Ontogenic changes in the digestive enzyme activities and the effect of different starvation duration on the digestive enzyme activities of larval red swamp crayfish (Procambarus clarkii)

This study was performed to determine the effect of starvation and delayed feeding on activities of digestive enzymes and alkaline phosphatase (ALP) of larval red swamp crayfish (Procambarus clarkii), so as to reveal the tolerance to prolonged starvation and the recovery of digestive enzymes after delayed feeding in larval and juvenile P. clarkii. In the control group, activities of trypsin and ALP increased significantly (p < .05) with day‐age and then kept constant at 24 days after hatching (DAH) and 10 DAH, respectively, whereas the activities of amylase and pepsin increased firstly then decreased with day‐age, and the activity of lipase increased firstly then decreased and then increased again during the development period of juvenile P. clarkii (1–31 DAH). In the group with continuous starvation (CS), activities of pepsin and lipase both decreased (p < .05) after fasting, and the activities of pepsin, lipase and trypsin in the groups with delayed feeding all increased (p < .05) and recover to the levels of the control group after food supply. However, the activity of amylase increased (p < .05) in the CS group, and it decreased to normal level after food supply. The ALP activity did not significantly (p > .05) vary after starvation, whereas it decreased in the groups with delayed feeding after 1 day of food supply, and then increased back to the level similar with the control group. Results from this study could provide information for diet preparation and feeding regime in larval and juvenile red swamp crayfish culture.     

Direct visualization of the novel pathogen,Spiroplasma eriocheiris,in the freshwater crayfish Procambarus clarkii (Girard) using fluorescence in situ hybridization

Spiroplasma eriocheiris is the first spiroplasma strain known to be pathogenic to freshwater crustaceans. It has caused considerable economic losses both in the freshwater crayfish Procambarus clarkii (Girard) and in some other crustaceans. The monitoring of the pathogen in crustacean populations and study of its behaviour in the laboratory require the development of reliable diagnostic tools. In this article, we improved microscopic identification of S. eriocheiris by combining in situ hybridization with specific fluorescently labelled oligonucleotide probes. The established fluorescence in situ hybridization (FISH) allowed simultaneous visualization, identification and localization of S. eriocheiris in the tissues of diseased crayfish P. clarkii and exhibited low background autofluorescence and ideal signal‐to‐noise ratio. With the advantages of better tissue penetration, potentially more specific and stable, we designed three species‐specific oligonucleotide probes utilizing the sequences of 16S‐23S rRNA intergenic spacer regions (ISRs) of S. eriocheiris. Positive hybridization signals were visualized in haemocytes and connective tissues of hepatopancreas, cardiac muscle and gill from diseased crayfish. This unique distribution pattern matched the pathological changes when diagnosed by H&E staining and indicated that S. eriocheiris probably spread throughout the tissues in P. clarkii by hemokinesis. This assay will facilitate our understanding of the pathogenesis of S. eriocheiris and enhance the early diagnosis of the novel pathogen.     

Dietary calcium requirement of red swamp crayfish (Procambarus clarkia)

To investigate dietary calcium requirement of red swamp crayfish (Procambarus clarkia), six semi‐purified diets were formulated to contain different concentrations of calcium (2.7(control group), 6.1, 11.9, 17.6, 23.5 and 29.1 g/kg calcium). Each diet was hand‐fed to triplicate of 15 crayfish with average initial body weight (6.22 ± 0.87) g for 8 weeks. The results showed that weight gain rate (WGR) significantly increased and feed conversion ratio (FCR) significantly decreased from 11.9 to 23.5 g/kg groups (p < .05). Protease activities in intestine and hepatopancreas and parathyroid hormone concentrations in serum significantly decreased with increasing dietary calcium levels (p < .05), while calcium and phosphorus contents in exoskeleton, calcium content in muscle and calcitonin concentrations in serum significantly increased (p < .05). The activities of lipase and amylase in intestine and hepatopancreas, serum alkaline phosphatase and total vitamin D concentrations in serum had significant increase as dietary calcium content increased up to 11.9–17.6 g/kg (p < .05). The inorganic phosphorus content in 29.1 g/kg group was significantly lower than those in other groups (p < .05). Broken‐line model analysis based on WGR and quadratic curve model analysis based on FCR showed that optimal dietary calcium requirement of red swamp crayfish ranged from 12.7 to 17.1 g/kg.     

维生素E对克氏原螫虾生殖的影响

实验用不同的维生素E（VE）含量的饲料投喂克氏原螯虾亲虾，研究对其繁殖性能的影响，目的是为研制亲虾的人工饲料提供指导。结果显示，在添加VE量分别为基础饲料干重的0％、0．004％、0．02％、0．1％和0．5％的5个饲料组中，0．02％VE组不仅可以增加产卵雌虾的数量比例，而且也能提高克氏原螯虾雌虾的产卵量。随着VE含量的升高，产卵雌虾的比例呈下降趋势，单个雌虾的产卵量与0．02％VE组差不多，这表明饲料中适当添加VE可促进克氏原螯虾产卵，但饲料中VE含量过高对克氏原螯虾的繁殖产生负面影响。在饲料节添加适量的VE能够提高亲虾繁殖能力，方法简单，效果明显，适合规模化生产的需要，可以推广。     

Growth performance and muscle composition response to reduced feeding levels in juvenile red swamp crayfish Procambarus clarkii (Girard, 1852)

Overfeeding in aquaculture can lead to suboptimal growth and increased production costs. Red swamp crayfish, Procambarus clarkii, is one of the most noteworthy species cultured in China, but little information is available on the optimal feeding levels of this species, especially in ponds cultured with macrophytes. In this study, we tested the effects of five different feeding levels (20%, 40%, 60%, 80% and 100% satiation) of an artificial diet on growth performance and muscle composition of juvenile P. clarkii in 15 concrete ponds cultured with the macrophyte Hydrilla verticillata (three replicates for each treatment). The results showed that growth performance decreased only when feeding levels were below 60% satiation. Muscle composition analysis revealed that the moisture and ash contents of muscle did not vary significantly with feeding levels but that the lipid and protein contents of muscle significantly decreased when P. clarkii was fed to 40% satiation. Stable isotope analysis suggested a shift in crayfish diets to easily available H. verticillata when feeding levels decreased. With this study, we hope to encourage crayfish farmers to reduce feeding levels and increase natural food items such as macrophytes in cultured ponds to maximize crayfish yields and reduce production costs.     

Bacillus subtilis CK3 used as an aquatic additive probiotics enhanced the immune response of crayfish Procambarus clarkii against newly identified Aeromonas veronii pathogen

To identify the aetiology of the disease outbreak in cultured Procambarus clarkii, a dominant bacteria strain from the diseased crayfish hepatopancreas was isolated. It was identified as Aeromonas veronii based on biochemical identification and 16S rDNA sequencing. The symptoms of artificially infected crayfish were similar to those of naturally infected P. clarkii. The A. veronii was sensitive to norfloxacin, ciprofloxacin, ceftazidime, cefuroxime and ceftriaxone antibiotics, while resistant to penicillin, ampicillin, oxacillin and piperacillin. Virulence genes such as the enterotoxin genes (act, alt and ast) and haemolytic toxin genes (hlyA and aerA) were detected. After A. veronii infection, the connections between some liver tubules disappeared, the vacuoles appeared in the brush border and the mitochondria were enlarged. The antagonistic action of previous identified B. subtilis CK3 against A. veronii was also detected. The supernatant of B. subtilis CK3 exhibits a significant bactericidal effect on A. veronii. After B. subtilis CK3 immersion, the antioxidant enzymes and immune-related enzyme activities in hepatopancreas were significantly higher than control. The accumulative mortality caused by infection of A. veronii can be significantly reduced by adding B. subtilis CK3 into the aquatic water. These results demonstrated that B. subtilis CK3 could act as a water additive to improve the immune response of P. clarkii against newly identified A. veronii. The present study will provide a new way for the prevention and control of crayfish bacterial diseases and also provide technical support for the healthy cultivation of P. clarkii.     

饲料中赖氨酸水平对克氏原螯虾生长、体组成与消化酶活性的影响

克氏原螯虾(Procambarus clarkii)购自盱眙某龙虾养殖场,初始体质量为(7.60±0.01)g。以酪蛋白、明胶、小麦面筋粉和晶体氨基酸为蛋白源配制7组蛋白质水平为28.26%、能量为17.15 kJ.g 1的等氮等能半纯和饲料,各组赖氨酸质量分数分别为0.76%、1.00%、1.22%、1.47%、1.69%、1.89%和2.18%。实验虾分为7组,每组设4个重复,每个重复12尾虾,分别投喂以上7种饲料。养殖实验于室外网箱(规格为60 cm×60 cm×60 cm)中进行,饲养周期为60 d。结果表明,随着饲料中赖氨酸水平的升高,增重率、特定生长率和饲料效率均显著升高(P<0.05),分别在赖氨酸水平1.69%、1.69%和1.22%组时,三者达到最大值;当赖氨酸水平进一步升高,三者均呈下降趋势。各组的成活率为89.58%~97.92%,组间无显著差异(P>0.05)。全虾水分含量以赖氨酸水平1.22%组最低,显著低于0.76%组(P<0.05)。全虾粗蛋白和粗脂肪含量分别以赖氨酸水平1.89%组和1.69%组最高,均显著高于0.76%组(P<0.05)。各组之间的全虾灰分含量、肌肉组成、肌肉氨基酸组成和消化酶活性均无显著差异(P>0.05)。以增重率为评价指标进行一元二次回归分析,得出克氏原螯虾的适宜赖氨酸需求量为1.66%,占饲料蛋白的5.87%。本研究旨在通过确定克氏原螯虾对饲料中赖氨酸的适宜需求量,为其人工配合饲料的研制提供基础数据和理论依据。     

维生素E对克氏原螯虾生殖的影响

实验用不同的维生素E(VE)含量的饲料投喂克氏原螯虾亲虾,研究对其繁殖性能的影响,目的是为研制亲虾的人工饲料提供指导.结果显示,在添加VE量分别为基础饲料干重的0%、0.004%、0.02%、0.1%和0.5%的5个饲料组中,0.02%VE组不仅可以增加产卵雌虾的数量比例,而且也能提高克氏原螯虾雌虾的产卵量.随着VE含量的升高,产卵雌虾的比例呈下降趋势,单个雌虾的产卵量与0.02%VE组差不多,这表明饲料中适当添加VE可促进克氏原螯虾产卵,但饲料中VE含量过高对克氏原螯虾的繁殖产生负面影响.在饲料中添加适量的VE能够提高亲虾繁殖能力,方法简单,效果明显,适合规模化生产的需要,可以推广.     

克氏原螯虾的适宜蛋氨酸需求量

以酪蛋白、明胶、鱼粉、小麦面筋粉和晶体氨基酸为蛋白源配制了6组蛋白质水平为28.25%、能量为16.07 kJ/g的等氮等能的半纯合饲料, 各组蛋氨酸实际含量分别为0.39%、0.53%、0.71%、0.86%、0.98%和1.13%。以初始体质量为(9.80±0.15) g的克氏原螯虾(Proambarus clakii)为试验对象, 每组设4个重复, 每个重复12尾虾。养殖试验于室外网箱(规格为60 cm×60 cm×60 cm)中进行, 饲养周期为56 d。结果表明, 当饲料中蛋氨酸水平由0.39%升高至1.13%时, 克氏原鳌虾的增重率、特定生长率和饲料效率均显著升高(P<0.05)。当蛋氨酸水平为0.86%时, 三者均达到最大值。当蛋氨酸水平进一步升高时, 三者均呈下降趋势。试验各组的成活率为83.33%~91.67%, 但无显著差异(P>0.05)。当蛋氨酸水平为0.86%时, 全虾粗蛋白含量最高, 其显著高于0.39%组(P<0.05)。日粮蛋氨酸水平对全虾的水分、灰分和脂肪含量以及肝胰脏组成和肌肉氨基酸组成均无显著影响(P>0.05)。肌肉粗脂肪含量以蛋氨酸水平0.98%组最低, 其显著低于0.39%组(P<0.05)。饲料中蛋氨酸水平显著影响肠道蛋白酶的活性(P<0.05), 但对脂肪酶和淀粉酶活性均无显著影响(P>0.05)。以增重率为评价指标进行一元二次回归分析, 得出克氏原螯虾的适宜蛋氨酸需求量为0.94%, 占饲料蛋白质的3.36%。本研究通过分析饲料中不同蛋氨酸水平对克氏原螯虾生长、饲料利用、虾体组成、肌肉氨基酸组成和消化酶活性的影响, 确定克氏原螯虾的适宜蛋氨酸需求量, 以期为其人工配合饲料的研制提供基础数据和理论依据。

     

Inducing synchronous ovarian maturation in the crayfish,Procambarus clarkii,via eyestalk interventional injection as compared with eyestalk ablation and combined injection of serotonin and domperidone

We examined the feasibility of inducing synchronous ovarian maturation in the crayfish, Procambarus clarkii, via eyestalk interventional injection (EI) using a Bletilla striata polysaccharide (BSP) gelatin containing tranexamic acid (TRA), domperidone (DOM) or serotonin (5‐HT). In total, 360 females were randomly assigned to seven experimental groups and a control group. The average survival rate (SR%) and average synchronous ovarian maturation rate (SMR%) in survivors in the EI groups, EI‐TRA, EI‐DOM, EI‐5HT were compared with bilateral eyestalk ablation (BEA), unilateral eyestalk ablation (UEA), abdominal injection (AI) of DOM (0.5 mg crayfish^?1) alone (AI‐DOM) or combined with 5‐HT (0.5 mg crayfish^?1) (AI‐DOM+5‐HT). The experiment covered a prolonged period of 32 days until two ovigerous females were observed in BEA. EI‐DOM achieved a SMR (66.67 ± 9.62%) higher than the control (22.05 ± 3.06%) (P < 0.05), but lower than BEA (88.89 ± 11.11%), UEA (63.33 ± 18.56) and AI‐DOM+5‐HT (59.26 ± 10.14%) (P > 0.05). EI‐DOM also achieved a higher SR (53.33 ± 3.85%) than BEA (15.56 ± 2.22), UEA (24.44 ± 5.88) (P < 0.05) and AI‐DOM+5HT (51.11 ± 4.44) (P > 0.05). However, EI‐TRA (SR = 28.89 ± 4.44, SMR = 37.78 ± 2.22) and EI‐5‐HT (SR = 15.56 ±4.44, SMR = 22.22 ± 11.11) failed to induce significantly higher SMR than the control. These findings suggest that EI methods, such as EI‐DOM, may have positive characteristics for the development of inexpensive and less labour‐intensive techniques for induction of ovarian maturation in decapods.     

饲料蛋白质水平和加工工艺对克氏原螯虾生长性能、消化力和抗氧化性能的影响

为研究不同蛋白质水平和加工工艺饲料对克氏原螯虾(Procambarus clarkii)生长性能、消化力和抗氧化能力的影响,本研究采用4个蛋白质水平(28%、30%、32%和36%)和2种加工工艺即膨化饲料(EF)和硬颗粒饲料(PF)的4×2双因素实验设计,进行为期13周的养殖实验。结果显示,蛋白质水平和加工工艺对克氏原螯虾增重率和肝体比有显著的交互作用(P<0.05)。主效应结果显示,28%蛋白质可显著提高增重率和特定生长率,EF组的末重和增重率显著高于PF组(P<0.05)。蛋白质水平和加工工艺对肝胰脏的胰蛋白酶和淀粉酶活性有交互作用(P<0.05)。主效应结果显示,肝胰脏的胰蛋白酶、淀粉酶和肠道胰蛋白酶、淀粉酶均在28%蛋白质水平时最高,EF组肝胰脏胰蛋白酶、淀粉酶和肠道胰蛋白酶活力高于PF组(P<0.05)。蛋白质水平和加工工艺对肝胰脏的丙二醛和碱性磷酸酶活性有显著的交互作用(P<0.05)。主效应结果显示,肝胰脏碱性磷酸酶活性以及血清的碱性磷酸酶和酸性磷酸酶均在28%蛋白质水平时最低。PF组肝胰脏丙二醛含量显著低于EF组(P<0.05),碱性磷酸酶活性显著低于EF组(P<0.05)。4种模型(Logistic、Gompertz、Von Bertalanffy和Brody)均能很好地模拟克氏原螯虾体重和体长的生长曲线,拟合度均大于0.97,其中,以Logistic模型拟合克氏原螯虾体重和体长最优,且幂函数可以反映体重和体长之间的关系。综上所述,28%蛋白质水平和膨化加工工艺使克氏原螯虾具有最优的生长性能和消化力,28%蛋白质水平的硬颗粒饲料可提高克氏原螯虾的抗氧化能力。     

Growth responses of juvenile red swamp crayfish, Procambarus clarkii Girard, to several diets under controlled conditions

The growth responses of juvenile crayfish, Procambarus clarkii Girard, to diets based on common fresh feedstuffs available from agriculture and freshwater ecosystems were studied under controlled laboratory conditions. The dietary treatments were: fresh leaves and shoots of watercress, fresh potato tubers, fresh fish meat from mosquitofish, and all the possible combinations of those ingredients on alternate feeding days. After 22 days, the results showed a significantly higher growth rate (P= 0.05) in crayfish fed with mosquitofish meat. The differences were not significant between the other treatments, but those which included fish meat induced growth rates consistently higher than those based only on plant ingredients. This trend was maintained until the end of the experiment. These results suggest that a dietary supplement based on animal ingredients might significantly improve growth rate and yield, provided the economic feasibility of such a production process be confirmed in field trials. However, it was not clear whether this was because of protein, or lipid concentration and composition of mosquitofish meat. Further research must also clarify the role of vegetable ingredients in dietary composition, as well as the effect of microbial populations occurring during the decomposition of plant material on its nutritional properties for crayfish.     

饲料中蛋白质水平对克氏原螯虾生长性能及肌肉品质的影响

以初始平均体质量为(3.69±0.05)g的克氏原螯虾幼虾为研究对象,配制蛋白质水平分别为27.72％、30.09％、33.05％、35.31％和37.57％的5种等能、等脂的配合饲料,进行为期47 d的饲养试验,探索饲料中不同蛋白质水平对克氏原螯虾生长性能、肌肉质构特性、常规营养成分及氨基酸组成和含量的影响.结果 表...     

复方中草药对克氏原螯虾生长和脱壳的影响

研究了饲料中添加复方中草药(质量分数为1%,2%和4%)对克氏原螯虾(Procambarus clarkii)生长和脱壳的影响。结果显示,添加复方中草药后,虾的脱壳周期缩短,脱壳死亡率和非脱壳死亡率均明显降低(P<0.05),但各中草药添加组间差异不显著(P>0.05)。三个剂量的复方中草药添加组均能不同程度地提高虾的脱壳数、成功脱壳数、硬壳虾重和软壳虾重,与未添加组相比差异显著(P<0.05),其中2%组增加最为显著,分别比对照组高出71.1%、103.2%、20.3%、10.0%。回归分析表明,克氏原螯虾的特定生长率达到最大值时的中草药添加质量分数为2.74%。试验表明克氏螯虾饲料中复方中草药水平以2%左右为宜。     

Mixed model estmation of genetic and environmental correlations in red swamp crawfish Procambarus clarkii (Girard)

Total length, total weight, carapace length, carapace width, abdomen width, abdomen depth, chela length, chela width, abdomen meat weight and dressout percentage were measured on 752 individual crawfish Procambarus clarkii (Girard) from 39 full-sib families nested within 15 half-sib families. A nested mixed model analysis of variance and covariance was performed on all traits to obtain Henderson's method III variance and covariance components, as well as best linear unbiased estimators of the fixed effects of sex and male state of maturity. Genetic and environmental correlations were derived from mixed model variance and covariance components. Positive sire-based genetic correlations suggest that genetic gains in growth and processing yields, the major components of value of harvestable P. clarkii, may be pursued simultaneously in future breeding programmes. Data suggest that easily obtained external measurements are sufficiently correlated with growth and processing traits to allow for identification of potentially superior broodstock. Female, mature male and immature male levels of fixed effects, as well as estimated heritabilities for individual growth, body size and processing traits, were consistent with previous studies, further reinforcing the validity of data developed to date for use in planning P. clarkii breeding programmes.     

克氏螯虾壳聚糖抗疲劳作用研究

克氏原螯虾是当今中国人工养殖最广的淡水螯虾品种。近几年来,稻田养虾面积呈暴发式增长,但随着养殖规模的扩大,养殖环境的恶化,各种病害频发。为探明稻田养殖克氏原螯虾死亡的原因,实验取濒死病虾胃和肠组织进行病原菌分离,用生理生化实验、16S rDNA基因序列测定与系统进化分析鉴定优势菌;用回归感染和药敏实验评价优势菌毒力和药物敏感性,通过血清免疫相关酶活性变化探究优势菌的致病机理。结果显示,分离获得1株优势菌PCW-01,回归感染健康克氏原螯虾,出现与自然发病虾相同的病理症状,活力下降,胃部有积液,半致死量达0.60×10⁵ CFU/g,经理化分析和分子鉴定为维氏气单胞菌,对呋喃妥因、妥布霉素、头孢他啶等7种药物敏感。人工感染后的克氏原螯虾,体内血清谷丙转氨酶(ALT)和溶菌酶(LZM)活性极显著上升,总抗氧化能力(T-AOC)、总超氧化物歧化酶(T-SOD)和谷胱甘肽过氧化物酶(GSH-Px)活性极显著下降,丙二醛(MDA)含量略有上升。研究表明,优势菌PCW-01为致病菌,可失活抗氧化因子,破坏细胞膜结构,导致疾病发生。本研究结果揭示出了克氏原螯虾细菌性疾病的免疫应答规律,为其细菌性疾病的防控提供了科学依据。