Isolation and identification of algicidal substances produced by Brevibacterium sp. BS01 and algicidal mechanism of algicidal compound against Alexandrium tamarense

赤潮的控制与防治对于保证海洋环境安全和海洋生态健康是至关重要的。本论文主要研究了海洋杀藻放线菌BS01的种属关系；根据BS01杀藻活性物质的基本理化性质，分离纯化得到一个具有杀藻活性的化合物；同时探讨了塔玛亚历山大藻在BS01胞外分泌物胁迫下生理生化水平的响应；考察藻细胞死亡过程中细胞形态变化；氧化压力下藻细胞光合系统损伤情况；综合运用real-timePCR技术研究了藻细胞在杀藻活性物质作用下靶功能基因psbA、psbD、hsp、cox和cob转录水平变化。主要研究结果如下： （1）分离鉴定得到一株具高效稳定杀藻功能的放线菌，菌株为杆状，属于放线菌目，短杆菌属Brevibacteriums...Control of red-tide is becoming vital for marine security and ecological recovery. Marine actinomycetes are playing an irreplaceable role on marine biodiversity and stability of marine ecosystem, providing a possible path for controlling red-tide. In this paper, the typical harmful alga, Alexandrium tamarense (ATGD98-006), and algicidal actinomycete Brevibacterium sp. BS01 were studied as dominan...学位：理学硕士院系专业：生命科学学院_微生物学学号：2172009115212

高效溶藻放线菌BS01发酵培养基及发酵条件优化

从漳江口红树林区采集的沉积物样品中分离到1株放线菌菌株BS01 Brevibacterium sp.,其胞外活性产物对塔玛亚历山大藻Alexandrium tamarense具有明显的溶藻作用。采用单因素及均匀设计,通过摇瓶培养对BS01产溶藻活性物质的发酵培养基及发酵条件进行优化。结果表明,在可溶性淀粉为碳源、硝酸钠为氮源、装液量为40%、起始pH值为7.5、培养温度为28℃、转速为150r.min?1、振荡培养时间为48h的条件时,BS01发酵产物的杀藻活性最强。通过均匀设计进行最佳发酵培养基及培养条件优化的结果为:可溶性淀粉为20g.L?1,硝酸钠为0.5g.L?1,pH为7.7,温度为27.2℃。研究结果为杀藻活性物质高效提取及杀藻机制研究奠定了基础

Optimization of Fermentation Conditions of Marine Algicidal Bacterium Microbulbifer sp. BS03 by Uniform Design

采用均匀设计法设计和二次多项式逐步回归分析,对一株高效杀塔玛亚历山大藻微泡菌bS03(MICrObulbIfEr SP.)产杀藻活性物质的发酵培养条件进行优化.通过单因素实验筛选出碳源、氮源、PH、培养时间和接种量为显著影响因子,并对5个显著影响因子采用u15(155)水平对培养基进行优化.结果表明bS03最适发酵培养条件为:蔗糖8 g/l,蛋白胨10.50 g/l,初始PH值7.5,培养时间32 H,接种量3.00%.验证试验结果显示,在此条件下该菌发酵液的干重为4.725 g/l,较优化前增加了31.35%,ld50为0.768%,较优化前降低了25.14%.研究结果为杀藻活性物质以及杀藻机理的研究奠定了理论基础.The optimal parameters of fermentation conditions of marine bacterium Microbulbifer sp.BS03 producing algicidal substances against Alexandrium tamarense were obtained by using Uniform Design and quadric polynomial regression methods.Five main factors were achieved through single factor experiments,that is,carbon source,nitrogen source,pH,incubation time and inoculum size.The U15(155) design result showed the optimum conditions of experiment were as follows: sugar 8 g/L,peptone 10.5 g/L,initial pH 7.5,incubation time 32 h,and inoculum size of 3.00%.According to the verification experiment,under the optimal conditions,the dry biomass was 4.725 g/L which was increased by 31.35% and LD50was 0.768% which was decreased by 25.14% compared with those of the basic fermentation conditions.This outcome will help us to separate active substances from complex components in medium and thus to reveal the mechanism of algicidal activity in the future.国家自然科学基金项目(Nos.40930847;31070442);福建省自然科学基金项目(Nos.2012J01150;2010J01223);福建省教育厅项目(No.JA10232);莆田市科技计划项目[No.2011S09(4)]资助;theOpenFundoftheKeyLaboratoryoftheMinistryofEducationforCoastalandWetlandEcosystemsofXiamenUniversity(CWel0902)---

PPARγ激动剂罗格列酮对大鼠心肌缺血-再灌注心律失常的影响

【目的】探讨PPARγ激动剂对大鼠心肌缺血-再灌注心律失常的影响及相关机制。【方法】将雄性SD大鼠60只随机分成假手术（Sham）组、缺血再灌注（I／R）组、缺血再灌注＋罗格列酮（I／R＋Ros）组、缺血再灌注＋罗格列酮＋缓激肽B2受体拮抗剂HOE140（I／R＋Ros＋HOE140）组，每组15只。麻醉大鼠后，结扎大鼠冠状动脉前降支根部30min，再灌注40min，观察缺血再灌注前后心律失常发生情况，检测肌组织丙二醛（MDA）、超氧化物歧化酶（SOD）、总一氧化氮合酶（tNOS）、还原型一氧化氮合酶（eNOS）、诱导型一氧化氮合酶（iNOS）和一氧化氮（N0）水平。【结果】罗格列酮预处理组的大鼠恶性室性心律失常的发生时间（13min），持续时间（14．37s），室性早搏的发生次数（47次）和心律失常评分等指标均得到改善（P〈0．05）；提前应用Hoe140可部分或全部阻断缺血一再灌注时罗格列酮的抗心律失常作用（P〈0．05）；I／R＋Ros组与I／R组相比心肌组织eNOS、NO、和SOD活性水平明显提高，而iNOS活性和MDA水平显著下降（P〈0．051；HOE140可阻断此作用（P〈0．05）。【结论】PPARγ激动剂罗格列酮有抗大鼠心肌缺血再灌注心律失常作用，这可能是通过缓激肽-一氧化氮途径实现的

房室结慢径消融电生理特性与临床疗效关系探讨

【目的】探讨房室结折返性心动过速(AVNRT)慢径消融时心内电生理改变与临床疗效的关系。【方法】对94 例慢-快型AVNRT 病人采用射频消融慢径治疗, 分析消融前后电生理特征、消融时心内电生理改变和消融终点。【结果】94 例均消融成功, 其中75 例(80 %)慢径阻断, 19 例(20 %)慢径未阻断, 其中2 例(2 %)慢径未阻断者并有1 ～ 2 个心房回波;术 后随访(28 ±17)月, 4 例术后1～ 5 月复发, 经再次消融成功。术中5 例消融时呈持续交界心律, 经消融后仍然有心动过速发 作, 调整靶点后消融成功;4 例放电时有房室传导阻滞先兆, 用时间滴定法消融成功。【结论】慢径路消融的理想终点应该是 电刺激诱发的心动过速消失;消融时出现持续交界心律是安全的, 部分患者需调整消融靶点提高消融成功率;有房室传导阻 滞先兆者采用时间滴定法, 可避免永久性房室传导阻滞发生

室房间期差值对间隔部旁道与快径逆传心动过速的鉴别诊断价值

【目的】研究间隔部旁道参与的和房室结功能连续曲线性快径逆传的室上性心动过速的电生理特征, 为射频治 疗提供准确的定位依据。【方法】选择10 例间隔旁道介导的房室折返性心动过速(AVRT, 旁道组)与10 例房室结功能连续曲 线的房室结折返性心动过速(AVNRT, 房室结组)病人, 分别在与心动过速相同的频率起搏右心室及心动过速发作时, 测量希氏 束(HBE)、冠状窦近端(CS9 -10)导联记录的室房传导间期(VAVP和VASVT)及两者差值ΔVA(VAVP -VASVT)。【结果】①房室结组 心室起搏与心动过速时VA 间期差异显著(P 0.05);②心室起搏时旁道组与房室结组 的VAVP相比差异无显著性(P >0.05);③心动过速时VA 间期旁道组明显长于房室结组, 两者相比VASVT差异显著(P < 0.001);④旁道组ΔVA 与房室结组比较差异显著(P ≤0.001)。【结论】CS9-10导联ΔVA 是鉴别隐性间隔部旁道介导的AVRT 与AVNRT 的重要参考指标, 特别对房室结功能连续曲线性房室结折返性心动过速的鉴别有较高价值

Isolation,identification and characterization of algicidal bacterium BS03 against Alexandrium tamarense

作者简介: 李东( 1986—) ，男，E-mail: lidongbox2006@163． com; * 通讯作者( 责任作者) ，E-mail: wshwzh@ xmu． edu． cn Biography: LI Dong( 1986—) ，male，E-mail: lidongbox2006@163． com; * Corresponding author，E-mail: wshwzh@ xmu． edu． cn[中文文摘]从福建漳江口红树林区筛选出一株对产贝毒赤潮原因藻——塔玛亚历山大藻(Alexandrium tamatense)具有强溶藻能力的细菌,命名为BS03,通过生理生化及16S rDNA序列分析对该菌株进行鉴定,并探讨了菌株BS03对塔玛亚历山大藻的溶藻特性和溶藻代谢产物的初步性质.结果发现,菌株BS03属于微泡菌属(Microbulbifer sp.)相似性达99%;对塔玛亚历山大藻的杀藻效果具有一定浓度效应,在一定浓度范围内处理浓度越高,溶藻效果越好;菌株BS03对处于不同生长时期的塔玛亚历山大藻都表现出较好的杀藻效果,其中对处于延滞期的塔玛亚历山大藻表现出最佳的杀藻效果,处理96h后,抑藻率达98.17%;不同生长期菌株对塔玛亚历山大藻溶藻作用无明显差异;菌株BS03通过间接作用方式溶藻,所分泌的胞外活性物质的分子量小于1kDa,耐酸碱、具热稳定性,推测为非蛋白质、非核酸和非多糖类物质. [英文文摘] A bacterial strain，BS03，exhibiting strongest activity against the toxic dinoflagellate，Alexandrium tamatense，was isolated from the Zhangjiang Estuary Mangrove National Natural Reserve，China． Based on 16S rRNA gene sequences and biochemical and morphological characteristics，the strain BS03 was determined to be Microbulbifer sp． on the basis of 99． 0% similarity with reference strain sequences from the NCBI． The growth of A． tamatense was strongly suppressed by BS03 in all growth phases． With the strongest algicidal activity noted against harmful algae in the lag stage，the removal rate of A． tamatense reached 98.17% after 96 h． In addition，the initial concentration of the bacterial culture strongly affected the algicidal ability． The higher concentration of the bacterial culture was，the stronger algicidal activity to the A.tamatense.Furthermore，BS03 indirectly attacked A． tamatense by algicidal substances． The molecular weight of algicidal compounds were less than 1 kDa，and heat tolerant and stable in acidic and alkaline conditions． According to our results，we speculated the algicidal compounds could not be proteinaceous，nucleate and polysaccharide． These findings indicated that strain BS03 could be a potential bio － agent for future use in controlling harmful algal blooms．国家自然科学基金重点项目、面上项目(No.40930847,31070442);福建省自然科学基金(No.2010J01150);海洋公益性行业科研专项经费资助(No.201305016,201305041,201305022

Optimization of Fermentation Conditions of Marine Algicidal Bacterium Microbulbifer sp. BS03 by Uniform Design

通讯作者 Corresponding author (E-mail: [email protected]; [email protected])[中文文摘]采用均匀设计法设计和二次多项式逐步回归分析,对一株高效杀塔玛亚历山大藻微泡菌BS03(Microbulbifer sp.)产杀藻活性物质的发酵培养条件进行优化.通过单因素实验筛选出碳源、氮源、pH、培养时间和接种量为显著影响因子,并对5个显著影响因子采用U15(155)水平对培养基进行优化.结果表明BS03最适发酵培养条件为:蔗糖8 g/L,蛋白胨10.50 g/L,初始pH值7.5,培养时间32 h,接种量3.00%.验证试验结果显示,在此条件下该菌发酵液的干重为4.725 g/L,较优化前增加了31.35%,LD50为0.768%,较优化前降低了25.14%.研究结果为杀藻活性物质以及杀藻机理的研究奠定了理论基础.[英文文摘]The optimal parameters of fermentation conditions of marine bacterium Microbulbifer sp.BS03 producing algicidal substances against Alexandrium tamarense were obtained by using Uniform Design and quadric polynomial regression methods.Five main factors were achieved through single factor experiments,that is,carbon source,nitrogen source,pH,incubation time and inoculum size.The U15(155) design result showed the optimum conditions of experiment were as follows: sugar 8 g/L,peptone 10.5 g/L,initial pH 7.5,incubation time 32 h, and inoculum size of 3.00%. According to the verifi cation experiment, under the optimal conditions, the dry biomass was 4.725 g/L which was increased by 31.35% and LD50 was 0.768% which was decreased by 25.14% compared with those of the basic fermentation conditions. This outcome will help us to separate active substances from complex components in medium and thus to reveal the mechanism of algicidal activity in the future. Fig 7, Tab 5, Ref 20.国家自然科学基金项目(Nos.40930847,31070442); 福建省自然科学基金项目(Nos.2012J01150,2010J01223);; 福建省教育厅项目(No.JA10232);莆田市科技计划项目[No.2011S09(4)]资助

Advances in Algicidal Substances Produced by Algae-lysing Actinomycetes

作者简介: 傅丽君( 1975-) , 女, 福建莆田人, 副教授, 主要从事环境生态学研究. E-ma i:l lijun _fu@ s ina. com 通讯作者: 郑天凌( 1955-) , 男, 福建莆田人, 教授, 主要从事环境微生物研究. E-mai:l m icrozh@ xm u. edu. cn[中文摘要]赤潮是一种严重的全球性海洋自然灾害,近年来赤潮发生次数增多,发生区域扩大,危害加剧。放线菌是产生生物活性物质相对较多的一类微生物,溶藻放线菌的发现及其代谢产物多样性,在赤潮生消过程中起着越来越重要的作用,为微生物防治赤潮及新型杀藻剂的开发提供了可能。采用溶藻微生物进行赤潮和水华的防治已经被深入认识和充分肯定,成为当前赤潮研究的重点和热点。概述了溶藻放线菌发现及其活性物质分离筛选的研究进展以及赤潮防治的新方法,强调了放线菌及其代谢产物在生态环境中的重要作用,提出了利用放线菌防治赤潮的可能性及有效性,并对溶藻放线菌及其活性物质的研究趋势和应用前景进行了展望。[英文摘要]The dram atically increasing occurrence of harmful algal blooms (HABs) a ll over the world has at tracted a great dea l of a ttentions from researchers tying to mitigate the blooms. Some management strategies have been developed such as flocculation of microalgae with clay, biological agents. It is now fully convinced that blooms can be controlled and exorcised through microorganism, which has already become the hot issue. All we know is that actinomycete is a kind of microorganism producingmuch more bioactive substance relatively. Based on the var-ieties and high complexity of me tabo lites, actinomyces with algicida l features are playing more and more important role inHAB s. It is possible that alg icida l act inomyces will contribute to exploring new algicidal agents andm it igating the blooms. In th is paper, the find ing of a lgicidal actinomycetes and screening of bioactive substance, as well as new methods for controlling blooms are reviewed. Also, it focuses on the current deve lopment in the research o f a ig-icida l actinomycetes and algicida l substances and refers to possibility and sign if icance of algicidal actinomycetes maginiting blooms. A t las,t the paper prosoects the research trendency and application of research ing a lg icida l act-i nomycetes and relevant bioact ive substances. Th is implies exploring and find ingvarious mechan isms will be a long period and and will provide a better understanding of close beneficia l interactions between algae and actinomycetes in favor of the effect of theseing teractions on algal blooms.国家高技术研究发展计划项目“海洋微生物抑藻(有毒赤潮藻)活性化合物的发现与利用”(编号:2008AA09Z408);国家自然科学基金重点项目“我国近海海洋细菌的生态过程及关键菌群影响赤潮生消的机理”(编号:40930847);滨海湿地生态系统教育部重点实验室开放基金项目“基于有害赤潮消控的红树林湿地杀藻放线菌的筛选与作用方式定位研究”(编号:Cwel0902);福建省自然科学基金项目“高效抑藻放线菌对有毒赤潮藻的生理生态效应及作用机理研究”(编号:2010J01223)资