Genetic comparison between torafugu <Emphasis Type="Italic">Takifugu rubripes</Emphasis> and its closely related species karasu <Emphasis Type="Italic">Takifugu chinensis</Emphasis>

Abstract:   Sequence analyses of mitochondrial (mt) and nuclear genes were performed for genetic comparison between two Takifugu pufferfish species: torafugu T. rubripes and karasu T.  chinensis . With a sequence coverage of 20% in mtDNA, 640, 308, 344, 522 and 697 bp encoding mt 16S ribosomal RNA (rRNA), adenosine triphosphatase 6 ( ATPase 6 ), nicotinamide adenine dinucleotide dehydrogenase subunit 4 ( ND4 ), ND5 and cytochrome b (cyt b ), respectively, among 24 wild torafugu, 24 wild karasu and six hybrid-like samples, 15% of the torafugu identified by external color patterns showed nucleotide sequences consistent with karasu. Meanwhile, sequences of 60% karasu were consistent with those registered for torafugu (AJ421455). As for the hybrid-like samples, two possessed karasu-specific sequences in some base positions while torafugu-specific sequences in others. The remaining hybrid-like samples possessed torafugu-specific sequences. On the other hand, the mt control region did not show such type of consistency. Analysis of nuclear melanocortin receptor genes ( MC1R , MC4R ) among 54 samples showed 99–100% inter- and intraspecific sequence identity. Partial nuclear 18S  rRNA, complete internal transcribed spacer 1 ( ITS1 ), partial 5.8S  rRNA and ITS2 genes showed similar levels of identity, indicating a very low level of variation in their respective gene fragments between the two Takifugu species.     

Comparison between α-LNA and DHA in early developmental stages of <Emphasis Type="Italic">Takifugu obscurus</Emphasis> and <Emphasis Type="Italic">Takifugu rubripes</Emphasis>

ABSTRACT:   Rearing experiments were conducted to investigate the essential fatty acid requirements in the early developmental stages of river puffer Takifugu obscurus and tiger puffer T. rubripes using two n-3 series unsaturated fatty acids, α-linolenic acid (18:3n-3, α-LNA) and docosahexaenoic acid (22:6n-3, DHA), under salinity of 30 and 18.5–20.3°C. River and tiger puffer larvae used in this study were 15 and 14 days old after hatching, and their average body weights were 30.1 and 20.8 mg, respectively. The results on fatty acid requirements of these two species were evaluated from fish growth, survival, fatty acid composition of the fish body and activity test results. The DHA groups of both river and tiger puffer exhibited better survival and weight gain. However, there was no difference in the mean final body weights of river puffer between two dietary groups. Also, the DHA group of tiger puffer showed better results in the recovery test from anesthetic condition than that obtained in the LNA group. In an examination of the fatty acid compositions of the whole body, the LNA group containing no dietary DHA resulted in 0.5% DHA in tiger puffer and 1.1% DHA in river puffer . These results suggest that α-LNA from Artemia converted to eicosapentaenoic acid (20:5n-3, EPA) and to DHA successively by their fatty acid metabolism. Symptoms following essential fatty acid deficiency were not observed in any experimental groups. As river puffer did not represent a significant difference in the dietary effects between α-LNA and DHA treatment groups, its essential fatty acid requirement was assumed to be somewhat closer to that of the freshwater fishes in comparison with that of marine fishes, including tiger puffer.     

双酚A对暗纹东方鲀幼鱼类固醇合成酶基因表达的影响

为研究环境激素对养殖鱼类性别分化前后的内分泌干扰效应,分别将孵化后25 d(dph)的暗纹东方鲀(Takifugu obscurus)稚鱼和95 dph的幼鱼暴露于不同浓度双酚A(BPA)(0、50μg/L、200μg/L、1 000μg/L)3周和2周,用荧光定量PCR(QRT-PCR)方法检测编码性类固醇激素合成酶及相关因子的基因(STAR、CYP17a、CYP11b及CYP19a)表达。结果显示,BPA诱导了稚鱼STAR、CYP17a和CYP11b表达上调,且与浓度有正相关,而CYP19a表达在BPA 200μg/L和1 000μg/L浓度下分别暴露3周后显著下调。在幼鱼暴露组中,雌雄性腺发生不同的应答,精巢中STAR、CYP17a及CYP11b的表达出现先下调后上调,CYP19a则呈先受抑制后表达升高;在卵巢中STAR、CYP17a及CYP11b的表达先受抑制,随着时间的延长基因表达上调,CYP19a的表达受抑制。研究结果表明,BPA通过干扰性别分化初期暗纹东方鲀类固醇激素合成酶及相关因子的表达的而对性别分化产生影响;通过抑制雄性暗纹东方鲀中类固醇合成相关基因的表达而表现出内分泌干扰效应。对于雌鱼中类固醇合成相关基因的表达的影响则有波动性的变化,其作用机制仍需要进一步研究。     

暗纹东方鲀应激逃避行为及其 Mauthner 细胞的形态学观察

探讨暗纹东方鲀(Takifugu obscurus)这种具有胀气、佯死行为的鱼类是否具有快速逃跑行为,以及脑干Mauthner细胞是否是该行为的指令性神经元.研究发现,给予骤发的声音刺激(100 Hz,110 dB),暗纹东方鲀幼鱼和斑马鱼(Danio rerio)均出现快速逃跑行为.但是,与斑马鱼相比,暗纹东方鲀的快速逃跑行为发生概率低,且延迟时间长(P＜0.01).经10 μmol/L GABAA受体抑制剂药浴处理30 min后,与对照组相比,暗纹东方鲀快速逃跑行为发生概率显著增加(P＜0.05),反应延迟时间也显著降低(P＜0.01).组织学观察发现,暗纹东方鲀的Mauthner细胞形态特殊,胞体为椭圆形,细胞的长短轴比显著低于斑马鱼Mauthner细胞的长短轴比,且未见其与第八神经的直接联系.并且,在暗纹东方鲀延脑中未观察到Mid2cm、Mid3cm等神经元.因此,推测由于暗纹东方鲀Mauthner细胞欠发达,导致其快速逃跑能力较弱,故在应激逃避方面,需要进化出其他的防御方式,如胀气等,以保证种群的存活率.本研究为深入研究鱼类的应激反应的生理机制及经济鱼类的健康养殖提供了理论依据.     

菊黄东方鲀仔稚鱼的生长、发育及行为生态

在大批量苗种培育条件下,通过对菊黄东方鲀胚后发育阶段的观察、测量和摄影,研究了菊黄东方鲀仔、稚鱼的生长、发育及行为生态特点。结果显示,仔稚鱼发育分为前期仔鱼、后期仔鱼、前期稚鱼、中期稚鱼和后期稚鱼5个阶段,每个阶段由若干个时期组成,总共为17个时期。在盐度10.0～13.5、温度20.5～26.0 ℃条件下,鱼苗历时35 d 完成了从初孵仔鱼至夏花鱼种的发育。期间,鱼苗食性经历了开口和转食配合饲料2个关键时期。食物转换依次为卵黄、轮虫、卤虫幼体、中型枝角类、大型桡足类和枝角类、底栖生物和虾类幼体、配合饲料。同时,鱼苗完成了由水体中上层向中下层活动区域的转变。游泳行为经历了平游、巡游和池塘觅食巡游3个模式。鱼苗在后期仔鱼阶段特定生长率最低,为3.18%/d,中期稚鱼阶段最高,达13.31%/d。全长与日龄及积温均呈指数函数增长相关(y=ae^bx)。肛前长和体高相对于全长均呈有拐点的、可分段的异速生长相关(y=ax^b)。拐点分别出现在鱼苗16～17日龄、全长10.0 mm和16～17日龄、全长10.5 mm。在拐点前,肛前长、体高生长优于全长(b＞1)。拐点后则相反(b＜1)。根据研究结果,建议当菊黄东方鲀仔鱼4日龄开口时,采用S型褶皱臂尾轮虫,并保持轮虫密度(5～7 ind/mL);及时分池,时间在10～12日龄为好;下塘时间为20日龄左右;在鱼苗转食配合饲料后(35日龄),及时过筛、分级养殖等,提高菊黄东方鲀苗种培育存活率。     

基于径向基函数(RBF)神经网络的红鳍东方鲀体质量预测

基于表型性状预测红鳍东方鲀体重时,由于不同表型性状间的自相关、部分性状和体重之间的非线性关系以及线性回归方法自变量间的共线性,结果导致预测误差过大。为了解决这一问题,根据人工神经网络(Artificial Neural Networks, ANN)建模原理,采用径向基函数(Radial Basis Function, RBF) 模型,利用72个红鳍东方鲀样品的生长数据, 通过最近邻聚类算法, 构建了基于RBF神经网络的红鳍东方鲀体重预测模型,并采用线性回归检验法对所构建模型的可信度进行检验。结果发现,基于RBF神经网络预测模型的拟合优度为0.992,接近于1,而线性回归模型的拟合度为0.949。这表明: 基于RBF神经网络方法构建的预测模型消除了线性分析中自变量的共线性问题, 比线性回归预测模型的拟合度提高4.53%,预测精度高于线性回归方法。基于RBF神经网络体重预测模型的构建,为利用表型性状精确评估红鳍东方鲀的体重提供了一种新的方法。     

红鳍东方鲀两个群体的遗传结构及与经济性状的相关性分析

本文利用21对多态性微卫星标记对河北省红鳍东方鲀(Takifugu rubripes)良种场引进的日本群体和选育的秦皇岛群体进行了遗传结构分析.日本与秦皇岛两个群体的有效等位基因数分别为3.3697和2.8593,期望杂合度分别是0.6516和0.6073,多态信息含量分别是0.6009和0.5509.研究结果表明这两...     

不同性别沙塘鳢体重与生化组成和能值的关系

研究了体重增加时不同性别沙塘鳢鱼体生化组成和能值的变化。结果显示,雌性沙塘鳢的干物质、脂肪含量和能值随体重的增加而显著增加,蛋白质含量与体重没有显著相关性;雄性沙塘鳢的脂肪含量和能值随体重的增加而显著增加,蛋白质含量显著下降,干物质含量与体重没有显著相关性。随着体重增加,雌性沙塘鳢的脂肪、蛋白质含量和能值以及雄性沙塘鳢的蛋白质含量和能值都与各自鱼体的干物质含量呈显著的直线相关,雄性沙塘鳢的脂肪含量与干物质含量没有显著相关性。     

稻田养殖沙塘鳢对稻田水体及底泥微生物群落结构及多样性的影响

为了研究养殖沙塘鳢(Odontobutis obscurus)对稻田水体及底泥的微生物群落结构及多样性的影响,2015年于浙江海盐江南四阡现代农业公司进行了沙塘鳢的稻田养殖实验,利用DGGE(Denaturing Gradient Gel Electrophoresis)技术对养殖过程中的稻田水体及底泥中细菌的16S r DNA片段进行指纹图谱分析。DGGE条带测序分析结果显示:稻田水体及底泥共检测到包括α-变形菌亚门(Alphaproteobacteria)、β-变形菌亚门(Betaproteobacteria)、γ-变形菌亚门(Gammaproteobacteria)、δ-变形菌亚门(Deltaproteobacteria)、ε-变形菌亚门(Epsilonproteobacteria)、绿菌门(Chlorobi)、酸杆菌门(Acidobacteria)、厚壁菌门(Firmicutes)、放线菌门(Actinobacteria)、蓝藻门(Cyanobacteria)等细菌门类。多样性分析结果显示,养殖沙塘鳢的稻田底泥微生物DGGE条带数多于常规养殖稻田,养殖稻田的底泥Shannon多样性指数2.86,显著高于常规稻田的2.27,同时养殖稻田养殖沟底泥的Shannon多样性指数随养殖时间由2.56变化到2.16。PCA(Principal Component Analysis)及DGGE聚类分析结果显示,养殖稻田的水体及底泥微生物群落结构与常规稻田存在较大差异。结果表明,稻田养殖环境的微生态条件可能优于常规稻田。     

暗纹东方鲀线粒体COIII克隆及序列分析

克隆并测定了暗纹东方鲀线粒体基因组(m tDNA)含完整的细胞色素氧化酶亚基Ⅲ(CO III)及其下游70 bp的tRNAG ly序列。CO III基因编码框包含786个核苷酸,编码262个氨基酸的蛋白质。鱼类CO III的序列组成对A T核苷酸的偏倚程度比较低。通过暗纹东方鲀与GenBank中8个目12种鱼类的CO III序列同源性比较,显示暗纹东方鲀与这些鱼类的CO III基因具有较高的同源性。根据暗纹东方鲀与鲀目其他5种鱼的CO III基因序列所建立的进化树,与传统的分类地位相吻合。推定的tR-NA的二级结构具有典型的三叶草型结构。根据CO III序列构建的分子进化树分析,认为在采用线粒体基因序列进行分子进化分析时,应综合考虑物种遗传变异的特点。