草鱼、银鲫和青鱼捕捞后的应激反应

分别评价了捕捞对草食性(草鱼)、杂食性(银鲫)和肉食性(青鱼)鲤科鱼类血液指标(血浆史质醇、葡萄糖和乳酸浓度)、肝糖原含量和两种肝脏糖酵解酶(己糖激酶和丙酮酸激酶)活性的影响。结果显示:草鱼、银鲫和青鱼捕捞后血浆史质醇、葡萄糖和乳酸浓度均显著升高;草鱼和青鱼捕捞后2 h时肝糖原含量呈下降趋势,但银鲫捕捞前、后肝糖原含量未出现显著变化;捕捞前、后青鱼血糖浓度显著高于草鱼和银鲫。银鲫肝糖原含量显著高于草鱼和青鱼,其捕捞后血浆葡萄糖和乳酸浓度增加幅度较小,这意味着捕捞后银鲫应激反应强度相对较低。草鱼和银鲫捕捞后肝脏己糖激酶和丙酮酸激酶活性未发生显著变化,青鱼捕捞后2 h己糖激酶活性显著下降,这意味着捕捞应激后血糖升高未导致草鱼、银鲫和青鱼的肝脏糖酵解酶活性增强。     

热应激对大正三色锦鲤非特异性免疫指标及HSP70基因表达的影响

为探讨锦鲤的抗逆机理,在实验室条件下研究了持续热应激对大正三色锦鲤非特异性免疫指标及HSP70基因相对表达量的影响。分别于应激前、应激后2、6、10、14、18、22、26 h进行取样测定呼吸爆发、补体蛋白3(C3)、超氧化物歧化酶(SOD)、丙二醛(MDA)以及热休克蛋白70(HSP70)基因的相对表达量。结果发现,热应激后呼吸爆发降低,在应激后2、10、18～26h降低显著(P<0.05);热应激2 h后血清中C3含量略有上升,应激6～26 h过程中与应激前相比均下降,应激14～22 h显著地下降(P<0.05);热应激下血清中SOD有上升的趋势,但差异不显著(P>0.05);热应激下,血清中MDA浓度升高,应激10～18 h上升显著(P<0.05);应激后2、26 h HSP70基因的相对表达量与应激前相比显著性地上升了5.93倍、2倍(P<0.05),应激后6～22 h HSP70基因的相对表达量与应激前水平无显著性差异(P>0.05)。结果表明,热应激影响锦鲤非特性免疫指标,降低锦鲤非特异性免疫力;HSP70的表达受热应激调节,热应激下诱导合成的HSP70对锦鲤起到一定的应激保护作用。     

高碳水化合物日粮对异育银鲫血浆皮质醇含量和HSP70基因mRNA表达的影响

为了探讨高碳水化合物饲料是否会引起异育银鲫(Carassius auratus gibelio)的应激反应,选用体质量为(35.60±1.11)g的异育银168尾,随机分成2组,一组设为对照组(NS),投喂碳水化合物水平为35％的日粮;另一组设为高碳水化合物实验组(HS),投喂碳水化合物水平为50％的日粮,每组3个重复.在控温的循环水系统中饲养35 d和70 d后,测定异育银鲫血浆皮质醇激素含量及其肝、心脏、脾、肾中HSP70基因mRNA的表达量.结果显示,高碳水化合物实验组35 d时血浆皮质醇激素与对照组相比较显著升高(P＜0.05),肝和心脏HSP70基因mRNA表达较对照组明显增强(P＜0.05); 70 d时血浆皮质醇激素含量升高,且70 d时高碳水化合物组肝、心脏、脾和肾HSP70基因mRNA表达与对照组相比全部增强.各组饲养70 d后,饥饿24 h后重新投喂的0h、6h、12h、24 h、48 h,高碳水化合物实验组(HS)皮质醇含量在6h、48 h显著高于对照组(P＜0.05),在6h、12h、48 h肝HSP70基因mRNA表达与对照组相比均增强(P＜0.05),说明异育银鲫对50％碳水化合物含量的饲料的耐受性比较差,鱼体产生了应激反应.结果说明,当异育银鲫血清皮质醇激素含量升高时,其肝中HSP70基因mRNA的表达量却降低,反之亦然.本研究旨在为水生动物营养应激及营养免疫的相关研究提供新的思路,为开辟鱼类应激的营养调节提供科学依据.     

注射葡萄糖对吉富罗非鱼血浆生化指标、胰岛素和糖酵解关键酶的影响

研究注射葡萄糖对吉富罗非鱼(Oreochromis miloticus)生化指标、胰岛素和糖酵解酶的影响。选取体质量约80 g的吉富罗非鱼150尾,随机分为2个实验组,对照组腹腔注射0.7%的无菌生理盐水,处理组按照30 mg/100 g(体质量)的剂量腹腔注射葡萄糖。结果表明:(1)吉富罗非鱼在注射葡萄糖后1 h血糖达到最高水平,而后显著下降(P<0.05),3 h后降低到正常水平。注射葡萄糖可显著提高胆固醇的含量(P<0.05),并显著降低谷草转氨酶水平(P<0.05),但对血浆蛋白、甘油三酯和谷丙转氨酶水平没有显著影响(P>0.05)。(2)肝糖原在葡萄糖注射后的6 h达到最高水平,而后显著下降(P<0.05),肌糖原没有发生变化。(3)血浆和肌肉中的胰岛素均在葡萄糖注射后的3 h达到最高水平,而后显著下降(P<0.05),而肝中的胰岛素水平没有发生变化。(4)丙酮酸激酶的活力在葡萄糖注射后6h达到最高水平(P<0.05),己糖激酶的活力在注射葡萄糖后没有发生变化。结果表明,注射葡萄糖可显著提高吉富罗非鱼血糖水平,且维持时间较长,胰岛素含量和丙酮酸激酶(PK)活力的提高均相对延迟,而己糖激酶活力的不足又限制了葡萄糖的酵解反应,从而引起血浆胆固醇升高,并伴有持续的血糖和肝糖原升高,从而产生营养胁迫的生理反应,加重鱼类对葡萄糖的代谢负担。     

热应激对大正三色锦鲤非特异性免疫指标及HSP70基因表达的影响

为探讨锦鲤的抗逆机理,在实验室条件下研究了持续热应激对大正三色锦鲤非特异性免疫指标及HSP70基因相对表达量的影响.分别于应激前、应激后2、6、10、14、18、22、26 h进行取样测定呼吸爆发、补体蛋白3(C3)、超氧化物歧化酶(SOD)、丙二醛(MDA)以及热休克蛋白70 (HSP70)基因的相对表达量.结果发现,热应激后呼吸爆发降低,在应激后2、10、18～26 h降低显著(P＜0.05);热应激2h后血清中C3含量略有上升,应激6～26 h过程中与应激前相比均下降,应激14 ～ 22 h显著地下降(P＜0.05);热应激下血清中SOD有上升的趋势,但差异不显著(P＞0.05);热应激下,血清中MDA浓度升高,应激10～18 h上升显著(P＜0.05);应激后2、26 hHSP70基因的相对表达量与应激前相比显著性地上升了5.93倍、2倍(P＜0.05),应激后6～22 hHSP70基因的相对表达量与应激前水平无显著性差异(P＞0.05).结果表明,热应激影响锦鲤非特性免疫指标,降低锦鲤非特异性免疫力;HSP70的表达受热应激调节,热应激下诱导合成的HSP70对锦鲤起到一定的应激保护作用.     

普安银鲫(Carassius auratus gibelio)卵黄囊期脂蛋白脂酶和肝脂酶基因的表达及葡萄糖、维生素C对其的影响

为了探讨普安银鲫(Carassius auratus gibelio)卵黄囊期脂蛋白脂酶(Lipoprotein lipase, LPL)和肝脂酶(Hepatic lipase, HL)的表达特点及葡萄糖和维生素C溶液分别浸泡对它们的影响,本研究采用荧光定量PCR技术检测LPL和HL基因在普安银鲫卵黄囊仔鱼发育中的表达情况及mRNA表达水平,并检测了葡萄糖、维生素C对这两种基因mRNA表达量的影响。结果显示,LPL和HL基因在普安银鲫内源营养期、混合营养期和外源营养期均有表达,且LPL和HL mRNA表达量呈上升变化。葡萄糖组LPL和HL mRNA表达量呈上升变化,维生素C组也呈上升变化。在内源营养期、混合营养期和外源营养期,葡萄糖能显著上调LPL和HL mRNA的表达量(P<0.05);在内源营养期、混合营养期和外源营养期,维生素C能显著上调LPL mRNA的表达量,而HL mRNA的表达量在混合营养期和外源营养期显著高于对照组(P<0.05)。研究表明,普安银鲫发育至混合营养期时,机体内脂质分解代谢增强。适宜水平的葡萄糖、维生素C能诱导LPL和HL mRNA的表达。     

氨氮胁迫对凡纳滨对虾肠道免疫相关指标的影响

为研究急性氨氮胁迫对凡纳滨对虾(Litopenaeus vannamei)肠道免疫功能的影响,将对虾暴露于氨氮浓度为20 mg·L^-1的海水中72 h,测定了不同时间点肠道中抗病原感染指标如酸性磷酸酶(ACP)、碱性磷酸酶(ALP)、溶菌酶(Lys)、酚氧化酶原(proPO)以及抗氧化功能指标如总抗氧化能力(T-AOC)、超氧化物歧化酶(SOD)、热休克蛋白70(HSP70)、热休克蛋白90(HSP90)等变化。结果显示,与对照组相比,氨氮胁迫后:1)ACP和ALP活性均于6 h显著升高(P<0.05),随后于48～72 h显著低于对照组(P<0.05);Lys活性于24 h显著升高至最大值(P<0.05),随后于72 h显著低于对照组(P<0.05)。2)T-AOC和SOD活性均于6 h和12 h显著高于对照组(P<0.05),随后于48 h和72 h显著降低(P<0.05)。3)HSP70基因表达水平于24 h显著升高至最大值,随后虽有降低,但仍显著高于对照组(P<0.05);HSP90和proPO基因表达水平均于12...     

半滑舌鳎响应急性高温胁迫的生理变化和相关基因表达

夏季高温会引起半滑舌鳎(Cynoglossus semilaevis)的应激,甚至造成死亡,是工厂化养殖的重要影响因素之一。为探究高温胁迫对半滑舌鳎肝脏氧化损伤及热应激相关基因的影响,本研究选取半滑舌鳎一个全同胞家系为实验对象,通过连续升温达到高温胁迫条件(35 ℃)后,分别在0、3、6、12和24 h采集肝脏组织,进行苏木精–伊红染色法(HE)和TUNEL染色并观察细胞损伤情况,测定抗氧化酶活性及丙二醛(MDA)含量并检测应激相关基因heat shock protein family A member 1A (hspa1a)、heat shock protein 90 beta family member 1 (hsp90b1)和dual-specificity phosphatase 1 (dusp1)的表达变化。结果显示,急性高温胁迫会造成半滑舌鳎肝脏组织发生明显病理变化并出现细胞凋亡;超氧化物歧化酶(SOD)活性在高温胁迫6 h时显著高于对照组(P<0.05),谷胱甘肽过氧化物酶(GPx)活性在高温胁迫12 h时显著高于对照组(P<0.05),过氧化氢酶(CAT)活性在高温胁迫0 h时显著高于对照组(P<0.05),MDA含量在高温胁迫24 h时显著高于对照组(P<0.05);热休克蛋白基因hspa1a和hsp90b1分别在高温胁迫0 h和3 h时显著上调表达,热应激相关基因dusp1在高温胁迫3 h时显著上调表达。综上所述,急性高温胁迫下,半滑舌鳎肝脏发生氧化应激,短期内机体可调动抗氧化系统加速清除活性氧,并激活热应激相关基因表达。该研究可为解析半滑舌鳎对高温胁迫的响应机制、预防夏季高温大规模死亡的发生以及开展耐高温良种的选育等提供参考。     

Gln对中华鳖免疫应激后肝脏.脾脏及肠道HSP70表达的影响

本试验通过研究谷氨酰胺/脂多糖（Gln/LPS）对中华鳖肝脏、脾脏和肠道组织热应激蛋白HSP70表达的影响,考察Gln对中华鳖免疫应激的保护作用与其诱导组织HSP70表达的关系。试验选择体重接近的健康中华鳖24只,随机分为4组,分别腹腔注射0.7%Nacl、500 ugLPS/kgBW、100 mgGln/kgBW和500 ugLPS+100 mgGln/kgBW。采用免疫组化法检测中华鳖肝脏、肠道和脾脏中HSP70的表达,放射免疫法测定血浆皮质醇（COR）含量。结果表明：腹腔注射500 μgLPS/kgBW显著增强了中华鳖肝脏和脾脏组织HSP70的表达（P<0.05）,而补充100 mgGln/kg BW抑制了中华鳖肠道、肝脏和脾脏组织HSP70的表达。推测Gln对免疫应激中华鳖的保护作用与其诱导机体组织HSP70的表达无关。     

急性氨氮胁迫对黄颡鱼组织抗氧化酶活性及HSP70和HSP90基因mRNA表达水平的影响

为了探讨急性氨氮胁迫对黄颡鱼组织中抗氧化酶活性及HSP70和HSP90基因mRNA表达水平的影响,实验随机挑选了360尾黄颡鱼[初体质量(17.25±0.05) g],分别暴露于含有0(对照)、5.70(低浓度组)、28.50(中浓度组)和57.00 (高浓度组) mg/L总氨氮浓度的水体中,进行96 h的急性胁迫实验。实验开始后,分别于0、12、24、48和96 h取样。结果显示,氨氮胁迫发生后,低、中浓度组实验鱼肝脏中超氧化物歧化酶(SOD)活性呈先升高后降低趋势,而高浓度组则持续降低;低、中、高浓度组实验鱼肝脏中丙二醛(MDA)含量在胁迫开始后显著升高;3 h时,高浓度组实验鱼肝脏中SOD活性达到最低,而MDA含量最高;24 h后,高浓度组实验鱼肝脏中过氧化氢酶(CAT)活性显著升高;低、中、高浓度组实验鱼肝脏中HSP70基因的mRNA表达量呈先降低后升高趋势,而鳃中HSP70基因表达量持续升高,但脑中HSP70基因在0 h后显著降低;氨氮胁迫3 h时,低、中、高浓度组实验鱼肝脏和脑中HSP70基因表达量显著低于对照组,而在鳃中正好相反;相比HSP70基因,高氨氮浓度组实验鱼肝脏...     

Rare serotype occurrence and PFGE genotypic diversity of Streptococcus agalactiae isolated from tilapia in China

Previously, we reported 10 PEGE types of 85 tilapia Streptococcus agalactiae(GBS), which shifted from Streptococcus iniae in China, by using PEGE method. Presently, larger and more representative tilapia GBS were isolated, for the ?rst time in China, to characterize their serotypes and genetic diversities more precisely than had done before. 168 GBS strains were distributed in ?ve provinces of China, in which Guangdong, Guangxi and Hainan were the major ones, holding36.9%(62/168), 37.5%(63/168) and 19.6%(33/168), respectively. Serotypes, Ia, Ib and III, were observed in these strains and the most predominant one was Ia(95.2%), which mainly distributed in Guangdong, Guangxi and Hainan. Ia initially occurred in 2009, it shoot up to 32.1% in 2010,but decreased to 16.1% in 2011 before went up to 45.2% in 2012. Ib sporadically occurred during2007–2011, III onlyoccurred in 2012. 14 different PFGE types, including 4 new types(N, O,P and Q), were observed, in which B, D, F and G were the predominant types, holding 83.9%(141/168) of the total GBS strains. Ia corresponded to 11 PFGE types(A–H, N–P), in which type D predominated(51%). Ib represented 3 genotypes(I, J and Q) and III harbored only 2genotypes(N and F). Type N and Fsynchronously presented in Ia and III. In summary, the genetic diversity of tilapia GBS varied by serotypes and changed with geographical locations and years.Although Iastillpredominated, new rareserotypeIII alreadyoccurred in China.     

Growth hormone (GH) and reproduction: a review

Interaction between growth and reproduction occurs in many vertebrates and is particularly obvious at certain stages of the life cycle in fish. Endocrine interactions between the gonadotropic axis and the somatotropic axis are described, the potential role of GH being emphasised. A comparative analysis of these phenomena in mammals, amphibians and fish, suggests a specific role of GH in the physiology of puberty, gametogenesis and fertility. It also shows the original contribution made by studies on the fish model in this field of investigations.     

Purification and partial characterization of GtHs (cfLH and cfFSH) from Indian walking catfish (Clarias batrachus) (L.) and development of a homologous ELISA for cfLH

Two gonadotropins (GtH; Qa and Qb) were purified by gel filtration and ion exchange chromatography from the pituitaries of Indian walking catfish (Clarias batrachus). The presence of GtH during purification was assessed by in vitro oocyte maturation and in vivo steroidogenic activity, and their identities were determined by elution profiles, molecular weight, biological activities and yield. The molecular weights of Qa and Qb were 37 and 42 kDa, respectively, and composed of distinct subunits (Qa: 20 and 14 kDa and Qb: 26 and 18 kDa). Polyclonal antibodies raised against Qa immunostained Qa, Qb and pituitary GtH cells. A competitive Qa‐ELISA was developed whose sensitivity was 6.25 ng mL^?1 (1.25 ng well^?1) with intra‐ (3.5%) and inter‐ (12.4%) assay coefficients of variation. Displacement curves parallel to the standard were obtained with plasma and pituitary extracts of catfish, Qb and carp GtHII. The assay was validated by measuring the plasma Qa levels after LHRH treatment and in relation to ovarian growth in the female catfish during different reproductive phases. Based on the results, Qa and Qb corresponded to fish LH and FSH respectively. The findings will increase the knowledge of the mechanisms controlling fish reproduction and identification of sensitive phases in fish in captivity for hormonal manipulation.     

Controlled infection of Poecilia reticulata Peters (guppy) with Tetrahymena by immersion and intraperitoneal injection

Tetrahymena is a protozoan parasite, which infects guppy, Poecilia reticulata Peters, and causes substantial economical losses in commercial farms worldwide. Studies of guppy infected by Tetrahymena require standardized infection protocols. The LD50 for Tetrahymena infection of guppies by intraperitoneal (IP) injection was calibrated, and the level obtained was 946 parasites per fish. Guppy infection with Tetrahymena by immersion, imitating the natural route of infection via the integument, was studied under normal or stress conditions. Exposure to cold and netting (CNI) and to cold only (CI) followed by immersion exposure to 10 000 Tetrahymena per mL resulted in 22.5% and 19.2% mortality, respectively, as compared to 14.2% and 10% in groups that were netted only (NI) or non‐stressed (I). Histopathology revealed that immersion infection resulted in a systemic infection. Lysozyme levels, measured 3 weeks after infection, were significantly higher in the CNI group (288 μg per mg protein) compared with CI‐, NI‐ and I‐treated groups (94.5, 64 and 62.3 μg mg^?1, respectively). There was no evident parasite immobilization activity in body homogenates, suggesting no development of acquired immunity. Re‐infection by IP injection revealed no increase in protection in any of the treatment groups, mortality range of 56.3–75%, higher than in the non‐exposed control (40.6% mortality).     

Evaluation of a Fast Method Based on the Presence of Two Restriction Sites in the Mitochondrial ND5 (mt ND5) Gene for the Identification of Scomber Species

The purpose of this work was to evaluate the suitability of a method based on the presence of two restriction sites (for Hae III and Hindf I) in the mitochondrial NADH dehydrogenase subunit 5 (mt ND5) gene to identify Scomber species. The evaluation was performed on 144 reference and market samples by sequencing of the entire 505-bp fragment of the mt ND5 gene and of a 464-bp fragment of the Kocher fragment of the cytochrome b gene (mt Cytb). Sequence analysis of any of the two fragments allows the identification of each of the four Scomber species, but S. japonicus and S. colias had the same restriction sites at the ND5 amplicon and would not have been differentiated by this analysis. Similarly, loss of the Hae III site in some S. scombrus individuals would have misidentified them as not being Scomber. All the market products were correctly labeled except one acquired in Spain labeled as originating in the Atlantic and containing S. japonicus.     

Imported ornamental fish are colonized with antibiotic‐resistant bacteria

There has been growing concern about the overuse of antibiotics in the ornamental fish industry and its possible effect on the increasing drug resistance in both commensal and pathogenic organisms in these fish. The aim of this study was to carry out an assessment of the diversity of bacteria, including pathogens, in ornamental fish species imported into North America and to assess their antibiotic resistance. Kidney samples were collected from 32 freshwater ornamental fish of various species, which arrived to an importing facility in Portland, Oregon from Colombia, Singapore and Florida. Sixty‐four unique bacterial colonies were isolated and identified by PCR using bacterial 16S primers and DNA sequencing. Multiple isolates were identified as bacteria with potential to cause disease in both fish and humans. The antibiotic resistance profile of each isolate was performed for nine different antibiotics. Among them, cefotaxime (16% resistance among isolates) was the antibiotic associated with more activity, while the least active was tetracycline (77% resistant). Knowing information about the diversity of bacteria in imported ornamental fish, as well as the resistance profiles for the bacteria will be useful in more effectively treating clinical infected fish, and also potential zoonoses in the future.     

Effect of iodophor disinfection of non‐hardened Salmo trutta eggs on their bacterial and fungus load

This study investigated the efficiency of iodophor disinfection (135 ppm active iodine for 15–30 min) of non‐hardened Salmo trutta eggs against different groups of bacteria and against fungus. Egg samples were taken from non‐disinfected and from disinfected eggs, microorganisms were cultured on specific nutrient media and their mass was measured by turbidimetric methods. Bacteria and fungus mass of non‐hardened eggs could be reduced but not eliminated by iodophor disinfection with 135 ppm active iodine for 15 min. The extent of reduction was 47–65% (Experiment 1). The efficiency of disinfection increased with disinfection time as the reduction in bacteria and fungus mass was 40–55% after 15 min and 58–74% after 30 min (Experiment 2). Disinfection efficiency of iodophor solution diluted in water (reduction 49–57%) and of iodophor solution diluted in sodium chloride solution iso‐osmolar to the oocytes (reduction 52–61%) was similar (Experiment 3). The reduction in bacteria and fungus mass was persistent as it was 39–72% lower in embryos deriving from disinfected eggs than in embryos deriving from non‐disinfected ones (Experiment 4). In conclusion, the tested disinfection method is inadequate to eliminate pathogens completely but it could positively influence immune defence of eggs and embryos.     

Nitrogenous waste excretion and accumulation of urea and ammonia inChalcalburnus tarichi (Cyprinidae), endemic to the extremely alkaline Lake Van (Eastern Turkey)

The endemic, anadromous cyprinidChalcalburnus tarichi is the only fish species known to occur in alkaline Lake Van (Eastern Anatolia, Turkey). EightC. tarichi were maintained individually in Lake Van water (17 – 19°C; pH 9.8; 153 mEq·I^–1 total alkalinity; 22 total salinity) and tank water samples analyzed for 24 h in 2 to 4 h intervals. At zero time, < 1µM ammonia was present and urea was undetectable in the tank water; at 24 h, total ammonia and urea made up 114±32 and 35±25µM, respectively. Over the experimental period, ammonia-N and urea-N excretion averaged 1041±494 and 607±169moles·kg^–1 fish·h^–1, respectively. The extent of urea excretion was highly variable between specimens. Uric acid excretion was not detectable.Urea was present at high concentrations in all tissues and plasma (25 – 35moles·g^–1·ml^–1) of freshly caughtC. tarichi; total ammonia content of the tissues was by a factor of 1.9 (liver) to 3.0 (brain) lower. High arginase activity (2.4±0.2 U·min^–1·g^–1) was detected in the liver ofC. tarichi but ornithine carbamoylphosphate transferase, a key enzyme of the ornithine-urea-cycle, was absent. Ureagenesis is likely through degradation of arginine and/or uricolysis. High glutamine synthetase activity (11±0.6 U·min^–1·g^–1) and low ammonia content in brain suggest that, like other teleosts,C. tarichi has an efficient ammonia detoxification in the brain, but in no other tissue.Nitrogenous waste excretion at alkaline pH is discussed. The ability ofC. tarichi to excrete high levels of ammonia at extremely alkaline pH is unique among teleosts studied so far. The mechanism of ammonia excretion under Lake Van conditions remains to be elucidated.