Phylogenetic analysis of aerobic anoxygenic phototrophic bacteria and their relatives based on farnesyl pyrophosphate synthase gene

The study aims to reveal phylogenetic and evolutionary relationship between aerobic anoxygenic phototrophic bacteria (AAnPB) and their relatives, anaerobic anoxygenic phototrophic bacteria (AnAnPB) and nonphototrophic bacteria (NPB, which had high homology of 16S rDNA gene with AAnPB and fell into the same genus), and validate reliability and usefulness of farnesyl pyrophosphate synthase (FPPS) gene for the phylogenetic determination. FPPS genes with our modified primers and 16S rDNA genes with general primers, were amplified and sequenced or retrieved from GenBank database. In contrast to 16S rDNA gene phylogenetic tree, AAnPB were grouped into two clusters and one branch alone with no intermingling with NPB and AnAnPB in the tree constructed on FPPS. One branch of AAnPB, in both trees, was located closer to outgroup species than AnAnPB, which implicated that some AAnPB would be diverged earlier in FPPS evolutionary history than AnAnPB and NPB. Some AAnPB and NPB were closer located in both trees and this suggested that they were the closer relatives than AnAnPB. Combination codon usage in FPPS with phylogenetic analysis, the results indicates that FPPS gene and 16S rRNA gene have similar evolutionary pattern but the former seems to be more reliable and useful in determining the phylogenic and evolutionary relationship between AAnPB and their relatives. This is the first attempt to use a molecular marker beside 16S rRNA gene for studying the phylogeny of AAnPB, and the study may also be helpful in understanding the evolutionary relationship among phototrophic microbes and the trends of photosynthetic genes transfer.     

台湾海峡南部海域好氧不产氧光合异养细菌对上升流的响应

国内外关于好氧不产氧光合异养细菌(AAPB)和上升流之间关系的研究甚少。本文采用“基于蓝细菌校正的时序红外显微技术”研究了台湾海峡南部近岸上升流中心区AAPB对上升流变化的响应。研究结果发现,在上升流涌升的初始阶段,AAPB和总异养细菌丰度较低;随着上升流的发展,两者丰度均增加并在上升流的成熟期达到最高值;而当上升流衰退时,AAPB和总异养细菌丰度开始下降。在上升流发展过程中,AAPB丰度与叶绿素a浓度在一定范围内呈显著正相关,但同时受环境低磷浓度的限制,总异养细菌丰度与氮、磷、硅营养盐均有显著正相关,表明叶绿素a指示的浮游植物所释放的溶解有机碳和环境中的磷限制可能对AAPB起着更为直接和重要的作用,而营养盐则可能在总异养细菌对上升流的响应中起着重要作用。本研究有助于我们理解AAPB在碳及其他生源要素循环中的作用及其调控机制。     

南、北极近岸海水中好氧不产氧光合基因pufM的多样性

好氧不产氧光合(AAP)细菌因其具有利用溶解性有机物及光能的能力,在海洋碳循环及能量流动中发挥着重要的作用。该类细菌广泛分布在海洋环境中,其在不同生境中的多样性已被调查。但到目前为止,人们对于高纬度地区好氧不产氧光合细菌的认识还较为缺乏。有鉴于此,本研究基于编码光反应复合物上一个色素结合蛋白亚基的pufM基因,对北极王湾及南极乔治王岛近岸水体中夏季好氧不产氧光合细菌的多样性进行了检测。针对2个王湾站位和2个南极麦克斯维尔湾站位构建了4个pufM基因克隆文库,获得674个阳性克隆子。北极克隆子全部由α-变形细菌组成,而南极克隆子则包括α-变形细菌及β-变形细菌。来源于类似红细菌科的pufM基因在所有样品中皆占据优势。此外,与一株滴状亚硫酸盐杆菌中质粒编码的pufM基因存在亲缘关系的序列,在南、北极样品中均占优势。结果表明海洋环境中的pufM基因存在跨极甚至是环球分布。与此同时,南、北极序列之间的差异,也表明了极地地方种的存在。这些结果显示,作为好氧不产氧光合细菌的红细菌科在两极的近岸水体中具有重要的地位。     

一株多环芳烃芘降解菌的鉴定及其降解特性研究

从红树植物红海榄叶片中分离一株对芘具有较好降解作用的海洋细菌,命名为B11,并对其菌体特征、生长条件及降解效能等进行了系统研究。结果发现该菌株为革兰氏阴性菌,结合其生理生化特征和16S rDNA序列比对,表明此菌株属于交替单胞菌属(Alteromonas sp.)。菌株在芘的起始浓度为100 mg/L,pH为中性或偏碱性条件下,盐度35时对芘的降解作用最强。外加碳源对B11对芘的降解效果都具有一定的抑制作用,其中水杨酸的抑制作用更为显著。综合菌株B11的生长特性和降解效能,初步认为B11菌株较适合用于降解多环芳烃芘,可用于红树植物多环芳烃生态污染的原位修复。     

EROD activities of liver in Mugil so-iuy exposed to benzo (a)pyrene, pyrene and their mixture

Abstract-The effects on hepatic EROD (7-ethoxyresorufin O-deethylase) in Mugil so-iuy exposedto benzo(a)pyrene (BaP), pyrene and their mixtures of equal concentration were investigated, at con-centrations of 0.1, 1.0, 5.0, 10.0, 50.0 μg/dm~3, in experimental condition. Time-effects and dose-response of the biochemical indexs were observed. The results showed that the hepatic EROD activitieswere induced by the exposure of BaP, pyrene and their mixtures at high concentration. Dose-responseconnections were that the hepatic EROD activities were elevated with increasing concentration of the pol-lutants. The combined effect of BaP and pyrene at 1:1 concentration ratio on hepatic EROD activity wasantagonism.     

Cloning, expression and characterization of serine palmitoyltransferase (SPT)-like gene subunit (LCB2) from marine Emiliania huxleyi virus (Coccolithovirus)

The authors have isolated and characterized a novel serine palmitoyltransferase(SPT)-like gene in marine Emiliania huxleyi virus(EhV-99B1).The open-reading frame(ORF) of EhV99B1-SPT encoded a protein of 496 amino acids with a calculated molecular mass of 96 kDa and Ip 6.01.The results of sequence analysis showed that there was about 31%-45% identity in amino acid sequence with other organisms.The maximum likelihood phylogenetic tree suggested that the EhV99B1-SPT gene possibly horizontally transferred from the eukaryote.Hydrophobic profiles of deduced amino acid sequences suggested a hydrophobic,globular and membrane-associated protein with five transmembrane domains(TMDs) motifs.Several potential N-linked glycosylation sites were presented in SPT.These results suggested that EhV99B1-SPT was an integral endoplasmic reticulum membrane protein.Despite lower sequence identity,the secondary and three-dimensional structures predicted showed that the "pocket" structure element composed of 2α-helices and 4βsheets was the catalytic center of this enzyme,with a typical conserved "TFTKSFG" active site in the N-terminal region and was very close to those of prokaryotic organisms.However,the N-terminal domain of EhV99B1-SPT most closely resembled the LCB2 catalysis subunit and the C-terminal domain most closely resembled the LCB1 regulatory subunit of other organisms which together formed a spherical molecule.This "chimera" was highly similar to the prokaryotic homologous SPT.For a functional identification,the EhV99B1-LCB2 subunit gene was expressed in Escherichia coli,which resulted in significant accumulation of new sphingolipid in E.coli cells.     

涌流式培育的大獭蛤稚贝生长的初步研究

2004年11月至2005年1月利用涌流装置和浓缩扁藻进行了大獭蛤(Lutraria(Psam-mophila)maxima)稚贝的中间培育实验。实验所采用的涌流流量为500 mL/min,扁藻密度分别为8 870个/mL±512个/mL和3 368个/mL±557个/mL,稚贝的培育密度分别为20 000,35 000,50 000粒/m2。结果表明,过高的扁藻密度不利于大獭蛤稚贝的摄食,并会造成稚贝的死亡。只有在适当的扁藻密度和涌流流量条件下,大獭蛤稚贝才能正常生长。不同的稚贝培育密度对稚贝的生长存在影响,密度越高,生长越慢;水温低于15℃时,稚贝摄食停止,生长缓慢;稚贝在游离状态下与铺沙条件下的生长基本一致。     

军曹鱼生物学特性及苗种规模化繁育技术

简要介绍了军曹鱼(Rachycentron canadum)的形态、分布、食性、生殖、胚胎和仔稚鱼发育过程等生物学特性以及军曹鱼苗种繁育的技术工艺。选择2~3龄鱼作为亲鱼,在海区网箱中采用营养强化结合激素诱导的方法,使其在整个繁殖季节多次成熟产卵。在大亚湾地区其繁殖季节为4~10月,亲鱼繁殖的适宜水温为25~31℃、适宜盐度为30~34,大批量人工催产的受精率为30%~60%、孵化率为55%~82%。采用室外池塘进行种苗大规模培育,初孵仔鱼在水温26~32℃、盐度28~33条件下培育35~40 d,全长8~11 cm,达到商品苗规格。     

磺胺甲基唑和恩诺沙星在日本囊对虾体内的药代动力学研究

为了研究磺胺甲基唑(Sulfamethoxazole, 简称SMZ)和恩诺沙星(Enrofloxacin, 简称ERFX)在日本囊对虾(Marsupenaeus japonicus)体内的药物代谢动力学特点, 在水温21℃±0.5℃, 盐度29.91 的条件下, 应用萃取、液相测定和反相高效液相色谱法(RP-HPLC)对药物平均回收率、主要动力学参数及在3种组织中的半衰期进行了测定, 检测数据经药物代谢动力学软件3p97 进行研究。结果表明, SMZ 和ERFX 在对虾肝脏、肌肉和血淋巴的平均回收率分别为90.09％, 88.92％, 84.84％和89.14％, 86.04％,91.77％。药物代谢动力学分析表明, SMZ 单次腹部肌肉注射日本囊对虾, 其肝脏药时数据符合二室模型, 肌肉和血淋巴药时数据符合一室模型,半衰期为19.1284, 4.5799, 9.1855 h。ERFX 单次腹部肌肉注射日本囊对虾, 其肌肉药时数据符合二室模型, 肝脏和血淋巴药时数据符合一室模型。半衰期分别为15.16, 16.83, 17.19 h。通过两种药物的代谢特征比较分析表明, SMZ 在肌肉中吸收较ERFX 快, ERFX在血淋巴和肝脏中吸收较SMZ 快。根据SMZ 和ERFX 在21℃±0.5℃的条件下的主要动力学参数及3种组织内消除半衰期, 建议SMZ 在日本囊对虾的休药期不少于10 d, ERFX 的休药期不少于12 d。     

Rates of oxygen consumption and tolerance of hypoxia and desiccation in Chinese black sleeper(Bostrichthys sinensis) and mudskipper(Boleophthalmus pectinirostris) embryos

1Introduction Afossorialmodeoflifehasbeenadoptedbynu merousfishes(Gibson,1982).Burrowingmodeof lifeclearlyoffersanumberofadvantages,butburro wingfishmustalsobeadaptedtocopewiththespe cialproblemsthatthislifestylepresents.Fishthat constructburrowsininterti…     

不同剂型和剂量的维生素C对幼刺参生长的影响

在水温11.0～14.0℃条件下,将平均体质量2.29 g的幼刺参(Apostichopus japonicus)随机放入39个容积50 L塑料水槽中,投喂以玉米蛋白为蛋白源,分别添加0、500、1 000、2 000和4 000 mg/kg V_C-2-三聚磷酸酯(L-ascorby1-2-polyphosphate, LAPP)、V_C-棕榈酸酯(L-ascorby1 palmitate, LAP)和V_C -磷酸酯镁(L-ascorby1-2-monophosphate-magnesium, APM)的13种饲料.90 d的饲养情况表明,摄食添加V_C饲料的刺参的生长、蛋白质效率均显著高于摄食未添加V_C的幼参(P＜0.05).摄食添加LAPP饲料幼参的特殊生长率最高(0.21%/d),添加LAP(0.17%/d)的次之,添加APM(0.15%/d)的最低,分别比对照组(0.12%/d)高75.00%、41.67%和25.00%.摄食添加LAPP饲料幼参的平均蛋白质效率最高(12.57%),摄食添加LAP饲料的(7.76%)次之,摄食添加APM(6.86%)的最低,分别比对照组(5.52%)高127.72%、40.58%和24.28%.在3种剂型V_C组中,饲料系数由低到高依次为:LAPP＜LAP＜APM.饲料中LAPP、APM和LAP的添加量为2 000～2 500 mg/kg、1 000～1 500 mg/kg和2 000～3 125 mg/kg时,幼刺参对饲料蛋白的表观消化率最高,生长最快,饲料系数最低.     

重组别藻蓝蛋白(rAPC)的纯化与鉴定

报道了重组大肠杆菌表达的别藻蓝蛋白的下游生产工艺研究,并对产物进行了分析鉴定。工程菌株P1先在NBSBioFlo3000型5L自动发酵罐进行高密度发酵,融合蛋白获得了高效表达,且主要在细胞内以可溶形式存在。纯化前先将获得的菌体超声破碎,然后经硫酸铵沉淀、疏水层析和离子交换层析三步纯化,接着对纯化的重组别藻蓝蛋白(rAPC)进行SDS-PAGE、PAGE、Westernblot等电点分析。证明已获得纯度为96.4%的rAPC,其等电点为6.0,并且具有与天然APC相似的抗原性。     

织锦巴菲蛤(Paphia textile)过氧化氢酶基因cDNA全长克隆、序列同源分析与组织表达

Catalase is an important antioxidant protein that can protect organisms against various forms of oxidative damage by eliminating hydrogen peroxide. In this study, the catalase c DNA of Paphia textile(Pt CAT) was cloned using RTPCR and rapid amplification of c DNA ends(RACE). Pt CAT is 1 921 bp long and consists of a 5′-UTR of 50 bp, a 3′-UTR of 349 bp, and an ORF of 1 542 bp that encodes 513 amino acids with a molecular weight of 58.4 k D and an estimated isoelectric point of 8.2. Sequence alignment indicated that Pt CAT contained a highly conserved catalytic signature motif(~(61)FNRERIPERVVHAKGAG~(77)), a proximal heme-ligand signature sequence(~(352)RLFSYSDP~(359)), and three catalytic amino acid residues(H~(72), N~(145), and Y~(356)). Pt CAT also contains two putative N-glycosylation sites(~(34)NKT~(36) and ~(437)NFT~(439)) and a peroxisome-targeting signal(~(511)AQL~(513)). Furthermore, Pt CAT shares 53%–88% identity and 29%–89% similarity with other catalase amino acid sequences. Pt CAT m RNA was present in all tested organs, including the heart, digestive gland, adductor muscle, gonad, gill, and mantle, but its expression was highest in the digestive gland. High-temperature-induced stress produced two expression patterns of Pt CAT m RNA: first, an initial up-regulation followed by a down-regulation in the heart, digestive gland, and gonad and, second, consistent down-regulation in all other organs. These results demonstrate that Pt CAT is a typical member of the catalase family and might be involved in the responses to harmful environmental factors.     

Efficient detection of pathogen virus in sand dabs, Paralichthys olivaceus using loop-mediated isothermal amplification (LAMP)

Viral hemorrhagic septicemia virus (VHSV) and marine birnavirus (MABV) are the causative pathogens for some of the most explosive epidemics of emerging viral diseases in many Asian countries, leading to huge economic losses in aquaculture. Rapid molecular detection for surveillance or diagnosis has been a critical component in reducing the prevalence of pathogen infection. The loop-mediated isothermal amplification (LAMP) of DNA is currently one of the most commonly used molecular diagnostic tools, as it is simple, quick, and easy to amplify target DNA under isothermal conditions. In the present study, a novel and highly specific LAMP assay for the sensitive and rapid detection of VHSV and MABV infection in fish was developed. Using a set of synthesized primers matching a specific region of the genome, the efficiency and specificity of the LAMP assay were optimized in terms of the reaction temperature and DNA polymerase concentration, as they are the main determinants of the sensitivity and specificity of the LAMP assay. In particular, we demonstrated that our assay could be applied to efficient detection of VHSV and MABV infection in the wild fish, Paralichthys olivaceus. Our results demonstrate the simplicity and convenience of this method for the detection of viral infection in aquatic organisms.     

金钱鱼脑型芳香化酶基因cDNA 的克隆及表达分析

为了探讨Cyp19a1b基因表达与温度及鱼油添加量的相互关系,奠定金钱鱼人工繁育理论基础,作者采用RT-PCR和RACE法,克隆了金钱鱼(Scatophagus argue Linnaeus)脑型芳香化酶基因(Cyp19a1b);采用实时荧光定量PCR法,检测了不同温度(23、26和29℃)和不同水平鱼油(0、2%和6%)处理后二龄雌性金钱鱼脑垂体中Cyp19a1b mRNA表达。结果表明:Cyp19a1b cDNA全长2409bp,5′端非编码区164 bp,3′端(不包括poly(A))712 bp,开放阅读框(ORF)1506 bp,编码501个氨基酸,推测其蛋白质分子量为56.697 kDa。序列分析及分子系统进化树结果表明:金钱鱼Cyp19a1b氨基酸序列与黄锡鲷和舌齿鲈Cyp19a1b同源性较高,分别为86.2%、86.5%,与鲤鱼(Cyprinus carpio)、稀有鲫(Gobiocypris rarus)、斑马鱼(Danio rerio)、赤点石斑鱼(Epinephelus akaara)、大黄鱼(Larimichthys crocea)、南方大口鲇(Silurus meridionalis)和鲻鱼(Mugil cephalus)Cyp19a1b同源性为64.1%~84.4%,但与上述7种鱼性腺芳香化酶(Cyp19a1a)同源性低于63.5%,同源性分析结果与根据传统形态学和生化特征分类的结果相一致。荧光定量PCR结果显示,随处理时间的延长,26℃和29℃组金钱鱼脑垂体中Cyp19a1b mRNA的表达量都逐渐降低,但29℃组显著低于26℃组(P0.05),而23℃组在6周时则显著高于26℃和29℃组(P0.05),约为26℃组2倍的表达量;鱼油处理3个投喂组脑垂体中Cyp19a1b mRNA的表达量均逐渐降低,但鱼油对金钱鱼脑垂体中Cyp19a1b mRNA的表达量无显著影响(P0.05)。以上结果表明:随着卵巢发育或鱼油含量的提高,可降低雌性金钱鱼脑垂体中Cyp19a1b mRNA的表达,这种降低可能是E2反馈调节所致;水温高于26℃将抑制Cyp19a1b mRNA的表达。     

Effects of UVB on the expression of proliferating cell nuclear antigen (PCNA) in dinoflagellate Prorocentrum donghaiense Lu

PCNA has been considered as a useful marker for the estimation of growth rates of marine phytoplankton at the species level. Since dinoflagellates are noted for having many prokaryotic features in that they are the only eukaryotes to have permanently condensed chromosomes as well as lacking histones and a nucleosome, the sensitivity to UVB radiation and the validity of PCNA as a maker of growth rate in dinoflagellate need to be evaluated.Prorocentrum donghaiense Lu was investigated to valuate the UVB sensitivity in relation to cellular and molecular aspects of PCNA as a growth indicator.The effects of UVB radiation on PCNA were studied using the methods of western blots technology and whole-cell immunoflurescence with one mono-antibody. UVB changed the cell morphology, halted the growth and increased the cell size, even caused cell death to a certain extent after treatment with UVB radiation in P. donghaiense. Compared with the control, treating the algal cultures in exponential phases with UVB radiation for 24 h caused chromatin release and increases in protein levels of PCNA.     

栉孔扇贝体内麻痹性贝毒的累积与排出过程研究

通过室内摄食实验,研究了栉孔扇贝(Chlamys farreri)对麻痹性贝毒(PSP)产毒藻的敏感性及其累积和排出毒素的特征.有毒微小亚历山大藻(Alexandrium minutum)在短时间内可对栉孔扇贝的摄食有一定的抑制作用,且投喂毒藻的浓度越大,对摄食作用的抑制越明显.栉孔扇贝对PSP贝毒的累积能力很强,实验中,48 h后内脏累积毒素就高达5000μg STXeq100g-1,但其毒素排出速率较慢,在24d解毒阶段后内脏中的毒素还没有下降至国际贝毒食用安全标准水平的80μg STXeq 100g-1.各毒素组成比在整个累积与排除过程中有较大变化,GTX1,GTX4在累积阶段比例下降,同时GTX2,GTX3比例上升;内脏中GTX2:GTX3的比值逐渐增加.毒藻中PSP贝毒毒素组成和贝体中PSP贝毒毒素组成之间的主要差别是GTX4含量的减少和GTX1含量的升高;而扇贝粪便中的毒素组成却与对数生长期的毒藻极为接近.