparus aurata Linnaeus, 1758

Gilthead sea bream Sparus aurata Linnaeus, 1758 were subjected to repeated stresses, namely confinement and handling once a month for 5 consecutive months. Plasma cortisol concentration was determined as a primary stress response, and plasma glucose and osmolality, serum agglutination and growth rate were analysed as indicators of the secondary and tertiary effects of stress. The results of the mean values after the five trials showed that most indicators followed a consistent pattern with time. Cortisol and growth rate were positively correlated, and cortisol and osmolality were negatively correlated after stress. Analysis of the individual responses throughout all 5 months showed that 6–20% of the fish displayed a consistently high or low response depending on the parameter analysed.     

Physiological stress responses in spotted wolffish (Anarhichas minor) subjected to acute disturbance and progressive hypoxia

The spotted wolffish (Anarhichas minor) is considered a promising species for coldwater aquaculture. It is a sedentary, bottom-dwelling fish which exhibits a calm and “non-stressed” behaviour in captivity. There are, however, no reports on the physiological responses to stressors in this species. In the present study we investigated primary (cortisol secretion) and secondary (glucose mobilization) responses to common aquaculture stressors like disturbance and hypoxia. Pre-stress plasma cortisol levels were within those generally considered representative for unstressed fish (~ 10 ng ml^− 1), whereas basal glucose levels were unusually low (0.3–0.4 mM). After exposure to a short, but severe disturbance challenge (emptying the tank of water for 10 min), the increase in plasma cortisol level was slow and relatively weak, reaching a peak level of 25 ng ml^− 1 4 to 8 h after disturbance. When the fish were exposed to a gradually decreasing oxygen level in the tank, a significantly elevated plasma cortisol level (35 ng ml^− 1) was seen in the fish that remained in the tank until oxygen saturation had decreased to 20% oxygen saturation after 2.5 h. However, a two-fold higher plasma cortisol level (~ 70 ng ml^− 1) was seen in all fish exposed to reduced oxygen levels (60, 40 and 20% oxygen saturation) after 3.5 h recovery in normoxic water. Plasma glucose levels showed only moderate increases (~ 70%) following disturbance and hypoxia challenges. An in vivo injection of ACTH caused a strong elevation of plasma cortisol (peak level ~ 170 ng ml^− 1), demonstrating a high capacity for interrenal steroidogenesis in the spotted wolffish. The slow and relatively weak cortisol response to stressors, and low plasma glucose levels, may relate to the sedentary lifestyle of the spotted wolffish. The stress-response is characterised by a passive (reactive) coping style, which is considered adaptive for farming of this species.     

Immune responses of the sea cucumber Apostichopus japonicus to stress in two different transport systems

We report the effect of transport stress on the immune responses of the sea cucumber Apostichopus japonicus. The total coelomocytes counts, phagocytosis activity, reactive oxygen species production, acid phosphatase activity, phenoloxidase activity and catecholamine levels were measured in A. japonicus following both damp and water transportation methods. With both transport methods, stress resulted in significant elevation of catecholamine levels. Total coelomocytes counts increased following transportation, whereas phagocytic activity and reactive oxygen species production decreased. Damp transport significantly depressed acid phosphatase and phenoloxidase activity in A. japonicus. However, there were no significant differences in acid phosphatase and phenoloxidase activity after water transportation. These results show that transport stress induces modifications in the immune system of A. japonicus and that damp transport elicited more severe stress than water transport. This study increased understanding of the mechanism of transport stress inducing neuroendocrine responses and immune modification in A. japonicus. The information obtained should be helpful to optimize transport process in sea cucumber culture.     

拥挤胁迫后鲫鱼血液皮质醇和溶菌酶水平的变化及对病原的敏感性

分别在拥挤胁迫后第3、10、20、30天对鲫鱼(Carassiusauratus)血液皮质醇水平和血液溶菌酶水平进行了检测,并在每次采集血样后用嗜水气单胞菌(Aeromonashydrophila)进行人工感染以评估鱼体抵抗力。结果显示,血液皮质醇水平在胁迫后几天内显著升高(P<0.005),随后虽有所下降,但至第30天高密度组仍保持较高水平(216.32±11.91)ng/mL。实验初期,血液溶菌酶水平小幅升高,但经过长时间的拥挤后,高密度组血液溶菌酶活性受到极大抑制,显著低于同期对照组(P<0.01)。攻毒后的死亡率与胁迫时间成正相关,在最后一次采样期,高密度组死亡率高达58.3%,而同期对照组仅为16.7%,两者具统计学差异(P<0.01)。实验表明,经过拥挤胁迫后,鲫鱼血液皮质醇水平和溶菌酶水平均发生了变化,短期内,两者成正相关,而长期胁迫则表现为负相关。另外,拥挤胁迫也引起了鱼体抵抗力的下降。     

Identification of novel sRNAs involved in oxidative stress response in the fish pathogen Vibrio alginolyticus by transcriptome analysis

Vibrio alginolyticus as an important pathogen in aquaculture can encounter the oxidative stress produced by the immune system during infection. Previous studies showed that sRNAs have important functions in response to oxidative stress in bacteria; however, less of sRNAs related to oxidative stress response were identified in V. alginolyticus. In this study, a total of 749 novel sRNAs were identified by RNA sequencing; among them, 128 sRNAs were up‐ or downregulated in response to oxidative stress. In addition, 1,870 genes exhibited variation on mRNA levels in oxidative stress response. By analysing the target genes of the sRNAs, we concluded that these sRNAs could regulate expressions of genes responsible for iron transport, catalase, GSH‐dependent defence system, electron transferred and stress response. Moreover, the functions of the sRNAs are also seemed related to the pathogenicity in V. alginolyticus. Based on the results, we constructed the oxidative stress model in V. alginolyticus. This study provides us the first outlook of sRNAs function in oxidative stress response in V. alginolyticus. Furthermore, this study can help us to prevent and control this important opportunistic pathogen in aquaculture.     

Metabolic activity variations in the sea urchins (Paracentrotus lividus) treated with magnesium and subjected to handling stress and aerial exposure

This study assessed the impact of the use of magnesium chloride as a relaxant for the echinoid Paracentrotus lividus on the metabolic activity of individuals subjected to handling stress and aerial exposure. The variation in the gonad moisture content and biochemical composition was compared in sea urchins (P. lividus) treated with magnesium and untreated controls before transport and during 15 days of laboratory acclimation. Sea urchins treated with magnesium maintained higher levels of protein and carbohydrate throughout the trial in comparison with controls. However, the lipid level and fatty acid percentage remained stable during the experiment and no differences were found between treated or non‐treated sea urchins .     

Physiological stress responses to captivity in early developmental stages of the wedge sole Dicologoglossa cuneata (Moreau)

The stress responses in early growth stages of the wedge sole have been studied to determine whether the high cortisol levels described in juvenile fish are present from early developmental stages. Whole‐body cortisol, glucose and lactate contents, as well as biometric parameters in wedge sole larvae were measured at three different stocking densities. Stocking density affected growth‐related variables significantly, and larvae in lower stocking densities grew faster. Survival did not significantly differ among treatments. At hatching, the whole‐body cortisol concentration was 0.33 ± 0.01 ng g^?1 and varied significantly from 0 to 30 days after hatching (DAH) for each stocking density, though values remained stable for the remaining time in the low‐stocking density group. These hormone levels rose significantly (5.17 ± 2.43 to 22.10 ± 4.95 ng g^?1) at the end of the experiment, depending directly on the stocking density. Glucose and lactate‐body concentrations did not vary among treatments. We conclude that the stress responses of wedge sole larvae are detectable from 45 DAH and that stocking density already can be a stressor at that age. As described for juvenile stage, cortisol content values in wedge sole larvae under non‐stressful conditions are one of the highest among those reported in the literature. The captivity conditions could be responsible for this apparently stressful situation, though those values also could be normal in wild specimens.     

Capture and handling stress affects the endocrine and ovulatory response to exogenous hormone treatment in snapper, Pagrus auratus (Bloch & Schneider)

Sexually mature female hatchery‐reared snapper, Pagrus auratus (Bloch & Schneider) were captured from sea cages by handline and injected at first capture (control) or 24 h after capture, transport and subsequent confinement (delayed injection) with either saline, luteinizing hormone releasing hormone analogue, human chorionic gonadotropin, or 17α‐hydroxyprogesterone. Blood was sampled before hormone treatment and again after 168 h, and fish were checked daily for ovulation. Plasma levels of 17β‐estradiol (E2), testosterone (T), 17α, 20β dihydroxy‐4‐pregnen‐3‐one (17, 20βP) and cortisol were determined by radioimmunoassay. The ovulatory response was assessed from the proportion of fish ovulating, ovulation volume, egg quality and fertility. A delay in injection resulted in significantly lower plasma E2 and T levels in response to hormone treatment, smaller ovulation volumes, and poorer egg quality than in control fish. The results are consistent with the generally inhibitory effects of stress on reproduction in fish, and confirm the requirement to treat fish with hormones designed to induce ovulation, as soon as possible after capture and disturbance.     

Partial and total replacement of fish meal by marine microalga Spirulina platensis in the diet of Pacific white shrimp Litopenaeus vannamei: Growth,digestive enzyme activities,fatty acid composition and responses to ammonia and hypoxia stress

In the this study, we evaluated the effect of replacement of fish meal by a marine microalgae Spirulina platensis on growth, digestive enzyme activities, fatty acid composition and responses to ammonia and hypoxia stress in Pacific white shrimp Litopenaeus vannamei (2.6 ± 0.2 g). Experimental diets contained S. platensis at 0%, 25%, 50%, 75% and 100% replacement levels. After 8 weeks of feeding trial, growth parameters and proximate body composition were not significantly different among treatments (p > .05). Amylase and lipase activities did not show any significant differences between control group and other experimental diets (p > .05), while activities of trypsin and chymotrypsin were significantly higher in shrimp fed diet with 50% substitution of microalgae compared to control group. Fatty acid contents, particularly polyunsaturated fatty acids (PUFAs) including arachidonic acid (ARA), docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), were significantly higher in control diet compared to other experimental diets. On the contrary, the majority of fatty acids including the contents of PUFAs in the whole body of L. vannamei fed with different levels of S. platensis were significantly higher compared to those of control group. After 48‐h exposure to ammonia, survival per cent was not statistically different between all groups (p > .05), but in hypoxia challenge, the survival per cent of control group was significantly less than that of treatments fed diets contained S. platensis (p < .05). Altogether, o ur results demonstrated the effectiveness of S. platensis as a reliable protein source for substitution of fish meal in shrimp aquaculture.     

Dietary nucleotide supplements influence the growth,haemato‐immunological parameters and stress responses in endangered Caspian brown trout (Salmo trutta caspius Kessler, 1877)

A feeding trial was conducted to evaluate the effects of Optimûn, a commercial nucleotide (NT) product, on the growth, haemato‐immunological and serum biochemical parameters and stress responses of Caspian brown trout fingerlings (average initial weight of 12.26 g). A basal diet was supplemented with levels of 0 (control), 1.5, 2.5, 3.5 and 5 g NT kg^?1 to formulate five experimental diets. After 8 weeks of feeding trial, fish fed diet with 2.5 g NT kg^?1 had the highest final weight compared with other treatments. The fish fed diets with 2.5 g NT kg^?1 had higher blood protein, albumin, albumin/globulin ratio, red blood cells, white blood cells and lymphocyte content and lower alkaline phosphatase. The lysozyme activity in serum was found to be significantly (P < 0.05) greater in fish fed diet with 2.5 g NT kg^?1. Concerning both stressors (confinement and salinity stress), fish fed diet with 2.5 g NT kg^?1 had lower plasma cortisol and glucose levels. Thus, administration of 2.5 g kg^?1 of the Optimûn dietary nucleotide formula is recommended to promote growth and immunity as well as to enhance stress responses of Caspian brown trout.     

Effects of Sangrovit‐supplemented diet on growth performance,blood biochemical parameters,survival and stress resistance to salinity in the Caspian roach (Rutilus rutilus) fry

This study was carried out to show the effects of Sangrovit on growth, blood biochemical parameters, survival and resistance to salinity stress in the Caspian roach. Fish (1.24 ± 0.13 g) were divided into four groups fed diets containing Sangrovit at different levels: 0 (control), 0.05, 0.1 and 0.15% for 45 days. The results showed that there was a significant increase in final weight, weight gain and specific growth rate in those fish fed 0.15% Sangrovit diets compared with the control (P < 0.05). Food conversion rate of fish fed 0.05% Sangrovit diet was significantly (P < 0.05) lower than the control group. Condition factor was not significantly affected by Sangrovit dosage (P > 0.05). The blood glucose and cholesterol levels were significantly lower in treated groups compared with the control (P < 0.05). The highest total protein was obtained in the 0.15% group and it was significantly higher than in all other groups (P < 0.05). Challenges were carried out after 45 days of feeding, to determine the effect of Sangrovit on resistance to salinity stress. Blood samples were obtained four times after challenge (day 1, 3, 5 and 7) for evaluate haematocrit levels. On the first day, haematocrit increased significantly in all groups compared to day 5 and 7 (P < 0.05). The haematocrit levels decreased gradually in all groups from the third day. Before and after stress, no differences were observed for survival rate among the experimental diets (P > 0.05). These results indicate that the inclusion of 0.15% Sangrovit in the diet of the Caspian roach enhances feed efficiency, fish growth and blood biochemical parameters.     

低鱼粉饲料中添加牛磺酸对青鱼幼鱼生长、肠道修复及抗急性拥挤胁迫的影响

为研究低鱼粉饲料中添加牛磺酸对青鱼幼鱼[初始质量(5.90±0.03)g]生长、肠道修复及抗急性拥挤胁迫的影响,实验以青鱼正常鱼粉组(20%)为对照组,10%鱼粉组为负对照组(I_0组),在低鱼粉饲料中分别添加0.05%(I_(0.05)组)、0.1%(I_(0.1)组)、0.2%(I_(0.2)组)、0.4%(I_(0.4)组)牛磺酸,配制6种等氮等脂饲料,饲养8周。饲养实验结束后,参考生长结果,选择对照组、I_0组、I_(0.1)组、I_(0.4)组进行急性拥挤胁迫实验。结果显示:①与对照组相比,I_0组增重率显著下降,随着低鱼粉饲料中牛磺酸水平的升高,青鱼幼鱼增重率呈先上升后下降趋势,当牛磺酸添加量为0.1%时与对照组差异不显著;②相对于对照组,I_0组的绒毛高度降低、隐窝深度增加,杯状细胞数降低,添加牛磺酸使绒毛高度升高、隐窝深度降低,杯状细胞数和淋巴细胞数增多;③急性拥挤胁迫使青鱼幼鱼血清史质醇、血糖、血清溶菌酶(LSZ)、补体C3、超氧物歧化酶(SOD)和谷胱甘肽(GSH)含量均呈先升高后下降的趋势,其最大值出现在胁迫2 h或者8 h。在整个胁迫期间,I_0组史质醇和血糖含量高于其他各饲料组,I_0组溶菌酶、补体C3、SOD和GSH均低于其他各饲料组。研究表明,在低鱼粉饲料中添加牛磺酸可提高青鱼生长性能、改善肠道结构、增强青鱼的抗急性拥挤胁迫的能力。     

牛磺酸对红鳍东方鲀热应激转录调控机制的影响

为探究牛磺酸对红鳍东方鲀的热应激调控的影响,以初始体重为（32.28±0.20）g的红鳍东方鲀（Takifugu rubripes）作为研究对象,实验随机分为3组,每组设置3个重复,通过在低鱼粉饲料中分别添加3个水平的牛磺酸[0%（T1,对照）、1.0%（T2）和5.0%（T3）],配制3组实验饲料。养殖56 d后,水温（28±0.3）℃,急性热应激30 min,取肝脏。使用RNA-Seq测序技术对3组红鳍东方鲀肝脏转录组进行分析,并分别对3个转录组测序文库进行两两比较,设置显著差异基因筛选条件为P<0.05,共获得167个差异表达基因,其中上调基因111个,下调基因56个。GO功能分析发现,在T3vsT1组中,差异表达基因（DEGs）显著富集在蛋白质分解过程、丝氨酸型内肽酶活性、丝氨酸型肽酶活性、丝氨酸水解酶活性、内肽酶活性、L-氨基酸肽的肽酶活性和肽酶活性。KEGG富集分析发现,T2vsT1组中,这些DEGs主要参与细胞黏附分子、神经活性配体-受体相互作用通路;而T3vsT1组中,这些DEGs主要参与神经活性配体受体相互作用和代谢途径。选取3个显著差异表达基因进行实时荧光定量PCR（RT-qPCR）验证,结果证明,转录组测序分析可靠;饲料中添加牛磺酸后,在急性热胁迫条件下,红鳍东方鲀可通过细胞黏附分子和神经活性配体-受体相互作用通路调控机体对温度的响应;随着牛磺酸添加量的升高,红鳍东方鲀主要通过代谢调节、神经活性配体-受体相互作用通路调控机体对温度的响应。本研究旨为研究牛磺酸对红鳍东方鲀的热应激调控的影响和牛磺酸抗应激功能提供参考数据。     

Effect of L‐tryptophan supplemented diets on serotonergic system and plasma cortisol in Totoaba macdonaldi (Gilbert, 1890) juvenile exposed to acute stress by handling and hypoxia

Brain serotonin and cortisol play a central role in integrating the neuroendocrine response to stress. Previous studies in aquatic species show that dietary supplementation with the 5‐HT precursor L‐tryptophan amino acid is able to modulate this response. The aim of this study was to evaluate the effect of dietary tryptophan (Trp) on the serotonergic system and acute stress response in Totoaba macdonaldi. Four groups of fish were fed with one of four isonitrogenous and isocaloric diets formulated with different Trp content: 0.5% (1X TRP, control diet), 1.1% (2X TRP), 1.7% (3X TRP) and 2.3% (4X TRP). After a feeding period of 21 days, groups were exposed to acute stress by handling with a net or hypoxia. Feeding with Trp‐enriched diets resulted in increased 5‐HT content in telencephalon of undisturbed fish. Handling increased the 5‐hydroxyindole acetic acid (5‐HIAA) and serotonin turnover (5‐HIAA/5‐HT) in telencephalon of fish fed with 1X TRP and 2X TRP but was not significant for the fish fed 3X TRP and 4X TRP, and no significant increase was observed by hypoxia in both telencephalon and hypothalamus in any of the diets. An inverted U‐shaped response was observed in plasma cortisol increasing with 2X TRP after both handling and hypoxia and decreasing with diets rich in Trp, 3X TRP and 4X TRP. A higher increase was observed in haematocrit, plasma glucose and lactate in hypoxia than the handling groups. Our results indicate that Trp supplementation modulated cortisol response and mediated negative feedback of 5‐HT system in telencephalon after acute stress.     

Comparing the behavioural thermoregulation response to heat stress by Atlantic salmon parr (Salmo salar) in two rivers

Climate change is expected to increase the frequency and magnitude of extreme thermal events in rivers. The Little Southwest Miramichi River (LSWM) and the Ouelle River (OR) are two Atlantic salmon (Salmo salar) rivers located in eastern Canada, where in recent years, water temperatures have exceeded known thermal limits (~23°C). Once temperature surpasses this threshold, juvenile salmon exploit thermal heterogeneity to behaviourally thermoregulate, forming aggregations in coolwater refuges. This study aimed to determine whether the behavioural thermoregulation response is universal across rivers, arising from common thermal cues. We detailed the temperature and discharge patterns of two geographically distinct rivers from 2010 to 2012 and compared these with aggregation onset temperature. PIT telemetry and snorkelling were used to confirm the presence of aggregations. Mean daily maximum temperature in 2010 was significantly greater in the OR versus the LSWM (p = 0.005), but not in other years (p = 0.090–0.353). Aggregations occurred on 14 and 9 occasions in the OR and LSWM respectively. Temperature at onset of aggregation was significantly greater in the OR (T_onset = 28.3°C) than in the LSWM (T_onset = 27.3°C; p = 0.049). Logistic regression models varied by river and were able to predict the probability of aggregation based on the preceding number of hours >23°C (R² = 0.61 & 0.65; P₅₀ = 27.4°C & 28.9°C; in the OR and LSWM respectively). These results imply the preceding local thermal regime may influence behaviour and indicate a degree of phenotypic plasticity, illustrating a need for localised management strategies.