New Evidence to the Genetic Diversity of Trachurus japonicus in the Coastal Waters of Fujian Province Based on AFLP Makers

为研究福建近海竹荚鱼群体的遗传分化,对竹荚鱼闽东(30尾)和闽南(30尾)群体进行了扩增片段长度多态性(AflP)分析,8对选择性引物在2个群体60个个体中,共扩增出433条条带,其中多态位点286个.闽东和闽南群体的多态位点比例分别为63.28%、61.89%,nEI遗传多样性指数分别为0.171 8、0.172 2,SHAnnOn多样性指数分别为0.267 3、0.267 3.与其他鱼类对比显示,福建近海竹荚鱼群体的遗传多样性水平较高,说明其种质资源尚未遭到明显破坏;遗传分化系数(gST)和分子生物学方差分析(AMOVA)均显示竹荚鱼的遗传变异主要来源于群体内,而群体间无明显的遗传分化.基因流(nM)显示2个群体间基因交流频繁,无明显独立的遗传结构.群体的位点差异数分布和显性基因型频率分布显示2个群体有相似的群体遗传结构.结果表明,竹荚鱼闽东和闽南群体间无明显的遗传差异,可将福建近海的竹荚鱼划归同一个管理保护单元.近期扩张、较强的扩散能力和洋流可能是造成福建近海竹荚鱼群体间遗传同质性较高的原因.Given Trachurus japonicus resources declining,amplified fragment length polymorphism(AFLP)was employed to analyze the genetic diversity and variation of two populations of T.japonicus from Eastern and Southern Fujian coastal water in this study.AFLP bands were scored for presence(1)or absence(0)and transformed into 0/1binary character matrix.AMOVA and mismatch distribution analysis were performed in Arlequin,Nei genetic diversity and Shannon genetic diversity index were conducted in POPGEN,genetic distance between individuals were computed in Phyltools and the UPGMA tree was generated in MEGA.A total of 433loci ranging in size from 100bp to 1 000bp were detected from 60individuals using eight primer combinations,of which 286 were polymorphic.The proportion of polymorphic loci,the Nei genetic diversity and Shannon genetic diversity index of these two populations were 63.28%and 61.89%,0.171 8and 0.172 2,0.267 3and 0.267 3,respectively,revealing no significant difference(p>0.05)between the two populations.By comparison with that of other fishes,that the genetic diversity of T.japonicus in the coastal waters of Fujian province is above the middle level and the population genetic structure of T.japonicus had not been destroyed.Gst value,Shannon genetic diversity index and AMOVA analysis indicated that the genetic variation mainly came from individuals within populations.Nm showed that there was frequent gene flow between the two populations.The NJ tree based on genetic distance between individuals exhibited no correlation to geographic distribution.Mismatch distribution analysis and dominant gene frequency revealed that these two populations have the similar genetic structure.Based on the facts above,the results of AFLP analysis indicated that there is no significant genetic differentiation between two populations,and the recent expansion,annual migrations and larval drift in the ocean currents could be the reasons for the high homogeneity of genetic structure between two populations.The knowledge on genetic diversity and variation will be crucial to establish appropriate fishery management stocks for the species.福建省科技计划项目(2013N0005;2007I0005

Population Genetic Structure Analysis of Decapterus maruadsi from Fujian Coastal Waters

测定了福建近海蓝圆鲹(dECAPTEruS MAruAdSI TEMMInCk & SCHlEgEl)2个群体共60尾个体的线粒体dnA(MTd-nA)控制区序列,探讨了闽东和闽南群体遗传结构和遗传多样性.结果显示:获得长度为834~838bP的控制区部分序列,在所测的60个样本中,共检测到23个变异位点,定义了28个单倍型,平均单倍型多样性(H)为0.948±0.0145,核苷酸多样性(π)为0.004 99±0.002 79,核苷酸差异数(k)为4.173±2.104,提示了福建近海蓝圆鲹具有较高的遗传多样性水平.构建的单倍型邻接关系树没有明显的以地理群体为单位的家系式分支出现,单倍型网络图也未显示出单倍型和地理位置的对应关系.TAJIMA′S d和fu′S fS中性检验及核苷酸不配对分析暗示了蓝圆鲹闽南群体在63 000年前可能发生过扩张事件,而闽东群体符合中性理论进化,2个群体不同的历史动态可能是由于小冰河期海区间的气候差异和闽东群体的过度利用所致.分子方差分析(AMOVA)表明,遗传变异全部存在于群体内,2个群体间具有紧密的基因流和较低的遗传分化,群体间和群体内的kIMurA双参数遗传距离均较小.可知:福建近海蓝圆鲹遗传多样性较高,闽东和闽南群体的遗传结构相似,不存在显著的遗传分化.因此,建议将闽南渔场和闽东渔场作为一个种质资源评估、管理和保护单元.The partial mitochondrial DNA(mtDNA)control region sequences were amplified from two Decapterus maruadsi populations collected from coastal waters in Eastern Fujian(n=30)and Southern Fujian(n=30)and then sequenced to test the population genetic structure and genetic diversity.The results showed that a total of 23 variable sites were found in the aligned sequences of 834--838 bp,and 28 haplotypes were defined in 60 specimens.High levels of haplotype diversity(h=0.948±0.0145),low levels nucleotide diversity(π=0.004 99±0.002 79)and the pairwise nucleotide differences(K=4.173±2.104)were detected,indicating a high level of genetic diversity.Neighbor-joining(NJ) phylogenetic tree was constructed based on the haplotypes from the two populations.However,there was not any genealogy appearing in the haplotypes tree.The star burst structure of the haplotype network also suggested no population structure existed.The demographic history of D.maruadsi was examined by using neutrality tests and mismatch distribution analysis.Demographic expansion was observed in Southern Fujian,starting within the interglacial period before 63 000 a,whereas no population expansion was detected in Eastern Fujian.The regional climate variations caused by the Little Ice Age and overexploitation of Eastern Fujian were predicted to account for the distinct demographic history among clades.Analysis of molecular variance(AMOVA) by ARLEQUIN Version 3.1 revealed all variation occured within populations,which signaled no significant genetic structure throughout the examined range which was in conformity with Neighbor-joining phylogenetic tree and haplotype network.The pairwise Kimura 2-paramter genetic distances calculated by the MEGA Version 4.0 were also small between the two populations.The result showed that D.maruadsi had high genetic diversity and genetically homogeneous population structure between the two populations in Eastern Fujian and Southern Fujian,this might be emerge from strong diffusion capacity,big population size,population expansion and oceanographic current in Fujian coastal waters.The knowledge on genetic diversity and genetic structure will be crucial to establish appropriate fishery management stocks for the fish.福建省科技计划项目(2007I0005

Establishes Method of Avermectin in Four Tissues of Pseudosciaena crocea by High Performance Liquid Chromatography with Fluorescence Detection

建立了大黄鱼肌肉、肝脏、鳃和血液中阿维菌素的高效液相色谱-荧光检测(HPlC-fld)法.样品用乙腈提取,正己烷脱脂,经碱性氧化铝固相萃取(SPE)柱过滤,1-甲基咪唑和三氟乙酸酐的乙腈溶液在室温下避光衍生化,再经甲醇水解,进行HPlC-fld测定.结果表明,阿维菌素在0.5~50.0μg/l范围内线性良好,检测限为肌肉0.5μg/kg,其他组织1.0μg/kg.添加5.0~50.0μg/kg时,平均回收率为:肌肉81.2%~87.1%,肝脏68.9%~74.4%,鳃67.6%~80.7%,血液52.4%~60.3%,日内精密度和日间精密度分别为:肌肉4.6%~5.5%、6.1%~6.8%;肝脏5.5%~6.2%、7.6%~8.7%;鳃5.1%~6.3%、6.9%~7.5%;血液7.6%~8.9%、9.7%~11.6%,该方法能满足现行兽药残留分析要求.In order to understand the avermectin residues in the tissues of Pseudosciaena crocea,a method for the quantitative determination of this drug in fish muscle,liver,gill and blood sample was established using high performance liquid chromatography with fluorescence detection(HPLC-FLD).The procedures for sample treatment included extraction by acetonitrile,delipidation by n-hexane,clean-up by Basic Alumina solid phase extraction cartridge and derivation by 1-methylinidazole and trifluoroacetic anhydride inacetonitrile at room temperature away from light and hydrolysis by methanol.Samples were analyzed by HPLC coupled to a fluorescence detector.The results indicated that the calibration curves were linear within the working range from 0.5 to 50.0 μg/L.The limit of detection of the method for avermectin in muscle was 0.5 μg/kg,and 1.0 μg/kg for the other three tissues.When 5.0-50.0 μg/kg samples were added,the overall recoveries of avermectin in Pseudosciaena crocea muscle,liver,gill and blood sample were respectively 81.2%-87.1%,68.9%-74.4%,67.6%-80.7%,52.4%-60.3%.The intra-day and inter-day coefficient variations for muscle,liver,gill and blood sample detection were 4.6%-5.5% and 6.1%-6.8%,5.5%-6.2% and 7.6%-8.7%,5.1%-6.3% and 6.9%-7.5%,7.6%-8.9% and 9.7%-11.6%,which meet the current requirements for drug residue analysis.This research provides a reference for the detection of avermectin for the tissues of P.crocea.国家公益性(农业)行业专项(200903029

Observation on the Life History of Cryptocaryon irritans in Eastern Sea of Fujian and Research on Antiparasitic Effect of Acetic Acid

近年来,由刺激隐核虫(CryPTOCAryOn IrrITAnS)感染引发的“白点病“为闽东大黄鱼(PSEudOSCIAEnA CrOCEA)养殖业带来连续重大损失.大量针对该病的研究工作已经开展,但仍未找到治疗“白点病“安全、有效的方法.本实验从福建省福鼎市八尺门海域取得一种寄生虫,通过形态学及分子生物学相关鉴定方法,证明了所取寄生虫为刺激隐核虫.利用乙酸对大黄鱼幼鱼进行药物安全性评价,同时开展杀灭不同生活史阶段刺激隐核虫的时间效应与剂量效应实验.结果显示:刺激隐核虫包囊对乙酸的耐受力显著强于幼虫,乙酸在杀灭幼虫的同时,对大黄鱼幼鱼也有一定的刺激作用,因此在实际用药中应严格控制剂量.根据实验结果,建议在生产中防治“白点病“时,要以刺激隐核虫幼虫为目标,使用157.4Mg/l乙酸,累计处理2H,可以有效杀灭全部幼虫,又不会对鱼体造成危害,是一种治疗“白点病“的安全用药策略.Cryptocaryon irritans,a holotrichous marine ciliate protozoan that causing "white spot disease",is one of the most devastating parasites of marine fish in Fujian Province.In the present study,a strain of C.irritans-like parasite was isolated from Pseudosciaena crocea aquiculture areas in eastern Fujian province and then identified.The morphological features and biochemical characteristics of the parasites were identical to those C.irritans strains which had been reported in other places,this fact suggested that the sampled pathogen was C.irritans.In addition,the acute toxicity of acetic acid to juvenile P.crocea and C.irritans was studied.The results showed that C.irritans theronts were more sensitive to acetic acid than tomonts.Meanwhile,the susceptibility of acetic acid to juvenile P.crocea was an important factor which was considered in the process of killing C.irritans.The results indicated that dealt "white spot disease" infected P.crocea with 157.4 mg/L acetic acid for 2 h could kill C.irritans theronts effectively as well as unharmful to the hosts,it was a safe drug-using strategy for preventing and curing "white spot disease".公益性(农业)行业专项(200903029); 国家科技支撑计划项目(2011BAD13B08); 广西科技攻关计划项目(桂科攻1222013-2

Genetic polymorphism of mitochondrial control region and cyt b in Trachurus japonicus from Fujian coastal waters

测定了捕自福建近海闽东渔场和闽南渔场竹荚鱼(TrACHuruS JAPOnICuS)的线粒体dnA(MTdnA)控制区和细胞色素b基因(CyT b)序列。获得了长度为861--866 bP的控制区全序列。60个样本中共检测到66个变异位点和53个单倍型,平均单倍型多样性(H)为0.993,核苷酸多样性(π)为1.093。930 bP的CyT b部分序列共有37个变异位点,从41个样本中得到25个单倍型,平均单倍型多样性(H)和核苷酸多样性(H)分别为0.937和0.336。CyT b片段编码330个氨基酸,氨基酸序列无变异位点,仅有1个氨基酸单倍型。竹荚鱼闽南群体MTdnA序列的遗传多样性高于闽东群体。构建的单倍型系统树未出现明显的以地方群体为单位的家系式分支或者聚簇,群体间的遗传距离(0.01)也较小。分子方差分析(AMOVA)显示,2个群体的遗传变异绝大部分来自群体内部,群体间无显著遗传分化。中性检验和核苷酸不配对分析显示,福建近海竹荚鱼近期经历过种群扩张。种群扩张、扩散能力和东海环流可能促进闽东渔场和闽南渔场竹荚鱼群体间频繁的基因交流,并导致群体间较高的遗传同质性。Trachurus japonicus is a pelagic fish in Carangidae family mainly distributing in coastal waters of China,Japan and Korea.It has been an important commercially marine fish in China since 1960s.But the natural population genetics and resource quantity of T.japonicas have been declined rapidly due to over-fishing,degrada-tion of its natural habitats and so on in recent years,whereas little knowledge is available about its population ge-netic structure.The distribution of genetic diversity and genetic structure in wild stocks of T.japonicus seems emergent and the conservation strategies should be developed nowadays.The complete mitochondrial DNA con-trol region(861-866 bp) was amplified by means of PCR and sequenced from two T.japonicus populations col-lected from coastal waters in Eastern Fujian(n=30)and Southern Fujian(n=30).The result showed that 66 variable nucleotide sites were detected and 53 haplotypes were identified in two populations for control region sequence,but only three shared haplotypes were observed.The global haplotype diversity(h) and nucleotide diversity() were 0.993 and 1.093,respectively.Thirty-seven variable sites were recoded in the aligned sequences of partial cytochrome b gene(930 bp),and 25 haplotypes were recovered in 41 specimens.The average haplotype diversity(h) and nucleotide diversity() of two populations were 0.937 and 0.336,respectively.Cytochrome b gene en-coded 330 amino acids and the amino acid chain had only one haplotype.Each population had good haplotypic diversity and low nucleotide diversity in both mtDNA control region and cytochrome b,and the genetic diversity in Southern Fujian population was higher than that in Eastern Fujian one.Decapterus maruadsi was chosen as out-group in this study,and neighbor-joining(NJ) phylogenetic tree was constructed based on the haplotypes from the two populations of T.japonicus.The result showed that the two populations might have originated from one clade.Analysis of molecular variance(AMOVA) by ARLEQUIN version 3.1 revealed that the variation occurred mostly within the two populations for no significant population genetic structure existed throughout examined range,which was in consistent with neighbor-joining phylogenetic tree and haplotype network.The pairwise Ki-mura 2-paramter genetic distances calculated by MEGA version 4.0 were also small between the two populations.The demographic history of T.japonicus was examined by using neutrality tests and mismatch distribution analy-sis.The neutral test of Fu' Fs,which is devised specifically to detect population expansion and is sensitive to the presence of singletons in a sample,showed several highly significant negative values and the pairwise nucleotide differences distribution showed a singlet,indicating population expanded 62 000-205 000 years ago.The result showed that T.japonicus had low genetic diversity and genetically homogeneous population structure between the two populations,and this might be caused by population expansion,strong diffusion capacity and oceanographic current in East China.All these results would be very important and useful for making scientific strategy for the natural resource protection and genetic breeding of this fish.福建省科技项目(2007I0005

Analysis of genetic diversity of Decapterus maruadsi in the coastal waters of Fujian Province

以往研究表明,福建近海的蓝圆鲹分属2个地理种群,即东海西部种群和闽南—粤东近海地方种群。为研究这两个群系的遗传结构,对蓝圆鲹闽东(30尾)和闽南(32尾)种群进行了AflP分析,8对选择性引物在2个种群62个个体中,共扩增出563个位点,其中多态位点364个。闽东和闽南种群的多态位点比例、nEI遗传多样性指数和SHAnnOn遗传多样性指数分别为62.70%、58.97%,0.1875、0.1809和0.2878、0.2763。与其他鱼类对比显示,福建近海蓝圆鲹种群的遗传多样性水平高,说明种群遗传结构尚未遭到明显破坏;基因分化系数gST、SHAnnOn遗传多样性指数和AMOVA分析均显示蓝圆鲹的遗传变异主要来源于种群内,而种群间无明显的遗传分化。nM显示2个种群间基因交流频繁。种群的显性基因型频率分布和位点差异数分布显示2个种群有基本相同的群体遗传结构。结果表明,蓝圆鲹闽东和闽南种群间无明显的遗传差异,因此可将福建海域的蓝圆鲹划归同一个管理保护单元。较强的扩散能力及海洋环流可能是造成福建近海蓝圆鲹种群间遗传同质性较高的原因。Decapterus maruadsi in Fujian coastal waters has been facing considerable fishing pressure because the resources of main economic demersal fishes have been exhausted.The information of stock structure of different populations will give us theoretical guidance in planning management and conservation of natural resources and in genetic improvement programs.In this study,amplified fragment length polymorphism (AFLP) was used to analyze the genetic diversity and variation of two populations of D.maruadsi from eastern and southern Fujian coastal waters.AFLP bands were scored for presence (1) or absence (0) and transformed into 0/1 binary character matrix.AMOVA and mismatch distribution analysis were performed in Arlequin,Nei genetic diversity and Shannon genetic diversity index were conducted in POPGEN,genetic distance between individuals were computed in Phyltools and the UPGMA tree was generated in MEGA.A total of 563 loci ranging in size from 100 bp to 450 bp were detected from 62 individuals using eight primer combinations,of which 364 were polymorphic.The number of bands per primer combination varied from 54 to 84 and the polymorphic bands per primer combination ranged from 45.83% to 84.44%.The proportion of polymorphic loci,the Nei genetic diversity and Shannon genetic diversity index of these two populations were 62.70 % and 58.97 %,0.187 5 and 0.180 9,0.287 8 and 0.2763, respectively,revealing no significant difference (P >0.05) between the two populations.The results showed that the genetic diversity of these two seemed at the same level.Compared with other fishes,the genetic diversity of D.maruadsi in the coastal waters of Fujian Province is above the middle level,with a higher genetic diversity, indicated that the population genetic structure of D.maruadsi had not been destroyed.Wide range of habitats, unbiased sex ratio and the characteristics of its distribution could be the causes for the higher genetic diversity.Gst value,Shannon genetic diversity index and AMOVA analysis indicated that the genetic variation mainly came from individuals within populations.The UPGMA tree based on genetic distance between individuals exhibited no correlation between clades and geographic distribution.Nm showed that there was frequent gene flow between the two populations.Mismatch distribution analysis and dominant gene frequency revealed that these two populations have the same genetic population structure.The results of AFLP analysis indicated that there is no significant genetic differentiation between two populations.Therefore,it draws a conclusion that D.maruadsi in the costal waters of Fujian Province can be assigned to the same management unit.Annual migrations and larval drift in the ocean currents could be the causes for little genetic structure in the studied area.However,AFLP markers are inherited as dominant markers,further studies utilizing codominant markers are needed for a better understanding of the genetic diversity of this fish.福建省科技计划项目(2007I0005

令和元年台風19号及び台風21号による広域災害に関する総合研究

2020年9月1日(火), 於 : Zoomによるオンライン開催令和元年台風19号では, 東日本の非常に広い範囲で長時間の豪雨をもたらし, 大規模な洪水氾濫が発生すると共に, 台風による過去最多の士砂災害件数, 竜巻などの強風被害, 記録的な高潮・高波被害が発生し, 甚大な人的・物的被害が生じた。その約2週間後には, 日本南岸の低気圧と台風21号による大雨が発生し, 千菓県や福島県などにて洪水氾濫が発生した。本研究では, 科研費・特別研究促進費による突発災害調査研究により, 台風19号, 21号による広域災害について, 全国から多分野の研究者が集結した総合研究を実施した。本報はその成果の一部を取りまとめたものである。また, 今後に向けての提言も一覧できる形でまとめている