Results of comparative growing test of all-female and bisexual population in two-year-old common carp (Cyprinus carpio L.)

The aim of this study was to verify whetherall-female carp populations grow faster thanbisexual ones. All-female mirror and scalypopulations were obtained by crossingmasculinizated females (neomales) of Northernmirror carp (M72) with regular females of M72and scaly Ropsa carp (ROP) purebreds,respectively. All-female and bisexual progenyof both mirror and scaly carps were rearedseparately for one year. However, controlgroups of a distinct phenotype of scalinesswere put in ponds with tested fish to make anadjustment for fish weight, which could becaused by environmental influence. Nodifferences were found after eight weeks ofrearing between all-female and bisexualpopulations, but, all-female mirror carppopulation (M72) gained 29.7% more weight(P < 0.0001) by the end of the first year thanthe bisexual (68.5 g ± 15.8 against 52.8 g± 13.9; mean ± S.D., respectively).Before beginning the second growing season,each group was marked and stocked together intriplicate ponds. Corrected weight after secondgrowing season (P < 0.0062) was 9.9% higher inall-female mirror carps (395.9 g ± 98.6)compared to the bisexual (360.2 g ± 89.0).No significant differences were found incorrected weights between scaly all-female andbisexual crossbreds (522.4 g ± 102.4 and518.3 g ± 95.8, respectively). Nodifferences in survival of all-female andbisexual populations were observed.     

一种免疫诱导型鲤启动子的克隆与功能分析

为发掘适用于基因工程抗病育种的鱼类启动子,通过实时荧光定量PCR实验对鲤Rab GTP酶(Ras-associated binding-GTPases 1a3,Rab1a3)基因的表达模式进行了分析,证实该基因在鳃、头肾等与机体免疫防御功能密切相关的组织内转录水平较高,且免疫激活后转录显著增强,符合基因工程抗病育种所需的外源免疫基因转录模式。从鲤细菌人工染色体文库中,使用Rab1a3基因特异引物筛选获得包含该基因区域的文库克隆,测序获得该基因完整序列,以及上下游调控序列。通过生物信息学手段,预测到长度为1014 bp的鲤Rab1a3基因启动子序列,该启动子不具有典型的TATA盒或CpG岛特征,存在多个免疫相关转录因子结合位点。在草鱼肾组织细胞系内验证该启动子活性,结果显示,绿色荧光蛋白基因和萤火虫荧光素酶基因都能够在该启动子驱动下表达,证实该片段具有启动子活性,且启动子活性在受到免疫诱导后增强,双荧光素酶报告基因检测结果显示,该启动子活性在免疫刺激后增强至免疫刺激前的8.67倍。研究表明,鲤Rab1a3基因启动子有望被开发成为免疫诱导型的基因工程元件,驱动外源免疫基因在鱼体内适时表达,抵御外界病原感染,同时避免非必要条件下的过度表达形成生长负担。     

盐度对鲤能量收支的影响

以水蚯蚓为饵料，在温度为２７℃，盐度为淡水、３、５、７、９的条件下对幼鲤（初始体重为２．１９－３．３１ｇ）进行了摄食－生长实验，每一盐度下设４个摄食水平（饥饿－饱足），测定了最大摄食率、吸收率、特定生长率、转化效率和能量收支。结果表明：盐度对最大摄食率、特定生长率（ＳＧＲ）和转化效率有显著影响；盐度对排出废物所占比例（Ｅ／Ｃ）影响不显著，对代谢能所占比例（Ｒ／Ｃ）和生长能所占比例（Ｇ／Ｃ）有显著影     

鲤Rhbdd3蛋白的抗病毒功能

Rhbdd3(Rhomboid domain-containing protein 3)蛋白在哺乳动物天然免疫中发挥了重要作用,但水生动物中rhbdd3基因的确定序列及Rhbdd3蛋白的功能均尚未见报道。为研究鲤(Cyprinus carpio)的Rhbdd3蛋白在鱼类细胞中的功能,探讨其过表达对鱼类病毒感染的影响,本研究通过PCR扩增得到了鲤rhbdd3基因的编码序列,并将其克隆至pCI-neo载体上,构建了真核表达质粒pCI-rhbdd3。pCI-rhbdd3转染鲤上皮瘤细胞EPC(epithelioma papulosum cyprinid)和鲑囊胚细胞CHSE-214(chinook salmon embryo)后利用制备的特异性抗体进行Western blot,检测Rhbdd3蛋白的表达情况,并利用CCK-8试剂检测其过表达对细胞增殖的影响。转染后分别进行鲤春病毒血症病毒(SVCV)和传染性胰腺坏死病毒(IPNV)的感染实验,并利用间接免疫荧光、Western blot和RT-qPCR方法检测Rhbdd3过表达对SVCV和IPNV增殖的影响。结果显示,pCI-rhbdd3转染后Rhbdd3蛋白在EPC和CHSE-214细胞中得到了过表达,且Rhbdd3蛋白的过表达能显著抑制SVCV和IPNV的复制,但不影响两种细胞的正常活性。本研究为鱼类广谱抗病毒药物的开发提供了新的实验依据,也为鱼类抗病毒新品种的培育奠定了重要基础。     

鲤硒缺乏的病理学研究

将健康幼鲤360尾设置3个相同平行处理,每个处理用鱼120尾,随机分为四组,分别投以0 mg/kg、0.15 mg/kg、0.30 mg/kg和0.45mg/kg硒水平的饲料,各试验组的发病率和死亡率与日粮中硒含量高低呈负相关,其发病率分别为46.7%、33.3%、13.3%和0%,死亡率分别为26.7%、16.7%、6.7%和0%。病鱼出现特征性的“瘦背病”和脊柱弯曲等症状。组织学上最突出的变化表现为营养性肌病、营养性肝病、胰腺的变性、坏死等多器官组织的退行性变化。超微结构上,骨骼肌纤维、肝细胞和肾小管上皮细胞线粒体肿胀,嵴断裂,甚至发生溶解,整个线粒体呈囊泡状。硒缺乏幼鲤血液中谷草转氨酶（AST）、谷丙转氨酶（ALT）活性升高,谷胱甘肽过氧化物酶（GSH-Px）、过氧化物歧化酶（SOD）活性降低,丙二醛（MDA）含量升高。     

Effects of cyclic and regular feeding of a Quillaja saponin supplemented diet on growth and metabolism of common carp (Cyprinus carpio L.)

Fifteen common carp weighing 19.2±2.9 g were reared individually in chambers of a respirometer system which allowed feeding, and continuous measurement of oxygen consumption. The fish were divided into three groups of 5 each, and were given control feed (C group), control feed supplemented with 150 mg kg⁻¹ of Quillaja saponins (QS) (S150 group), and control and the saponin supplemented diet during alternate weeks (S150b group), for 56 days. Supplementation with QS at a level of 150 mg kg⁻¹ caused a significant (P<0.05) increase in metabolic growth rate, food conversion efficiency (FCE), protein productive value (PPV) and apparent lipid conversion (ALC) of carp, even as the metabolic rate was lower than the C group. The efficiency of energy utilisation indicated by parameters such as energy expenditure (EE), energy retention (ER) and amount of oxygen consumed per unit body mass gain (OPM) were also significantly (P<0.05) better in the S150 group. The performance of the S150b group was intermediate between the C and S150 group in this experiment. Possible reasons for the effects of dietary QS are discussed. It was concluded that there has to be continuous dietary supply of QS at the level of 150 mg kg⁻¹, for maximum positive effects of these substances on growth of common carp. This revised version was published online in August 2006 with corrections to the Cover Date.     

Stimulation of gonadal and germ cell development in larval and juvenile carp (Cyprinus carpio L.) by homologous pituitary extract

The data presented show that in larval carp, gonadal size has increased distinctly after treatment with homologous pituitary extract (PE). Moreover, the precocious onset of primordial germ cell proliferation and of sex differentiation into male and female gonads was induced. Body weight of treated and control specimens did not differ significantly from each other throughout the experiment. Treatment of juvenile carp with homologous PE led also to an increase in gonadal size without concomitant increase in body weight. Induction of precocious spermatogenesis was observed too. GTH-levels in control and PE-treated carp were monitored by means of a homologous radioimmunoassay, showing that in PE-treated carp the GTH-level was distinctly elevated. The possible role of pituitary hormones in larval and juvenile gonadal development is discussed.     

Testing of performance in common carp Cyprinus carpio L. under pond husbandry conditions I: top-crossing with Northern mirror carp

Performance tests of Northern mirror carp (M72) and three different crossbreds were conducted in semi‐intensive pond conditions from the larval stage up to market size. The aim of this study was to evaluate the possible utilization of new hybrid combinations (M72 × PL – Poho?elice mirror carp) and (M72 × Dor – Israeli mirror carp), and to compare them with a hybrid widely reared as a commercial product (M72 × M2 – Hungarian mirror carp) and with M72 purebred. The testing model was derived from similar studies carried out in the conditions of the Czech Republic, where an internal control of different scale cover is used to eliminate all non‐genetic effects. ancova revealed at market size significantly better growth in M72 × Dor (1 619±28.3 g; LS mean±SE) and M72 × PL (1475±29.8 g) compared with M72 (1278±26.3 g). Survival during the test of 3 years duration was significantly the best in the M72 × Dor hybrid as were the dressing out parameters measured at market size, even if these were significantly different only in proportion of fillets with skin (39.1±0.28%). Interestingly, the crossbred M72 × M2 commonly used by Czech fishermen showed for both, growth (1371±26.2 g) and survival, the lowest values among crossbreds and with regard to dressing out parameters mostly among all tested groups.     

Reciprocal hybrids of tench Tinca tinca (L.) × bream Abramis brama (L.), and tench × carp Cyprinus carpio L., and some characteristics of their early development

Reciprocal hybrids between tench Tinca tinca (L.) and carp Cyprinus carpio L., and tench and bream Abramis brama (L.) were produced artificially. The survival of all these hybrids during embryogenesis was quite high. The highest survival rate (over 60%) at the eyed stage was observed for tench and bream hybrids (both sex combinations). The hatching rates of these hybrids were also over 60%. The number of larvae with some abnormalities (i.e. deformed body) was low. In contrast, the hatching rates of tench and carp hybrids were very low (0.2%). From over 1000 fertilized eggs, only three specimens started swimming, and only one specimen survived to juvenile stage. Embryos of hybrids and their parental species differ in morphological features. These differences were also visible in the juveniles. Body parameters of juvenile hybrids produced from three species had intermediate values in comparison to parental fish.     

Influence of supplemental cereal feeding on the content and structure of fatty acids during long-lasting storage of common carp (Cyprinus carpio L.)

Supplemental cereal feeding (maize, wheat and triticale compared with a control group with natural food only) and its effect on fatty acid (FA) expression in the flesh during long-lasting storage of common carp (Cyprinus carpio) was investigated. The fish were cultured in earthen ponds in the Trebon region (Czech Republic). The content of fatty acid was investigated in the flesh of carp during 8 months of long-lasting storage without additional feeding. Sixty common carp in their third year of life were used for the analyses. The weight of the fish (marketable fish) ranged from 1,358 g to 2,221 g. Polyunsaturated fatty acid (PUFA, n–3) content and composition in fish flesh were determined by gas chromatography (VARIAN 3300). Supplemental cereals caused lower levels of PUFAs and n–3 PUFAs in fish fat. The content of these fatty acids did not decrease, even during 8 months of fish storage. The average percentage of PUFAs in total fat from edible parts was: for maize 13.7% ± 1.58%, for wheat 11.6% ± 1.17% and for triticale 10.7% ± 1.00%. The percentage of n–3 PUFA for maize was 2.5% ± 0.36%, for wheat 3.38% ± 0.44% and for triticale 3.1% ± 0.39%.     

Influence of nutrition on the lipid quality of pond fish: common carp (Cyprinus carpio) and tench (Tinca tinca)

Like marine fish freshwater fish are an important source of essential fatty acids for human nutrition. However, the fatty acid composition of pond fish can vary considerably and strongly depends on that of the ingested food. Investigations on the fatty acid composition of common carp (Cyprinus carpio) and tench (Tinca tinca) have shown that different methods of rearing and feeding cause substantial variations in the proportions of the n-6 and n-3 polyunsaturated fatty acids of these fish species. Carp reared on the basis of natural food in ponds exhibit high contents of n-6 as well as n-3 fatty acids in their muscle triacylglycerols. On the other hand carp fed supplementary wheat in ponds resulted in somewhat lower levels of these essential fatty acids. High amounts of n-3 fatty acids can be found in carp fed high-energy diets containing high levels of fish oil. Analogous results were obtained in experiments with tench reared under different nutritional conditions. While rearing on the basis of only natural food in ponds as well as feeding supplementary wheat yielded in similar levels of n-3 and n-6 polyunsaturated fatty acids, higher contents of n-3 fatty acids were recorded in tench fed pellets. High levels of n-3 polyunsaturated fatty acids in foodstuffs have positive effects on human health. Experiments with different cultured fish species proved that the fatty acid composition of the edible parts can be influenced by the diet. Therefore, a finishing diet with a suitable fatty acid profile can be used to improve the nutritional quality of fish products of farmed origin.     

Artificial spawning of carp, Cyprinus carpio (L.)

The effect of reproduction was investigated on females of Hungarian strain W, French strain F, and their cross‐breed 1X whose ovulation was stimulated with carp pituitary (0.3 mg kg^?1and, after 12 h, 2.7 mg kg^?1) or Ovopel (one‐fifth of a pellet per kg and, after 12 h, one pellet per kg). It was found that in the case of Ovopel treatment, the percentage of spawning females of strain F and the cross‐breed 1X was higher than in the hypophysed fish compared. The applied ovulation stimulators did not significantly affect the weight of obtained eggs, whereas the significant (P ≤ 0.01) effect was recorded with respect to the quality of eggs after 12‐, 24‐, 36‐ and 48‐h of incubation. After Ovopel stimulation, the quality of eggs was better. The origin of the females had no statistically significant effect on the weight of eggs although the yield of eggs from fish of strain W was much smaller than that from females of strain F and the 1X cross‐breed. The interaction between the ovulation stimulator and the provenance of the females was significant (P ≤ 0.05) for the percentage of live embryos after 48‐h of incubation of eggs. Eggs of the best quality (and highest weight) were obtained from fish of strain F and cross‐breed 1X treated with Ovopel. In females of strain F that spawned within 6 and 10 h after the second Ovopel injection, the effect of the ovulation time on the weight of eggs was non‐significant. It was significant with respect to the percentage of egg fertilization and of live embryos after 36‐h of incubation (P ≤ 0.01 and P ≤ 0.05 respectively). The better quality of eggs (and their higher weight) was recorded when this time was shorter.     

First detection of spring viraemia of carp virus in common carp (Cyprinus carpio L.) affected by a septicaemic disease in Mexico

Spring viraemia of carp (SVC) is an infectious disease responsible for severe economic losses for various cyprinid species, particularly common carp (Cyprinus carpio carpio). The causative agent is the SVC virus (SVCV), a member of the Sprivivirus genus, Rhabdoviridae family, and a List 1 pathogen notifiable by the World Organization for Animal Health. This study describes the diagnosis of an SVCV pathogen isolated in October 2015 from wild common carp inhabiting a natural lagoon in central Mexico. While neither an epidemic nor fish mortalities were reported, the collected killed specimens exhibited clinical signs of disease (e.g., exopthalmia, moderate abdominal distension and haemorrhaging, as well as internal haemorrhages and adhesions). Histological results of injuries were consistent with the pathology caused by SVCV. This finding was supported by the isolation of a virus in EPC and BF‐2 cells and subsequent RT‐PCR confirmation of SVCV. The phylogenetic analyses of partial SVCV glycoprotein gene sequences classified the isolates into the Ia genogroup. These findings make this the first report of SVCV detection in Mexico, extending the southern geographical range of SVCV within North America. However, since this pathogen was detected in fish inhabiting a natural body of water without tributaries or effluents, it is difficult to estimate the risk of SVCV for other wild/feral cohabitating cyprinid species in the lagoon. The status of this virus is also unknown for other bodies of water within this region.     

锦鲤microRNA-137系统进化、靶基因验证及表达分析

为了探究microRNA-137(miR-137)与锦鲤体色形成的关系,实验对已报道的14种鱼类miR-137前体序列(precursor miR-137, pre-miR-137)进行比对,利用最大似然估计法(Maximum Likelihood Estimate, MLE)构建系统进化树,同时荧光定量PCR (quantitative realtime PCR, qRT-PCR)分析其在体色发生阶段和不同组织中的相对表达水平,并预测其靶基因,随后对靶基因进行GO和KEGG分析,最后通过荧光素酶实验验证miR-137-3p与靶基因的靶向关系,并分析miR-137-3p与靶基因mRNA的表达变化。结果显示,锦鲤与鲤的pre-miR-137序列相似度最高,且pre-miR-137序列在所有鱼类中具有较高的保守性。对miR-137靶基因进行GO和KEGG分析,多个靶基因富集在色素沉积、色素细胞分化等信号通路。定量结果显示,锦鲤出膜后11 d(day post hatching, dph) miR-137-3p表达量达到最高,随后显著降低;miR-137-3p在锦鲤不同组织中均有表达,在眼和肌肉中表达量较高,在皮肤和鳍条等色素细胞存在的组织中也有较高表达,且白色组织表达量显著高于红色组织。荧光素酶实验结果显示,miR-137-3p可结合在mitfa 3′-UTR上并抑制其表达,但对sprb并无显著抑制作用;miR-137-3p与mitfa和sprb在不同发育阶段存在显著负相关,但在组织中不存在相关性。本研究发现,miR-137在鱼类进化过程中高度保守,与锦鲤体色形成具有一定的相关性,且可靶向调控mitfa参与体色的形成,以上结果为进一步探讨miR-137在锦鲤体色形成中的作用提供基础数据。     

Effects of feeding time,water temperature,feeding frequency and dietary composition on apparent nutrient digestibility in rainbow trout Oncorhynchus mykiss and common carp Cyprinus carpio

A series of experiments was conducted to evaluate the effects of feeding time (daytime vs nighttime feeding), dietary fat content (8 and 20%), feeding frequency (trout 1–3 times/day, carp 2–7 times/day), water temperature (trout 18 and 11°C, carp 25 and 17°C), on the apparent nutrient digestibility in rainbow trout and common carp. The feeding time had little effect on the macronutrient digestibility in both species. In trout, starch digestibility decreased with the decrease of water temperature and with increase of feeding frequency, but in carp, increase of the feeding frequency markedly decreased the macronutrient digestibility and phosphrrus absorption of the high fat diet. Fat digestibility of the beef tallow diet decreased relative to the pollock oil diet in carp, without affecting the phosphorus absorption. Inclusion of raw starch, the digestibility of which was lower than that of gelatinized starch, increased the phosphorus absorption in carp. These results suggest that reduction of water temperature and increase of feeding frequency notably decreased starch digestibility in trout while in carp, the effects of feeding frequency and water temperature on macronutrient digestibility and phosphorus absorption are notable for a high fat diet.     

Pharmacokinetics of chloramphenicol in carp (Cyprinus carpio L.) after drug administration

Chloramphenicol (CAP) is a banned substance in fish farming; yet, residues are sometimes detected in farmed fish products. Therefore, it is important that tissue distribution and elimination rates of CAP are measured to allow monitoring of fish farms. The tissue distribution and residue of CAP in carp (Cyprinus carpio L.) after muscle injection was investigated. Chloramphenicol concentrations were determined using high‐performance liquid chromatography. Pharmacokinetics were used for the studies of CAP residue and the results showed that the CAP was easily assimilated in the liver, serum and gill, with the absorption rates of 6383, 3.02 and 1.39 h^?1, respectively, following a single‐dose injection of 80 mg kg bw^?1. The elimination half‐lives (t_1/2) were 22.28, 15.47, 14.87, 9.28 and 5.32 h for the liver, serum, gill, muscle and kidney respectively. The t_1/2 after 5 days of multi‐doses of repeated CAP injection with 40 mg kg bw^?1 was 1.09, 0.887, 0.872, 0.476 and 0.617 days for the five tissues respectively. The results validated that CAP in the liver, serum and gill could be markers for CAP residue analysis.     

Substrate concentration affects the in vitro metabolism of 17-hydroxyprogesterone by ovaries of the carp,Cyprinus carpio

Carp ovarian tissue was incubated with ³H-17-hydroxyprogesterone in the presence of 0, 0.1, 1, 10, and 100 μg ml⁻¹ unlabeled 17-hydroxyprogesterone. The pattern of metabolites formed showed a marked variation with substrate concentration. Formation of glucuronide and sulphate conjugates was important only at low substrate concentration. At high substrate concentration (10 and 100 μg ml⁻¹) 17,20α-dihydroxy-4-pregnen-3-one was the major metabolite, but at intermediate concentrations polar 7α-hydroxypregnanetetrols predominated. The results support the hypothesis that at low substrate concentrations conjugating, 5α-reducing and 7α-hydroxylating enzymes, of high activity but low capacity, act as scavengers to deactivate any steroids formed during the relatively low pituitary gonadotrophin secretions which are necessary for oocyte development, but that during the prespawning gonadotrophin surge when high levels of substrate are present these enzymes are saturated and 17,20α-dihydroxy-4-pregnen-3-one (17,20αP) becomes the major ovarian steroid. The possible role of 17,20αP during oocyte final maturation requires further examination.     

镉对鲤血清促性腺激素和生长激素的影响

采用ＣｄＣｌ２浸没法处理鲤，研究镉对鲤血清促性腺激素（ＧｔＨ）和生长激素（ＧＨ）水平的影响，结果表明，较镐浓度镉（ＣｄＣｌ２９ｍｇ／Ｌ）处理引起血清ＧｔＨ水平降低，ＧＨ水平升高，提高水中钙浓度，能使由镉引起的ＧｔＨ水平的降低恢得取正常水平，并对镉引起的ＧＨ水平升高起延迟作用，镉处理使鲤对ＬＨＲＨ－Ａ刺激ＧｔＨ分泌的反应性下降，提高水中钙浓度也不能恢复正常，实验结果提示，镉引起的ＧｔＨ和ＧＨ的变化可