孔雀石绿及其代谢物隐色孔雀石绿在鳜鱼体内组织的分布及消除规律

为研究在养殖模式下孔雀石绿及其代谢物隐色孔雀石绿在鳜鱼体内各组织的分布及消除规律,将鳜鱼以5 mg/L的孔雀石绿溶液浸泡2 h后饲养于池塘中,采用高效液相色谱-串联质谱法分析鳜鱼背肌、腹肌、内脏、腮、皮肤等5种组织中孔雀石绿及其代谢物隐色孔雀石绿的浓度水平。结果表明,孔雀石绿在鳜鱼5种组织中的浓度由高到低的顺序为:皮肤＞内脏＞鳃＞腹肌＞背肌,在皮肤组织中残留时间最长,在背肌、腹肌、内脏、腮、皮肤中分别于58、90、20、58 d和120 d时未被检出；隐色孔雀石绿在鳜鱼5种组织中浓度由高到低的顺序为:内脏＞皮肤＞背肌＞腹肌＞鳃,在皮肤组织中残留时间最长,在背肌、腹肌、内脏、腮、皮肤中分别于270、290、230、230 d和300 d时未被检出。试验期间平均水温为25.6 ℃,在此养殖条件下,孔雀石绿及其代谢物隐色孔雀石绿在鳜鱼各组织中消除至少需要7 680 ℃·d。     

加快鳗鲡体内残留孔雀石绿降解速度药物的初步研究

本试验选择已具孔雀石绿残留的鳗鱼进行研究。降解药物按试验设定的剂量拌饵投喂，用高效液相色谱法测定样品中孔雀石绿和隐性孔雀石绿的含量。结果表明：复配药物能起到加快鳗鲡体内孔雀石绿降解速度的作用；降解药物Ⅰ和降解药物Ⅱ均能加快鳗鲡体内隐色孔雀石绿的消除速率，而且效果相近；起主要降解作用的药物是腐植酸，维生素C和654—2只起辅助作用或不起作用。     

孔雀石绿浸泡几种鱼类受精卵及苗种后的降解规律研究

为研究孔雀石绿在几种鱼类受精卵、苗种体内的降解规律,用孔雀石绿溶液对受精卵和苗种进行浸泡处理,然后检测显性孔雀石绿及其代谢物隐性孔雀石绿的残留量。试验结果:用10.0 mg.L-1的孔雀石绿浸泡异育银鲫、团头鲂的受精卵10 min,至仔鱼孵出后第31、45 d,在两者中均未检出显性孔雀石绿(检出限为O.5μg/kg);至第45 d,两者中均未检出隐性孔雀石绿。用10.0 mg/L的孔雀石绿分别浸泡异育银鲫的鱼苗、鱼种10、25 min,至浸泡后第10、11周,在两者中均未检出显性孔雀石绿;至第16、38周,两者中均未检出隐性孔雀石绿。用1.0 mg/L的孔雀石绿浸泡黄颡鱼鱼种30 min,至浸泡后第12周,未检出显性孔雀石绿;但至第52周,隐性孔雀石绿仍有5.34μg/kg的残留量。结果表明,在水产动物体内,隐性孔雀石绿的降解速度要明显慢于显性孔雀石绿。     

养殖用水中孔雀石绿和结晶紫及其代谢物的测定

建立了用高效液相色谱紫外、荧光检测器同时检测养殖用水中的孔雀石绿、结晶紫及其代谢物隐色孔雀石绿和隐色结晶紫的分析方法。孔雀石绿和结晶紫用紫外检测器588 nm波长检测;隐色孔雀石绿和隐色结晶紫用荧光检测器检测,激发波长:265 nm,发射波长:360 nm。孔雀石绿和结晶紫的线性范围5.0~2500μg/L;隐色孔雀石绿和隐色结晶紫的线性范围1.0~1 000μg/L,四种物质的检出限均为0.5μg/L,回收率均在70%以上,RSD15%,此方法也可用于地下饮用水中孔雀石绿、结晶紫、隐色孔雀石绿和隐色结晶紫的检测。     

小口脂鲤对几种常用药物的敏感性试验

在水温 2 6～ 30℃下 ,敌百虫等十种常用药物对小口脂鲤夏花鱼种的安全浓度分别为 :敌百虫 0 .0 4 2 mg/ L、甲醛 18.2 0 mg/ L、硫酸铜合剂 0 .98mg/ L、孔雀石绿 0 .0 35mg/ L、高锰酸钾0 .93mg/ L、鱼虫克星 0 .0 37mg/ L、强氯精 0 .13mg/ L。呋喃唑酮、土霉素、亚甲基蓝的试验最高浓度分别为 6 4 mg/ L、6 4 mg/ L、8mg/ L,在此浓度下 ,2 4 0小时内小口脂鲤夏花鱼种全部存活。以安全浓度作为衡量标准 ,小口脂鲤夏花鱼种对上述药物的敏感性依次为 :孔雀石绿 >鱼虫克星 >敌百虫 >强氯精 >高锰酸钾 >硫酸铜合剂 >甲醛。其中孔雀石绿、敌百虫的安全浓度仅为常用浓度的 10 %～ 30 % ,故不宜使用。鱼虫克星、强氯精的安全浓度较低 ,在疾病防治中可谨慎使用。     

孔雀石绿降解菌Enterobacter sp.B-20的分离、鉴定和降解特性研究

从受孔雀石绿污染的淡水鱼养殖池塘底泥中,筛选到一株孔雀石绿(C23H25Cl N2,malachite green,MG)高效降解菌,经16S r RNA基因序列及系统进化树分析,初步鉴定为Enterobacter sp.B-20。采用硼氢化钾(KBH4)还原-高效液相色谱法对Enterobacter sp.B-20的孔雀石绿降解特性进行研究。结果显示,孔雀石绿对菌株的生长有一定抑制作用,24 h内菌株对5~20 mg·L~(-1)孔雀石绿降解率超过97%,12 h内对1~40 mg·L~(-1)孔雀石绿降解率超过82%。菌株的孔雀石绿降解能力随着p H的增加而显著提高,最适降解p H为9。Enterobacter sp.B-20具有较强的耐盐性,在10~40 g·L~(-1)的氯化钠(Na Cl)质量浓度下具有稳定的孔雀石绿降解特性。在高浓度金属离子条件下,Enterobacter sp.B-20表现出较强的耐性及稳定的孔雀石绿降解能力,0.1~1 mmol·L~(-1)铁离子(Fe3+)、0.1~0.5 mmol·L~(-1)铜离子(Cu2+)、0.1 mmol·L~(-1)锰离子(Mn2+)、0.1~1 mmol·L~(-1)铅离子(Pb2+)均能显著提高菌株的孔雀石绿降解率。结果表明,Enterobacter sp.B-20具有在复杂养殖水环境下降解孔雀石绿的应用潜力。     

阳性饵料鱼投喂模式下孔雀石绿及代谢物无色孔雀石绿在鳜体内的残留消除规律

为了解鳜(Siniperca chuatsi)食用含有孔雀石绿(malachite green,MG)的饵料鱼后,其体内MG及其代谢物无色孔雀石绿(leucomalachite green,LMG)的残留消除规律,以期为孔雀石绿的监管提供基础数据,本实验模拟了自然养殖条件,对饲养于池塘网箱中的鳜连续投喂10 d经1 mg/L MG溶液浸泡2 min后的鲮(Cirrhinus molitorella),投喂量为5%(m/m),于停药后0、6、12 h及1、3、6、10、15、20、30、40、50、70、90、120、150和180 d采集鳜肌肉样品,各采集点随机取6尾以上鳜,采用液相色谱串联质谱法测定鳜肌肉中MG和LMG的残留量。结果发现,阳性饵料鱼投喂结束后,鳜体内未检出MG,LMG残留浓度也较低,在0 h鳜肌肉中LMG的浓度为8.62μg/kg,随后缓慢降低,10 d时鳜肌肉中检测不到LMG。本研究表明,养殖过程中,阳性饵料鱼体内的MG会通过食物链传递给鳜,造成鳜体内LMG检出,建议执法部门对鳜的饵料鱼进行监控,以确保鳜的食用安全。     

呋喃唑酮降解菌株的分离鉴定及降解特性研究

从污染鱼塘底泥中筛选分离出一株能有效降解低浓度呋喃唑酮的细菌F5,经16S rDNA同源性序列分析,鉴定为铜绿假单胞菌(Pseudom onas aeruginosa)。30℃静置避光培养30 d后,该菌株对0.75、1.0和2.5 mg/L呋喃唑酮的降解率分别为58.8%、64.1%和38.2%。温度为20~25℃更有利于该菌株的降解作用,对1.0 mg/L呋喃唑酮降解率均能达85%。碳源和氮源浓度的提高均能促进该菌株的生长和对呋喃唑酮的降解。在0.5 mg/L低磷浓度下,该菌株生长受抑制但降解活性增加,对1.0 mg/L呋喃唑酮降解率达71.8%。     

湛江港区底泥悬浮物对2种海洋动物的致死效应研究

为评价湛江沿海涉海工程建设过程中引起的悬浮物质对附近海区海洋动物的影响,研究了底泥液相及不同浓度底泥悬浮物对川纹笛鲷和尖吻鲈的致死效应。试验结果表明,在8d的试验时间里,底泥水相对川纹笛鲷和尖吻鲈均无致死效应。底泥悬浮物浓度≤160mg/L时,川纹笛鲷和尖吻鲈的死亡率均处于较低水平。底泥悬浮物浓度≥2560 mg/L时,川纹笛鲷死亡率达75%,尖吻鲈死亡率达70%。通过概率单位法计算得出川纹笛鲷的8d半致死浓度为716.14 mg/L,尖吻鲈的8d半致死浓度为1105.096 mg/L。     

高效液相色谱-荧光检测法测定鳗鲡中孔雀石绿和结晶紫残留

本研究建立了高效液相色谱-荧光检测法测定鳗鲡(Anguilla japonica)中孔雀石绿和结晶紫残留量的新方法.样品中残留的孔雀石绿或结晶紫用硼氢化钾还原为其相应的代谢产物隐色孔雀石绿或隐色结晶紫,用乙腈、乙酸铵缓冲溶液提取,二氯甲烷液液萃取,PRS固相萃取柱净化,反相色谱柱分离,荧光检测器检测,外标法定量.孔雀石绿和结晶紫混合标准工作溶液质量浓度范围为1～600 μg/L时,该方法的线性关系极好,相关系数均为0.999 9.该方法定量检出限为0.4 μg/kg.当空白样品中孔雀石绿和结晶紫添加水平为0.4～100 μg/kg时,孔雀石绿回收率为70.0%～90.6%,结晶紫回收率为73.0%～87.2%,相对标准偏差均小于10%.     

Association between the interannual variation in the oceanic environment and catch rates of bigeye tuna (Thunnus obesus) in the Atlantic Ocean

The environmental processes associated with variability in the catch rates of bigeye tuna in the Atlantic Ocean are largely unexplored. This study used generalized additive models (GAMs) fitted to Taiwanese longline fishery data from 1990 to 2009 and investigated the association between environmental variables and catch rates to identify the processes influencing bigeye tuna distribution in the Atlantic Ocean. The present findings reveal that the year (temporal factor), latitude and longitude (spatial factors), and major regular longline target species of albacore catches are significant for the standardization of bigeye tuna catch rates in the Atlantic Ocean. The standardized catch rates and distribution of bigeye tuna were found to be related to environmental and climatic variation. The model selection processes showed that the selected GAMs explained 70% of the cumulative deviance in the entire Atlantic Ocean. Regarding environmental factors, the depth of the 20 degree isotherm (D20) substantially contributed to the explained deviance; other important factors were sea surface temperature (SST) and sea surface height deviation (SSHD). The potential fishing grounds were observed with SSTs of 22–28°C, a D20 shallower than 150 m and negative SSHDs in the Atlantic Ocean. The higher predicted catch rates were increased in the positive northern tropical Atlantic and negative North Atlantic Oscillation events with a higher SST and shallow D20, suggesting that climatic oscillations affect the population abundance and distribution of bigeye tuna.     

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 performance,digestive enzyme activity and immune response of Japanese sea bass,Lateolabrax japonicus fed with fructooligosaccharide

In this experiment, a feeding trial was performed to determine the effects of fructooligosaccharide (FOS) on growth performance, digestive enzyme activity and immune response of Japanese sea bass, Lateolabrax japonicus juveniles (initial weight 38.3 ± 0.5 g), and the fish were examined following feeding with six levels of FOS (0, 0.5, 1, 2, 4 and 6 g/kg) for 28 days. Significant enhancement of weight gain (WG) and specific growth rate (SGR) was found in fish fed 1 g/kg FOS incorporated diets (p < .05), while the feed conversion ratio (FCR) in the 1, 2 g/kg FOS groups reduced significantly compared with the control (p < .05). Besides, the crude lipid in the 4, 6 g/kg FOS groups increased significantly compared with the control (p < .05). On the other hand, the erepsin and lipase activities significantly elevated in intestine of fish fed 2 g/kg FOS (p < .05) and the lysozyme activity in serum of fish fed 2 g/kg FOS were significantly higher than that in the control (p < .05). Moreover, the alkaline phosphatase activities in serum of fish fed 0.5, 1, 2 g/kg FOS were significantly higher than in control (p < .05). Regression analysis showed that the relationships between dietary FOS levels and either SGR, FCR, erepsin or lysozyme activities were best expressed by regression equations, and the optimal inclusion levels are 1.37, 1.80, 3.06, 3.11, 1.93 and 1.80 g/kg for SGR, FCR, erepsin, lipase, lysozyme and total superoxide dismutase activities, respectively. Overall, this study revealed that FOS incorporated diets could beneficial for L. japonicus culture in terms of increasing the growth, digestion and immune activities. Under the present experimental condition, the optimal supplementary level of FOS in the diet of L. japonicus is 1–3 g/kg.     

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.     

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).     

The optimal dietary protein level of juvenile silver sillago Sillago sihama at three dietary lipid levels

A 5 × 3 factorial growth trial was conducted to evaluate optimal dietary protein and lipid levels (dietary protein level, DP; dietary lipid level, DL) for juvenile Sillago sihama (S. sihama) (2.0 ± 0.02 g, initial weight). Fish were fed 15 diets containing 5 DPs (350, 400, 450, 500 and 550 g/kg) and 3 DLs (60, 90 and 120 g/kg) for 8 weeks. The interaction between proteins and lipids significantly influenced the feed conversion ratio, condition factor, body composition, antioxidant indices and lipase activity (p < .05). DP 450 g/kg showed the highest average final body weight. DPs 500 and 550 g/kg significantly decreased the protein efficiency ratio (p < .05). DL 120 g/kg showed the highest percentage weight gain. The low feed conversion ratio was found in diets P45L12, P55L9 and P55L12. Diet P45L12 showed high superoxide dismutase activities. DP 450 g/kg showed the lowest average malondialdehyde content. Lipase activity was increased by increasing DP (p < .05) with a fall at DP 550 g/kg. Under the present experimental conditions, the optimal DP for S. sihama was 450 g/kg under the DL 120 g/kg.     

Involvement of gonadal steroids in final oocyte maturation of white perch (Morone americana) and white bass (M. chrysops): in vivo and in vitro studies

Plasma estradiol-17 (E₂), testosterone (T), 17,20-dihydroxy-4-pregnen-3-one (DHP) and 17,20,21-tri-hydroxy-4-pregnen-3-one (20-S) levels were measured by radioimmunoassay (RIA) in white perch (Morone americana) and white bass (M. chrysops) that were induced to undergo final oocyte maturation (FOM) with human chorionic gonadotropin (hCG). Plasma DHP levels increased in females of both species in association with oocyte germinal vesicle migration (GVM) and germinal vesicle breakdown (GVBD) and decreased thereafter. Plasma 20-S levels also increased with oocyte GVM in white bass, but were several-fold lower than DHP levels. Circulating E₂ and T levels were greatest during GVM and GVBD in both species and decreased to low levels during oocyte hydration and ovulation. Follicles from white perch and white bass which received a priming injection of hCG in vivo, produced both DHP and 20-S in vitro after exposure to hCG and their oocytes underwent GVBD. Ovarian incubates from unprimed fish of either species produced only E₂ and T and their oocytes did not complete GVBD. Oocytes from unprimed bass, but not perch, matured when follicles were exposed to hCG in vitro. Both trilostane and cycloheximide blocked in vitro production of DHP and 20-S and oocyte GVBD by white perch follices. DHP and 20-S were equipotent inducers of FOM in the GVBD bioassay. None of several other structurally-related steroids tested were effective within a physiological range of concentrations. These results indicate a role for DHP and 20-S in the control of FOM in white perch and white bass.