酶联免疫(ELISA)法测定水产品中呋喃唑酮代谢物AOZ的残留

采用MaxsignalTM AOZ快速检测试剂盒检测水产品中呋喃唑酮代谢物。通过对精密度、回收率和检测限等项目的测定,验证了方法的可行性。呋喃唑酮代谢物标准液在0．05ng／mL～4．5ng／mL范围内具有良好的线性,微孔板孔间变异系数为3．2％～7．7％。样品批内变异系数为5．2％～8．4％,批间变异系数为6．3％～8．9％,平均回收率为94．5％～98．4％。试剂盒的理论检测下限为0．05μg／Kg。该方法灵敏度高,重复性好,适用于水产品中呋喃唑酮代谢物残留的检测。     

柱后衍生高效液相色谱法测定水产品中河豚毒素含量

采用柱后衍生高效液相色谱法检测水产品中的河豚毒素含量，选用河豚Takifugu）和织纹螺（Nassarius）为主要研究对象，样品先由0.1%乙酸提取，经过C18固相萃取柱净化。以庚烷磺酸钠为离子对试剂，应用反相离子对液相色谱分离河豚毒素，采用在线柱后衍生系统，在110 ℃高温和4 mol·L^-1 NaOH强碱条件下，将河豚毒素衍生化成具有荧光特性的C9碱，荧光检测器定量检测。河豚毒素在5～1 600 ng范围内呈良好的线性相关，相关系数r=0.999 7；1、2、8 μg·g^-1，3个浓度水平添加样，日内和日间的回收率为80.9%～91.2%、相对标准偏差为1.93%～5.57%；方法定量限为1 μg·g^-1；从添加样色谱图上看，河豚毒素目标峰附近没有干扰峰，定性准确性好。采用柱后衍生高效液相色谱法与小鼠生物法检测同一阳性河豚样，检测结果一致性好。实验结果表明，柱后衍生高效液相色谱法适用于水产品中河豚毒素的检测。     

肌注和口服氟苯尼考在中华鳖体内残留分析及药代动力学

研究不同给药条件下，氟苯尼考在中华鳖体内的残留及药代动力学特征。健康中华鳖160只，随机分为2组，按30 mg·kg^-1剂量分别单次肌注或口服氟苯尼考，高效液相色谱法测定中华鳖血浆和肌肉药物残留浓度，利用3P87药代动力学软件分析数据。肌注和口服时均符合一室开放模型；肌注给药的动力学方程C=16.72(e^-0.15t-e^0.52t) ，主要药代动力学参数：AUC=76.45 μg·mL^-1·h^-1，吸收半衰期（T_1/2Ka）=1.31 h，半衰期(T_1/2Ke)=4.48 h，最高血药浓度C_max=7.09 μg·L^-1；口服给药的动力学 方程为C=39.99(e^-0.19t-e^0.4t)，主要药代动力学参数：AUC=109.42 μg·mL^-1·h^-1，吸收半衰期（T_1/2Ka）=1.73 h，半衰期(T_1/2Ke)=3.63 h，最高血药浓度C_max=10.64 〗μg·L^-1。实验结果表明，氟苯尼考口服情况下，在中华鳖体内吸收快，血药浓度高，维持时间长，生物利用度高；药物在肌肉中消除缓慢。     

达氟沙星在健康和鳗弧菌感染牙鲆体内的药物代谢动力学比较

牙鲆随机分为3组，单剂量静注（健康）和口灌（健康对照和鳗弧菌感染）达氟沙星，进行健康和鳗弧菌感染牙鲆比较药动学和生物利用度的研究。药物浓度用高效液相色谱法测定，数据采用MCP-KP自动化药动学分析程序进行分析。结果表明，健康牙鲆静脉注射达氟沙星(5 mg·kg^-1)后，血药经时过程符合无吸收一室开放模型。口灌给药(10 mg·kg^-1)健康及感染牙鲆的血药浓度与时间关系均符合一级吸收一室开放模型；肝脏和肾脏中药物浓度与时间关系均符合一级吸收二室开放模型。与健康牙鲆药动学参数相比较，达氟沙星在鳗弧菌感染牙鲆中的药时曲线下面积由256.07 mg·h^{-1·L-1降为209.18 mg·h-1·L-1，最大血药浓度由5.699 μg·mL-1降低为2.932 μg·mL-1，消除半衰期由27.758 h延长为46.195 h，生物利用度由71.21%降低为58.17%。}     

达氟沙星对史氏鲟红细胞抗氧化功能及微核形成的影响

2004年5月至7月间，用20 mg·kg^-1、50 mg·kg^-1和100 mg·kg^-1剂量对史氏鲟经口灌服达氟沙星20 d，分别于第5、10、15 和20天定红细胞SOD、CAT、Na⁺、K⁺ATP酶、GST、GSHPX等抗氧化酶活性及丙二醛（MDA）含量，以评价达氟沙星对史氏鲟抗氧化防御功能的影响，同时进行红细胞微核率和总核异常率的测定。结果表明实验第5天各实验组红细胞SOD活性均显著高于对照组；各实验组红细胞中CAT的活性明显高于对照组；红细胞Na⁺、K⁺ATP酶活性没有明显的改变。实验第20天时100 mg·kg^-1剂量组GSHPX活性明显高于对照组及其他剂量组（P<0.05）；GST活性及MDA含量没有明显的影响。达氟沙星在实验剂量内对史氏鲟红细胞核微核率没有明显的影响，但核异常率呈明显升高。     

海湾扇贝对海水中镉的富集规律研究

本文利用原子吸收分光光度法研究镉暴露实验中海湾扇贝不同器官组织富集、排放镉的过程及影响因素。水体中镉通过被动扩散进入海湾扇贝鳃，根据吸附等温线并结合直线拟合结果得海湾扇贝鳃对镉的单层饱和结合量为2.12 μmol·g^-1 干重。鳃对镉的富集量和富集速率与水相中镉浓度成正相关关系，最高浓度组（2 μmol·L^-）的镉富集量和富集速率分别为(1.5±0.52) μmol·g^-1 干重和(0.375±0.13) μmol·（g·d）^-1。随着养殖时间的延长，海湾扇贝鳃、内脏和肌肉镉富集量逐渐增加，14 d（336 h）后，内脏和鳃中镉富集达到平衡，但肌肉中镉富集是否达到平衡，尚需进一步研究。盐度对海湾扇贝鳃、内脏和肌肉镉富集速率无显著影响（P>0.05）。富集在海湾扇贝体内的镉不断被排出，其中鳃的排放率最大，达到了38.3％，其次是内脏、肌肉。     

饲料中添加硫酸锌对奥尼罗非鱼幼鱼生长合机体抗氧化功能的影响

试验以硫酸锌为锌源,评价不同锌水平对奥尼罗非鱼幼鱼（Oreochromis aureus♂×Oreochromis niloticus♀）生长和抗氧化能力的影响。体质量为(4.13±0.32)g罗非鱼随机分配在18个水族箱中,每箱20尾,每3个箱为1个处理组,分别以添加锌为0、20、40、80、160和320 mg•kg^-1的6种饲料投喂,日投喂率为鱼体重5%～9%。8周的试验结果表明：20mg•kg^-1锌饲料组的增重率和鱼体蛋白含量显著高于其它各组(P＜0.05),蛋白质效率、饲料转化率也明显高于其它各组;全鱼脂肪含量随着饲料锌水平的升高而升高,但320mg•kg^-1锌饲料组降低;20mg•kg^-1锌饲料组,脊椎骨中锌离子浓度达到最大值且显著高于0mg•kg^-1锌饲料组(P＜0.05);20mg•kg^-1与40mg•kg^-1锌饲料组的血液中红细胞数量显著高于其它各添加组(P＜0.05),20mg•kg^-1锌饲料组的血液中血比容显著高于0mg•kg^-1组(P＜0.05),与其它各组没有显著差异;肌肉中,硫代巴比妥酸反应产物(TBARS)(Thiobarbituricacid reactive substances)的量,锌为0mg•kg^-1饲料组显著高于其它各组(P＜0.05)。综上所述,饲料中锌添加量为20mg•kg-1饲料促进了罗非鱼生长,使鱼体抗氧化功能增强。     

气相色谱法测定水产品中氯霉素残留前处理方法的比较

研究了气相色谱法测定水产品肌肉组织中氯霉素残留的前处理方法.在综合分析比较现有国内外有关气相色谱法及气-质法检测动物源性食品中氯霉素残留前处理方法的基础上,对加标溶剂选择及样品提取方式、净化方法、SPE小柱活化方法对回收率的影响,硅烷化的时间等进行了比较研究.结果表明用蒸馏水作加标溶剂,回收率高,平行性较好,可代替甲醇作为加标溶剂.采用乙酸乙酯作为提取剂,用正己烷脱脂,水溶液直接过SPE小柱(小柱依次用5mL甲醇和5mL水活化)净化,硅烷化试剂衍生20～30min后测定的前处理方法,步骤少,试剂省.该方法0.1～20 μg·kg-1加标水平的回收率为67%～115%,适合水产品中氯霉素残留分析.     

盐度驯化对史氏鲟鳃Na+/K+ ATP酶活力、血清渗透压及离子浓度的影响

对史氏鲟在盐度驯化过程中鳃Na⁺/K⁺ATP酶活力、血清渗透压及血清离子（Na⁺、K⁺、Cl^-）浓度进行了检测和分析，探讨了史氏鲟驯化过程中血清渗透压调节机制。研究表明：史氏鲟在不同盐度（10、20、25）下经过驯化，鳃Na⁺/K⁺ATP酶活力显著高于对照组鳃Na⁺/K⁺ATP酶活力(P<0.05），其活力是对照组的2～2.5倍。驯化过程中，3种不同盐度阶段下鳃Na⁺/K⁺ATP酶活力首先表现为下降，随着驯化时间的延长，活力逐渐增加，最后下降并趋于平稳。血清渗透压也随盐度的增加而上升，盐度10时最高，达到(328.77±26.78) mmol·kg^-1，此后逐渐下降并稳定在290 mmol·kg^-1左右，略高于淡水中血清渗透压。不同盐度下，血清渗透压和鳃Na⁺/K⁺ATP酶活力的变化趋势相同。3种不同盐度下史氏鲟血清K⁺浓度平均值保持在3.00～3.30 mmol·L^-1之间，与对照组相比无显著差异（P>0.05）。3种盐度下血清Na⁺和Cl^-浓度变化趋势基本一致，随着盐度的增高而增高，盐度20时达到最高。盐度20以下血清Na⁺和Cl^-含量没有显著差异（P>0.05）。史氏鲟血清渗透压调节可以分为3个阶段：一是应激反应阶段，主要表现为鳃Na⁺/K⁺ATP酶活力受到抑制，陡然下降；二是主动调节阶段，鳃Na⁺/K⁺ATP酶被重新激活，且活力逐渐上升；三是适应阶段，鳃Na⁺/K⁺ ATP酶趋于平稳。     

几种化学物质对西施舌幼虫附着和变态的诱导技术

采用添加不同化学诱导物的方法，研究了肾上腺素（EPI）、氨基丁酸(GABA)、L-多巴（L-DOPA）、Ca²⁺对西施舌眼点幼虫附着和变态的诱导作用，确定了化学诱导物 最佳诱导浓度。结果表明,EPI对西施舌附着和变态的诱导效果最为显著，当EPI浓度为10^-4 mol·L^-1时，附着率及变态率分别为88.3％及92.8％，同时成活率也高达98.14％；GABA亦有较好的诱导效果，当GABA浓度为10^-5 mol·L^-1，附着率及变态率分别为86.9％及87.6％；L-DOPA能诱导西施舌眼点幼虫变态，当L-DOPA浓度为10^-6 mol·L^-1时，变态率达73.3％，附着率达84.9%. Ca²⁺最适宜浓度为20 mmol·L^-1时，附着率、变态率及成活率分别为86.1％、42.8％和87.4％。以上4种化合物最适宜浓度诱导作用时，西施舌幼虫的附着率和变态率均显著高于对照组。实验还对Ca²⁺在西施舌大规模人工育苗中的诱导作用进行探讨。     

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.