灭幼脲Ⅲ号与除虫脲在实验室鱼水系统中归趋的比较研究

本研究选择当年草鱼为试验对象,放入水族箱内,分为试验组和对照组。每组两个平行样,灭幼脲Ⅲ号(CCU)和除虫脲(DFB)的暴露浓度分别都为5和50μg/L。未放鱼的水族箱作为对照。实验开始后,分别在0,6,24,48,72和120h后取样分析两种母体化合物浓度。CCU和DFB在水中的溶解度,用标准柱提取法测定为0.21mg/L和0.23mg/L(温度19℃±1℃),试验鱼体內类脂含量用1987年OECD法测定为1％(湿重计),水样与鱼样中的DFB与CCU,分别在样品纯化步骤后用高效液相色谱法进行测定。本实验结果表明:无论对照箱还是养鱼箱中,同DFB相比,水中CCU都消失得更快,同时它在鱼体内的积累值较低(BCF<100),说明CCU比DFB具有更强的生物可降解性。两种化学品在鱼体内的积累过程与水中暴露浓度有关。     

四氯乙烯和镉对草鱼的单一与联合毒性效应

以静水生物测试法研究了四氯乙烯和重金属镉对草鱼的单一与联合毒性,同时采用Marking相加指数法对二者的联合毒性进行了评价.单一毒性试验表明:四氯乙烯对草鱼24、48、72和96 h的半数致死浓度(LC₅₀)分别为49.12、41.68、36.37和34.30 mg·L^-1;镉对草鱼24、48、72和96h的LC₅₀分别为45.58、34.81、28.63和24.05mg·L^-1;二者对草鱼均有毒性,而且为高毒,镉的毒性大于四氯乙烯.联合毒性试验表明:二者毒性比为1∶1,暴露时间为24、48、72和96 h时四氯乙烯和镉的LC₅₀分别为24.63、12.54、9.88和7.08 mg·L^-1以及17.11、8.71、6.87和4.92 mg·L^-1,相加指数AI(additive index)分别为0.14、0.81、0.95和1.43,联合作用结果为协同效应,并且随着时间的增加,协同作用增强;二者浓度比为1∶1, 暴露时间为24、48 、72和96 h时四氯乙烯和镉的LC₅₀分别为17.00、11.18、10.61和9.19 mg·L^-1,AI分别为0.39、0.70、0.51和0.54,联合作用为协同作用.     

玉屏风多糖合生元对热应激草鱼免疫性能的影响

为研究不同浓度的玉屏风多糖合生元对热应激草鱼免疫性能的影响, 选360尾健康草鱼Ctenopharyngodon idella (20±2) g, 随机分为4组。其中Ⅰ组为对照组, 试验组(Ⅱ、Ⅲ和Ⅳ组)在饲喂基础日粮的基础上分别添加0.08%、0.12%和0.20%玉屏风多糖, 并在饲喂前按复合益生菌体积(mL)/饲料重(g)为 1%的剂量添加复合益生菌拌料, 整个试验期为21d。在饲养试验结束后, 对草鱼进行热应激48h处理, 在0h、6h、24h和48h的4个时间点采集样品并测定血清、肝脏生化指标和肝脏相关基因表达量。结果表明: Ⅲ组草鱼血清中谷丙转氨酶(ALT)和谷草转氨酶(AST)在各时间点均显著低于对照组(P<0.05); 同时, 草鱼肝脏中超氧化物歧化酶(SOD)、总抗氧化能力(T-AOC)、谷胱甘肽过氧化物酶(GSH-Px)和谷胱甘肽还原酶(GR)在各时间点均不同程度高于对照组, 其中SOD在0h和24h(P<0.05), GR在0h和 48h均显著高于对照组(P<0.05), 而MDA均低于对照组, 其中在24h和48h显著低于对照组(P<0.05); 草鱼肝脏中Nrf2、Cu/Zn-SOD、GSH-Px、GR、HSP70和MHC Ⅱ相对表达量在各时间点均不同程度高于对照组, 其中GR和MHC II相对表达量在各时间点均显著高于对照组(P<0.05), 而Keap1和IL-8在各时间点均在不同程度低于对照组, 其中Keap1在各时间点均显著低于对照组(P<0.05), IL-8均差异不显著(P>0.05)。总之, 随着热应激时间的延长, 草鱼的血清和肝脏生化指标、肝脏相关基因表达量均产生了不同程度的变化, 说明热应激后草鱼产生了强烈的应激反应。在饲料中添加0.12%的玉屏风多糖合生元最有利于提高草鱼的抗氧化应激能力, 有效缓解肝脏氧化损伤, 具有较好的免疫调节作用。     

五氯酚对斑马鱼胚胎的毒性效应研究

采用斑马鱼胚胎发育技术,对环境激素类物质五氯酚的毒性进行测定,结果表明,五氯酚(PCP)对胚胎的特定作用时间段是卵产出至发育6 h之内;PCP对胚胎发育有明显的抑制作用,会造成胚胎发育的畸形或死亡,不同时间染毒产生的可观察毒理学终点各异;随着PCP对发育48 h斑马鱼胚胎作用时间的减短,其致死效应敏感性降低,其中0 hpf组的LC0值最小,为70.8μg·L^-1,24hpf组LC0值最大,为831.8μg·L^-1;斑马鱼胚胎对孵化后0时染毒的PCP最为敏感,PCP对胚胎产生急性毒性效应的敏感指标:心胞囊肿、血液循环障碍、无心律＞孵化率降低＞停滞发育作用;斑马鱼胚胎最敏感的指标为48 h血液循环障碍和48 h半致死效应.     

敌百虫对中国花鲈的毒性效应

为了给中国花鲈(Lateolabrax maculatus)病害防治提供根据,研究了不同浓度的敌百虫对中国花鲈鱼苗和幼鱼的急性毒性,检测了幼鱼肝脏的谷胱甘肽(GSH)、丙二醛(MDA)含量及超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷丙转氨酶(GPT)、谷草转氨酶(GOT)和乙酰胆碱酯酶(AChE)活力,测定了暴露在不同浓度敌百虫中幼鱼的红细胞总核异率和微核率.结果表明:敌百虫对中国花鲈鱼苗24、48、72和96h的半致死浓度分别为7.76、3.02、1.74和1.58 mg·L-1,安全浓度为0.14 mg·L-1;对幼鱼24、48、72和96 h的半致死浓度分别为10.96、6.15、3.98和2.89 mg·L-1,安全浓度为0.58 mg·L-1.濒死的中国花鲈肝脏组织有溶解性坏死灶.幼鱼在不同浓度的敌百虫中暴露48 h,随着敌百虫浓度的升高GSH含量先下降后恢复再下降;MDA含量先上升后恢复;各组SOD活力与对照组比较无显著的差异;CAT活力先下降后恢复;GPT活力维持在低水平;GOT活力在1.35和2.70 mg·L-1实验组下降;AChE活力先下降后略回升.GSH和MDA含量以及CAT、GPT、GOT和AChE活力对敌百虫胁迫敏感而SOD活力则不敏感;幼鱼在敌百虫浓度为1.45mg·L-1的实验组中暴露96 h,其红细胞的总核异率与对照组比较显著升高(P＜0.05);在0.73 mg·L-1实验组中暴露96 h,红细胞的微核率与对照组比较显著升高(P＜0.05).     

亚致死剂量高效氯氰菊酯对蚯蚓体内生化指标的影响

采用接触滤纸法, 测定蚯蚓经亚致死剂量高效氯氰菊酯染毒24 h、48 h 和72 h 后其体内蛋白含量、超氧化物歧化酶(SOD)活力、过氧化氢酶(CAT)活力和丙二醛(MDA)含量的变化, 探讨不同染毒时间和暴露浓度对蚯蚓的毒性效应。结果表明, 在高效氯氰菊酯亚致死剂量下, 染毒24 h 时蛋白含量在50 mg·L^-1 达到最大值, 而染毒72 h 时蛋白含量在5 mg·L^-1 达到最大值, 表现为短时间内促进蛋白合成而长时间下抑制。高效氯氰菊酯对SOD 和CAT 活力的诱导作用明显, SOD 活力在低浓度激活, 高浓度抑制; CAT 活力呈现出先升高后降低的趋势; MDA 含量诱导作用不明显,短时间内随暴露浓度升高而升高, 但随染毒时间延长趋于正常水平。因此SOD 和CAT 可作为蚯蚓受到氧化胁迫的指示指标, 而MDA 无法作为指示指标。此外, 各生化指标对高效氯氰菊酯毒性效应的敏感性存在差异, SOD 活力影响最大, CAT 次之, MDA 最小。     

取代苯化合物生物富集系数的估算

为了用气相色谱保留时间(t_R)估算取代苯类化合物在鱼体内的生物富集系数(BCF),测定了草鱼对氯苯、邻氯甲苯、对二氯苯、间二氯苯、邻二氯苯、硝基苯、对氯苯胺、对硝基氯苯、4-氯联苯9种取代苯类化合物的BCF和不同取代苯化合物在6种不同极性固定相上的t_R,建立了t_R与BCF的一元回归方程及t_R一阶分子连接性指数(¹X^V)与BCF的二元回归估算方程.统计检验表明,两类估算方程在不同固定相上的平均相关系数分别为0.952和0.965,均呈极显著相关(P＜0.01),可用于估算取代苯类化合物在草鱼体内的BCF,估算结果不受固定相极性的影响.     

胆固醇结石患者肝脏LCRα基因的蛋白差异表达及意义

目的:研究胆囊胆固醇结石患者肝脏的核受体基因:肝脏x受体a(liver Xreceptor a,LXRα)、法尼醇受体(famesoid Xreceptor,FXR)、人类固醇异生物受体(steroid xenobiotic receptor,SXR)及肝受体同类物1(liver receptor homolog-1,LRH-1)的蛋白表达,探讨胆固醇结石病的发病机理.方法:23例胆囊胆固醇结石患者(胆石组),12例无胆石症的胆囊息肉患者为对照(对照组).测定胆石胆固醇成分和胆汁脂类成分(胆固醇、磷脂和胆汁酸),并计算胆汁总脂和胆汁胆固醇饱和指数.Western-Blot法测定肝脏LRH-1、FXR、SXR及LXRa基因的蛋白表达量.结果:胆石组胆汁呈胆固醇过饱和;胆汁胆固醇摩尔百分比浓度较对照组升高,P<0.01;胆汁总脂较对照组明显下降,P<0.05;胆汁中胆汁酸和磷脂成分2组比较差异均无统计学意义(P>0.05).胆石组LRH-1蛋白表达高于对照组(0.88±0.05vs 0.69±0.03),P<0.05,LXRa、FXR和SXR表达2组差异无统计学意义(P>0.05).结论:人类肝脏LRH-1的蛋白表达增高与胆囊胆固醇结石形成有关.     

微生态制剂对肝内胆汁淤积性肝损伤兔模型胆汁和血清TNF-a、IL-6的影响

目的:通过研究异硫氰酸萘酯(ANIT))所致的急性肝内胆汁淤积性肝损伤兔模型血清和胆汁中TNF-a、IL-6的代谢变化,来进一步探讨微生态制剂-乐托尔对该模型的作用机制.方法:按200 mg/kg一次性灌服ANIT来诱发新生白兔急性肝内胆汁淤积性肝损伤病变,分别检测中毒后24、48 h及用乐托尔散剂治疗后血清和胆汁中总胆红素(TB)、丙氨酸转氨酶(ALT)、肿瘤坏死因子-a(TNF-a)、白细胞介素-6(IL-6)的值.结果:在乐托尔组中发现,灌服ANIT 48 h后血清和胆汁中的TNF-a、IL-6、ALT、TB值均较中毒48 h组明显降低(P值均小于0.05);各组胆汁中TNF-a、IL-6的浓度均明显高于血清中浓度.结论:微生态制剂能够改善肝功能,增加胆汁的排泄量,使炎性细胞因子的产生量显著减少.     

菲对摇蚊幼虫的急、慢性毒性效应

依照OECD 235和218指导准则,选用溪流摇蚊(Chironomus riparius)幼虫为受试生物,以存活率、羽化时间为评价指标,通过暴露于加标水中(48 h)和加标沉积物中(28 d)两种方式研究菲对摇蚊幼虫的急、慢性毒性。结果表明:对于急性毒性实验,在加标水中菲对摇蚊幼虫的LC50为1.1 mg·L~(-1)(24 h)和337.2μg·L~(-1)(48 h)。对于慢性毒性实验,在加标沉积物中菲对摇蚊幼虫的28 d-LC50为24.9 mg·kg~(-1);孔隙水中菲对摇蚊幼虫的28 dLC50为10.4μg·L~(-1);另外,随沉积物中菲浓度增加,幼虫的平均羽化时间提前,当沉积物中菲浓度达到5 mg·kg~(-1)及以上时,受试幼虫的平均羽化时间比对照组及低浓度组显著提前(P0.05),在菲浓度为10 mg·kg~(-1)时比对照组提前了3.8 d,而提前时间在大于5mg·kg~(-1)的各浓度处理之间无显著差异(3.7~4.2 d)。     

Induced Gynogenesis in Grass Carp (Ctenopharyngodon idellus) Using Irradiated Sperm of Allotetraploid Hybrids

Grass carp (Ctenopharyngodon idellus) eggs were activated by UV-irradiated diploid sperm of allotetraploid hybrids derived from red crucian carp (♀)?×?common carp (♂) and then duplicated by cold shock in 4-6°C water for 10-12 min. Different cold shock initiation times resulted in two types of diploid gynogenetic grass carp: meiotic gynogenetic (meiG) and mitotic gynogenetic (mitG). Over a 5-year period, a total of 17,170 meiG and 1,080 mitG fry were produced and 6,862 meiG and 372 mitG grass carp survived. The gynogenetic fish were confirmed by morphological characteristics, chromosome examination, and microsatellite DNA analysis. The morphological traits of the gynogenetic grass carp were similar to those of wild diploid grass carp. Normal gynogenetic fish were identified as diploid with 48 chromosomes by chromosomal metaphases examination, while nonviable abnormal embryos were detected as haploid with 24 chromosomes. Microsatellite DNA analysis indicated that after one generation of gynogenesis, the genetic purity of meiG and mitG grass carp was significantly increased over that of wild grass carp. In addition, both meiG and mitG grass carp groups were 100% female, and 88% of these showed normal ovary development. Thus, the sex determination mechanism in female grass carp was homogamety. The ability to establish pure all-female groups of meiG and mitG grass carp should be a valuable contribution to both fish genetics and grass carp breeding.     

草鱼鳃介导草鱼呼肠孤病毒免疫应答

采用草鱼呼肠孤病毒腹腔注射草鱼, 通过定量RT-PCR检测了12个抗病毒免疫相关基因在鳃中不同时间点的表达模式, 以了解鳃对内源性病毒的免疫应答。模式识别受体基因CiTLR3、CiTLR7、CiTLR22、CiRIG-I、CiMDA5、CiLGP2、CiNOD1和CiNOD2, 以及干扰素基因CiIFN-I的表达在注射病毒后12h、24h、48h及72h基本都上调。IgM基因的表达仅在72h上调。接头分子CiMyD88和CiIPS-1基因的表达在早期下调（6h）, 然后逐渐上升。为了证实病毒感染的可靠性, 通过RT-PCR检测了病毒VP4基因。结果表明草鱼鳃在抗病毒免疫方面发挥着重要作用。       

池塘循环水养殖对混养草鱼食用品质的影响

为探究养殖模式对草鱼(Ctenopharyngodon idellus)鱼肉品质的影响,实验对两种养殖模式下(传统池塘养殖和池塘循环水养殖)草鱼的食用品质进行了差异比较。结果表明,循环水养殖草鱼肌肉白度和弹性高于传统池塘养殖的草鱼,且其不饱和脂肪酸和必需氨基酸含量均显著高于传统池塘养殖;其中, n-3和n-6系列不饱和脂肪酸含量显著高于传统池塘养殖草鱼,是其1.2倍,使得草鱼的肌肉品质和营养品质更佳。此外,循环水养殖可降低草鱼中带有土腥味和青草味的己醛、庚醛和1-辛烯-3-醇等物质的含量,且鲜味核苷酸(IMP)含量高于传统池塘养殖草鱼,使得草鱼的风味更鲜美、浓厚。研究表明,循环水养殖草鱼的肌肉品质和营养特性均优于传统池塘养殖。     

Accumulation and metabolism of 2,4,6-trichlorophenyl-4'-aminophenyl ether by carp

1. Accumulation of 2,4,6-trichlorophenyl-4'-aminophenyl ether (CNP-amino) in carp was investigated for 14 days. CNP-amino in carp could be divided into free CNP-amino [CNP-amino(I)] and bound CNP-amino [CNP-amino(II)]. 2. The bioconcentration factors (BCF) of CNP-amino(I + II) in carp were 90 +/- 38 (mean +/- SD, n = 3) for muscle, 402 for liver, 501 for kidney and 5368 for gallbladder after 14 days exposure. 3. 2,4,6-Trichlorophenyl-4'-acetamide phenyl ether (CNP-acetamide) was detected as metabolites of CNP-amino in muscle and viscera of carp.     

Bioconcentration and excretion of diazinon, IBP, malathion and fenitrothion by carp

1. Bioconcentration and excretion of diazinon, IBP, malathion and fenitrothion were studied for carp (Cyprinus carpio L.). 2. The concentrations of these pesticides in muscle and viscera of the carp reached plateaus in 12-48 hr exposure. 3. The average values of bioconcentration factors (BCF) for diazinon were 20.9 in muscle, 60.0 in liver, 111.1 in kidney and 32.2 in gallbladder over the 168 hr exposure period. Similarly, those values were 4.3-26.7 for IBP, 2.7-17.3 for malathion, and 36.0-157.1 for fenitrothion. 4. The excretion rate constants of malathion (hr-1) were 0.13 for muscle, 0.12 for liver, 0.08 for kidney and 0.06 for gallbladder. Those of diazinon, IBP and fenitrothion (g.ng-1.hr-1) were 0.002-0.024 for muscle, 0.001-0.020 for liver, 0.0004-0.004 for kidney and 0.002-0.023 for gallbladder, respectively.     

Computerized analysis of motility, motility patterns and motility parameters of spermatozoa of carp following short-term storage of semen

A computer-aided semen analysis system was used to assess the % motile cells following storage of carp semen in 11 different buffers at 2, 5 or 22° C. BWW and TLP were the most suitable storage buffers because carp semen stored at 5° C in these buffers following activation showed no significant decrease in % motile spermatozoa up to 24 h. But, in most of the other buffers (Fish Ringer, Cytomix, Cortland, FRT, Mannitol, FPS, NAS and TSM) the motility potential was lost by 2 h. Storage was best at pH 6–9 and at 5° C. Carp spermatozoa exhibit three distinct motility patterns, namely 'linear', 'circular' and 'haphazard', the proportion of spermatozoa with a particular motility pattern depending on storage buffer and time. All spermatozoa with a linear trajectory had high VSL, STR and LIN; those moving in circles had low VSL, STR, LIN and BCF and those with a haphazard trajectory were distinct in that they had the highest ALH and their VSL, STR, LIN and BCF were higher than the circular moving spermatozoa and lower than the spermatozoa exhibiting linear trajectory. The study also demonstrates a pronounced time-dependent decrease in VCL, VAP, VSL and ALH of carp spermatozoa following activation with water or low osmolality solutions. This study provides for the first time data related to seven motility parameters of carp spermatozoa and demonstrates how these parameter values could be used to evaluate quality of carp milt following storage in different buffers. It confirms that carp spermatozoa exhibit linear or circular trajectories and provides evidence for a third type of trajectory described as haphazard. All three motility patterns could be discriminated objectively on the seven motility parameters.     

Structural conservation and food habit-related liver expression of uncoupling protein 2 gene in five major Chinese carps

The full-length cDNA of grass carp (Ctenopharyngodon idellus) and silver carp (Hypophthalmichthys molitrix) uncoupling protein 2 (UCP2) was obtained from liver. The grass carp UCP2 cDNA was determined to be 1152 bp in length with an open reading frame that encodes 310 amino acids. Five introns (Intron 3, 4, 5, 6 and 7) in the translated region, and partial sequence of Intron 2 in the untranslated region of grass carp UCP2 gene were also obtained. Gene structure comparison between grass carp and mammalian (human and mouse) UCP2 gene shows that, the UCP2 gene structure of grass carp is much similar to that of human and mouse. Partial UCP2 cDNA sequences of bighead carp (Aristichthys nobilis) and mud carp (Cirrhinus molitorella), were further determined. Together with the common carp (Cyprinus carpio) UCP2 sequence from GenBank (AJ243486), multiple alignment result shows that the nucleotide and amino acid sequences of the UCP2 gene, were highly conserved among the five major Chinese carps that belong to four subfamilies. Using beta-actin as control, the ratio UCP2/beta-actin mRNA (%) was determined to be 149.4 +/- 15.6 (common carp), 127.4 +/- 22.1(mud carp), 96.7 +/- 12.7 (silver carp), 94.1 +/- 26.8 (bighead carp) and 63.7 +/- 16.2 (grass carp). The relative liver UCP2 expression of the five major Chinese carps, shows a close relationship with their food habit: benthos and detritus-eating fish (common carp and mud carp) > planktivorious fish (silver carp and bighead carp) > herbivorous fish (grass carp). We suggest that liver UCP2 might be important for Chinese carps to detoxify cyanotoxins and bacteria in debris and plankton food.     

基于Miseq测序技术分析黄颡鱼不同养殖模式下池塘微生物群落结构多样性

利用Miseq测序技术分析了黄颡鱼Pelteobagrus fulvidraco (Richardson)三种养殖模式[(黄颡鱼单养、黄颡鱼混养草鱼(Ctenopharyngodon idellus, Valenciennes)和黄颡鱼混养草鱼、鳙(Aristichthys nobilis, Richardson)、鲢(Hypophthalmichthys molitrix, Valenciennes)]下养殖水体及底泥微生物群落结构多样性。结果显示, 12个样本共获得了655100条合格16S rDNA序列, 每个样本获得的有效序列数目为41256—77508, 可归纳为608—3686个分类操作单元。α多样性指数结果表明, 在三种养殖模式下末期水体的微生物群落丰度和多样性均高于养殖初期, 黄颡鱼混养草鱼模式下水体的群落丰度最高(Chao1指数: 2199.25, ACE指数: 2374.60), 黄颡鱼混养草鱼、鲢、鳙模式下水体微生物群落多样性最高(Shannon指数: 4.88, Simpson指数: 0.021)。样本聚类分析及PcoA主坐标分析结果表明, 黄颡鱼单养及黄颡鱼、草鱼混养模式下的水体和底泥的菌群组成及群落结构相似性较高, 黄颡鱼、草鱼、鲢、鳙混养模式下样本单独形成一类。从门水平的比较分析发现, 所有样本检测到细菌有8门, 其中变形菌门(Proteobacteria)的物种丰度在所有样本中占绝对优势地位。放线菌门(Actinobacteria)在养殖末期升高成为水体中占比第二的优势门, 尤其是黄颡鱼混养草鱼模式下(37.6%)。黄颡鱼在单养模式下, 养殖末期水体拟杆菌门(Bacteroidetes)细菌含量均明显增加(1.95%到10.98%)。从属水平的比较分析发现, 各样本之间的优势属组成有较大差异, 其中黄颡鱼在单养模式下水体样本中气单胞菌属(Aeromonas)在养殖末期丰度占优势(5.81%)。从微生物群落结构和多样性的角度考虑, 黄颡鱼、草鱼、鲢、鳙混养是一种值得推荐的养殖模式。     

草鱼Noxa基因克隆及其应答GCRV入侵的表达响应

研究通过获得草鱼Noxa基因(Cinoxa)全长cDNA, 进行序列分析和进化树构建, 使用定量PCR (qPCR)的方法研究其在GCRV刺激下的表达模式。研究发现, 草鱼Noxa基因的蛋白序列与斑马鱼Noxa基因具有高度相似性。通过分析草鱼和斑马鱼的Noxa基因的蛋白三维结构(3D)模型, 研究发现两者具有高度一致的蛋白三维结构模式, 该模型与高等哺乳类中的Bcl-2家族蛋白中C端结构域同源。对Cinoxa在GCRV刺激下的表达模式的研究表明, Cinoxa在GCRV感染后中肾、脾脏和头肾中表达发生显著性变化, 攻毒后24h和120h出现显著上调表达。研究表明草鱼Noxa为斑马鱼Noxa的同源基因, 并且参与了草鱼对GCRV入侵的应答反应, 为深入研究鱼类Noxa应答病毒入侵的功能和转录调控机制奠定了基础。