两种沼虾溶菌酶基因ORF的克隆和罗氏沼虾溶菌酶基因的组织表达(英文)

分别提取罗氏沼虾和日本沼虾血细胞总RNA,RT-PCR扩增获得特异性cDNA片段,纯化后克隆到T载体上。序列测定表明所克隆的两种沼虾溶菌酶基因的开放阅读框(ORF)为477bp,共编码158个氨基酸,包括溶菌酶成熟肽140个氨基酸残基和信号肽18个氨基酸残基。同源性分析表明,罗氏沼虾和日本沼虾溶菌酶基因的碱基序列及推测氨基酸序列高度同源,分别为99.4%和98.1%。两种沼虾溶菌酶基因的碱基序列和推测氨基酸序列与Gen-Bank上其他对虾溶菌酶的同源性达83.0%和80.0%以上。两种沼虾溶菌酶都具有c-型溶菌酶典型的两个酶活性位点(Glu51)和(Asp68),以及8个保守结构氨基酸残基Cys,且在101、106和107位上缺少Asp,因而推测本实验所克隆的两种沼虾溶菌酶基因属c-型溶菌酶基因的非钙结合亚型。以PCR法制备罗氏沼虾溶菌酶基因的生物素标记探针,斑点杂交检测感染弧菌后溶菌酶基因mRNA在各组织中的转录水平,结果表明受感染6h后在眼、肌肉、鳃、肝胰腺、肠管中的表达量均有升高,其中在肝胰腺中的表达量最高,约为对照组的560%。在不同感染时间里,肝胰腺中该基因表达量有较大的变化:感染后3h表达量最低,24h后表达量升至最高,大约为对照组的430%,48h时的表达量又有所下降,但仍明显高于对照组(约为330%)。受弧菌感染后罗氏沼虾溶菌酶基因转录的上调证明溶菌酶基因在非特异性免疫中的直接作用,同时表明肝胰腺可能在沼虾的免疫防御过程起重要作用。       

复方中草药对罗氏沼虾碱性磷酸酶(AKP)和血清蛋白的影响

为了探究中草药添加剂对罗氏沼虾(Macrobrachium rosenbergii)免疫功能的影响,以罗氏沼虾为对象,在水温20～30℃、pH 7～8的试验条件下,往饲料中分别添加质量分数为0%、1.0%、1.5%、2.0%、2.5%、3.0%的复方中草药制剂,连续饲喂25 d。结果显示:随着饲料中复方中草药质量分数的提高,罗氏沼虾的生长指标、血清中碱性磷酸酶(AKP)活性、血清蛋白含量均呈现先上升后下降的趋势,试验组相较于对照组有很大提高;中草药质量分数的提高引起的血清中蛋白含量的提高远远大于肌肉中蛋白的含量,在各试验组中2.5%的复合中草药制剂的添加量效果最好。试验证明,添加复方中草药制剂对罗氏沼虾的免疫活性、生长性能均有明显的促进作用。该结果能够为罗氏沼虾的健康养殖提供基础资料。     

日本三宅岛火山土壤中硫代硫酸盐氧化菌的分离、生物学特性及鉴定

【目的】利用培养法从日本三宅岛火山土壤(堆积年限131年)中分离到一株能氧化分解硫代硫酸盐的细菌MU2A-22T。【方法】用培养法对该菌株MU2A-22T进行了生理生化性质以及分类学位置上的确定。【结果】菌株MU2A-22T为革兰氏阴性,短杆状或球状。理化性质表明该菌株能利用葡萄糖、L-阿拉伯糖、葡萄糖酸盐、己二酸酯、dL-苹果酸钠、硫代硫酸钠(最适浓度为2.5 mmol/L)为唯一碳源进行自养生长。最适生长温度为25°C 30°C,最适pH为6.0 8.0。菌株MU2A-22T的16S rRNA序列与菌株Paracoccus solventivorans 6637T亲缘关系最近,序列相似性为97%,编码核酮糖-1,5-二磷酸羧化酶/加氧酶(Rubisco)的基因也被确定。对Paracoccus属内几种近缘菌的脂肪酸分析,证明菌株MU2A-22T中含有Paracoccus属的特征氨基酸,其中含量大于10%的分别为C18:1(74.7%)和C18:0(12.1%)。DNA-DNA杂交实验表明,菌株MU2A-22T与Paracoccus solventivorans 6637TDNA的相似度为49.3%。MU2A-22T菌株G+C含量为66.5%66.7%。【结论】菌株MU2A-22T为Paracoccus属内的一新种菌(登录号GQ452286),命名为Paracoccus scorialis sp.nov.。     

暗黑菌(Atribacteria)的环境分布和功能特征

暗黑菌门包括OP9和JS1两大分支,成员大多为未培养微生物,在自然环境中广泛分布,并且在部分环境如厌氧海洋沉积物、地热环境以及油藏中为优势微生物。基于基因组信息的研究表明,暗黑菌为严格的厌氧微生物,同时具有降解糖类、小分子酸、短链正构烷烃的能力,在地球碳循环过程中可能扮演着重要的角色。然而,由于缺乏相应代表性的纯培养菌株,对于暗黑菌的生理生化功能推测仍有待验证。文章概述了暗黑菌的发现及发展历史,分析了其环境分布和多样性,简述了目前提出的三种代谢方式,提出了未来暗黑菌的研究发展方向。     

大叶藻(Zostera marina)海草床沉积物细菌和古菌丰度及组成的垂直剖面特征

【背景】海草床是重要的"蓝碳"生态系统,对全球碳汇有重要贡献。海草床沉积物剖面的垂直梯度特征显著,表层呈现氧化态,富含活性有机质,而深层呈还原态,以惰性有机质为主。【目的】探究这种垂直特征如何影响微生物的丰度和群落分布。【方法】利用荧光定量PCR和16SrRNA基因高通量测序等手段,测定了山东荣成天鹅湖大叶藻海草床不同深度(5、10、15、20、25和30 cm)沉积物中细菌和古菌丰度、多样性和群落结构的变化。【结果】细菌和古菌16S rRNA基因拷贝数随深度的增加而降低,在沉积物5cm深处,细菌的16SrRNA基因拷贝数显著高于20cm和30cm层(ANOVA,P<0.05)。深度对细菌和古菌α多样性指数没有显著影响(P>0.05)。细菌中相对丰度最高的是变形菌门,其次是绿弯菌门,拟杆菌门,浮霉菌门等,其中δ-变形菌和浮霉菌的相对丰度随深度显著增加(P<0.05)。古菌群落中深古菌门比例最高,在25cm深处达到70%以上;其次是乌斯菌门、洛基古菌门、广古菌门和奇古菌门等。奇古菌门比例随深度增加而显著降低(P<0.05),其他古菌类群在不同深度间差异不显著(P&...     

Mn(Ⅱ)氧化细菌Pseudomonas aeruginosa L3的分离、鉴定及氧化特性

【背景】Mn(Ⅱ)氧化细菌是一类可以沉积和氧化Mn(Ⅱ)从而形成固态锰氧化物的细菌,在生物地球化学和环境修复领域中引起了广泛关注,但目前所研究的Mn(Ⅱ)氧化模式菌株中大多来源海洋,土壤源Mn(Ⅱ)氧化细菌涉及较少。【目的】丰富土壤源Mn(Ⅱ)氧化细菌的来源与物种多样性,同时也为方铁锰矿型Mn₂O₃的潜在应用提供新菌种。【方法】从湖南省株洲市已关停的清水塘冶炼工业园区内筛选得到一株具有Mn(Ⅱ)氧化能力的铜绿假单胞菌(Pseudomonas aeruginosa) L3,并就其分离纯化、鉴定、生长曲线、pH变化、Mn(Ⅱ)氧化特性及锰氧化物制备等方面进行系统研究。【结果】Pseudomonas aeruginosa L3的菌液离心后的上清液及提取的绿脓菌素(pyocyanin, PYO)均具有Mn(Ⅱ)氧化能力,与菌液相比,上清液对Mn(Ⅱ)氧化的能力更强。进一步利用X射线衍射(X-ray diffraction, XRD)对Pseudomonas aeruginosa L3产生的锰氧化物进行晶相分析,发现该锰氧化物在2θ为32.951°和5...     

罗氏沼虾良种选育研究进展

罗氏沼虾（Macrobrachium rosenbergii）是世界重要的经济虾类,畅销国内外市场,备受消费者的喜爱。中国是罗氏沼虾养殖大国,年均养殖产量占世界一半以上。种质资源是我国罗氏沼虾产业发展的原动力,处于整个产业链的最上游,并支撑着整个产业的发展。世界多国为了壮大产业规模,也纷纷开展了罗氏沼虾种质资源库的建设与良种选育。本文回顾了我国自主选育的罗氏沼虾主要品种及品系,综述了大规模家系的构建以及不同来源群体的遗传多样性分析,并对国外的良种选育现状和趋势进行了概述,最后结合国内外的研究进展,提出了良种选育的研究要点与方向,以期为今后的种质创新研究提供思路和借鉴。     

Distribution of dopamine and octopamine in the central nervous system and ovary during the ovarian maturation cycle of the giant freshwater prawn, Macrobrachium rosenbergii

Dopamine (DA), octopamine (OA) and serotonin (5-HT) are the key neurotransmitters that control gonadal development in decapod crustaceans. 5-HT stimulates, while DA and OA delay gonadal development in Macrobrachium rosenbergii. In the present study, we have further investigated the distribution patterns of DA and OA in the central nervous system (CNS) and ovary during various stages of the ovarian maturation cycle of this giant freshwater prawn. DA- and OA-immunoreactive neurons and fibers were distributed extensively in several regions of the brain, subesophageal ganglion (SEG), thoracic ganglia and abdominal ganglia. In the brain, the two neurotransmitters were present in neurons of clusters 6, 7, 11, 17, and nearby neuropil regions. In the SEG, thoracic ganglia and abdominal ganglia, immunoreactive neurons and fibers were found along the midline and in several neuronal clusters around each neuropil region. Staining for DA and OA was more intense in the thoracic ganglia than in other parts of the CNS. In the ovary, DA- and OA-immunoreactivities were present at high intensity in early oocytes. The presence of DA- and OA-immunoreactivities in neural ganglia as well as ovary suggests that DA and OA may also be involved in the reproductive process, particularly ovarian development and differentiation of oocytes in this species.     

亚硝酸盐胁迫对罗氏沼虾血细胞及其抗氧化酶活力的影响

【背景】亚硝酸盐是虾类集约化养殖过程中最常见的毒性污染物之一,研究亚硝酸盐胁迫对罗氏沼虾血细胞的毒性以及抗氧化酶在抗胁迫防御中的作用,能够为罗氏沼虾养殖过程中的亚硝酸盐中毒防治提供理论参考。【方法】以不同浓度(0、1、5和10 mg·L~(-1))的亚硝态氮(NO_2~--N)对罗氏沼虾进行胁迫,于胁迫后的0、6、12、24和48 h取样,应用流式细胞术检测血细胞活性氧(ROS)含量和细胞凋亡率,同时测定血细胞总数(THC)和胞内抗氧化酶活力。【结果】1 mg·L~(-1)NO_2~--N在48 h内对血细胞ROS含量、凋亡率和THC均无显著影响。5 mg·L~(-1)NO_2~--N胁迫24 h,血细胞ROS含量显著上升,THC显著下降,胁迫48 h凋亡率显著提高。10 mg·L~(-1)NO_2~--N胁迫6 h,血细胞ROS含量和凋亡率均显著上升,胁迫12 h THC显著下降。血细胞的超氧化物歧化酶(SOD)、过氧化氢酶(CAT)和谷胱甘肽过氧化物酶(GPx)的活力均不同程度地被NO_2~--N胁迫所诱导,CAT活力主要在胁迫前期提高,而GPx活力在胁迫后期提高。【结论与意义】亚硝酸盐存在浓度和时间毒性效应,一定浓度的亚硝酸盐会诱导虾血细胞产生ROS,这些ROS的过量产生诱导了血细胞发生凋亡,继而导致THC下降,这一氧化胁迫过程可能是亚硝酸盐对罗氏沼虾产生细胞毒性的重要机制之一。抗氧化酶活力的诱导表明抗氧化酶在亚硝酸盐胁迫过程中发挥防御作用。     

Spermatogenesis and distinctive mature sperm in the giant freshwater prawn, Macrobrachium rosenbergii (De Man, 1879)

The structures of differentiating male germ cells in the testis of the giant freshwater prawn, Macrobrachium rosenbergii, were studied by light and electron microscopy. Based on ultrastructural characteristics, the developing male germ cells are classified into 12 stages, including spermatogonia, six phases of primary spermatocytes (leptotene, zygotene, pachytene, diplotene, diakinesis and metaphase), secondary spermatocyte, three stages of spermatids and mature sperm. During spermatogenesis, the differentiating germ cells have characteristics similar to those of other invertebrates, but they exhibit some unique characteristics during spermiogenesis. In particular, an early spermatid has a round nucleus with highly condensed heterochromatin, appearing as thick interconnecting cords throughout the nucleus. In contrast to most invertebrates and vertebrates, the chromatin begins to decondense in one-half of the nucleus at the mid spermatid stage. In the late spermatid, the chromatin becomes almost entirely decondensed with only a small crescent-shaped heterochromatin patch remaining at the anterior pole of the nucleus. Mature sperm possess an everted umbrella-shaped plate with a spike covering the anterior pole of the nucleus, whose chromatin is totally decondensed as only small traces of histones H3 and H2B remain. The acrosome appears at the ruffled border of the spike plate as small sac-like structures. Few mitochondria remain in the cytoplasm at the posterior pole.     

A cytosolic glutathione s-transferase,GST-theta from freshwater prawn Macrobrachium rosenbergii: molecular and biochemical properties

Glutathione S-transferases play an important role in cellular detoxification and may have evolved to protect cells against reactive oxygen metabolites. In this study, we report the molecular characterization of glutathione s-transferase-theta (GST-θ) from freshwater prawn Macrobrachium rosenbergii. A full length cDNA of GSTT (1417 base pairs) was isolated and characterized bioinformatically. Exposure to virus (white spot syndrome baculovirus or M. rosenbergii nodovirus), bacteria (Aeromonas hydrophila or Vibrio harveyi) or heavy metals (cadmium or lead) significantly increased the expression of GSTT (P < 0.05) in hepatopancreas. Recombinant GST-θ with monochlorobimane substrate had an optimum activity at pH 7.5 and 35 °C. Furthermore recombinant GST-θ activity was abolished by the denaturants triton X-100, Gua-HCl, Gua-thiocyanate, SDS and urea in a dose-dependent manner. Overall, the results suggest a potential role for M. rosenbergii GST-θ in detoxification and possibly conferring immune protection.     

Diversity, host, and habitat specificity of oomycete communities in declining reed stands (Phragmites australis) of a large freshwater lake

We studied the diversity of oomycetes in a declining reed belt (Phragmites australis) of Lake Constance, Germany, using conventional baiting with specific reed and standard oak baits, accompanied by molecular techniques. Apart from an Aphanomyces sp. and a Phytophthora sp., baiting from reed rhizosphere samples from flooded, as well as drier, littoral sites revealed only Pythium spp. A total of 67 oomycete isolates was classified according to PCR-RFLP banding patterns and ITS sequencing, and 18 different sequence types could be separated. The majority of these seemed previously unknown species, as indicated by the degree of similarity to those deposited in nucleotide databases. Species communities in both flooded and drier habitats or both reed and oak baits clearly differed from one another, and only few species occurred in both dry and flooded sites, or in both oak and reed baits. A frequently occurring group of related Pythium species appeared to be specifically associated with reed, and these were the only species that proved pathogenic towards this host in vitro. Our study proved that unexplored natural ecosystems harbour diverse communities of oomycete species with specific habitat and host preferences within close-by, but ecologically contrasting, sites. Among the species isolated, those associated with the predominating plant might accumulate and thus may be reed pathogens of considerable importance.     

林龄对刺槐人工林土壤团聚体、有机碳和细菌群落的影响

探究刺槐人工林土壤团聚体、有机碳及细菌群落的变化特征,可为提升土壤质量与功能提供理论依据。以黄土高原沟壑区不同林龄刺槐人工林(8年生、18年生和30年生)和刚撂荒的农耕地(CK)为研究对象,利用最适湿度筛分法获得不同粒径团聚体,测定其有机碳含量和细菌群落结构特征。结果表明:(1)各林龄土壤团聚体均以>0.25 mm粒径为主,含量为92.74%-95.78%。土壤团聚体稳定性随着林龄的增加显著提高,以30年生林地最高,团聚体平均重量直径(MWD)、几何平均直径(GMD)分别较CK显著增加48.19%和91.38%(P<0.05)。(2)各林龄均以<0.25 mm粒径有机碳含量最高。土壤有机碳含量和>2 mm粒径团聚体有机碳含量均随着林龄的增长而显著增加。(3)团聚体细菌群落主要由变形菌门(Proteobacteria)、放线菌门(Actinobacteria)和酸杆菌门(Acidobacteria)组成。随着林龄的增加,各粒径团聚体中放线菌门相对丰度先降低后增加,18年生林地最低。酸杆菌门变化趋势与放线菌门相反。变形菌门无明显变化。土壤有机碳、pH、全氮和全磷是影响团聚体细菌群落的主要因素。(4)>2 mm和2-0.25 mm粒径团聚体中,己科河菌门(Rokubacteria)对团聚体稳定性影响最大,既对团聚体稳定性产生直接正效应,又通过增加有机碳的含量间接提高团聚体稳定性。在<0.25 mm粒径团聚体中,有机碳含量对其稳定性的影响最大,有机碳含量越高,团聚体稳定性越强。综上,营造刺槐人工林有利于提高土壤团聚体稳定性,促进土壤有机碳的积累,可作为该区域生态恢复的有效措施。在生长发育过程中,人工林可通过改变土壤团聚体细菌群落和有机碳含量从而影响团聚体的稳定性。     

Effects of eyestalk ablation on growth and food conversion efficiency of the freshwater prawn Macrobrachium lanchesteri (de Man)

Bilateral eyestalk ablation in the freshwater prawn Macrobrachium lanchesteri results in high mortality, while unilateral eyestalk ablated prawns exhibited a high survival rate. There was marked increase in the growth of bilateral eyestalk-ablated prawns (47.70 mg/prawn) as compared to those that were unilaterally ablated (19.19 mg/prawn).     

Diversity of sulfate-reducing bacteria from an extreme hypersaline sediment, Great Salt Lake (Utah)

The diversity of sulfate-reducing bacteria (SRB) inhabiting the extreme hypersaline sediment (270 g L(-1) NaCl) of the northern arm of Great Salt Lake was studied by integrating cultivation and genotypic identification approaches involving PCR-based retrieval of 16S rRNA and dsrAB genes, the latter encoding major subunits of dissimilatory (bi) sulfite reductase. The majority (85%) of dsrAB sequences retrieved directly from the sediment formed a lineage of high (micro) diversity affiliated with the genus Desulfohalobium, while others represented novel lineages within the families Desulfohalobiaceae and Desulfobacteraceae or among Gram-positive SRB. Using the same sediment, SRB enrichment cultures were established in parallel at 100 and at 190 g L(-1) NaCl using different electron donors. After 5-6 transfers, dsrAB and 16S rRNA gene-based profiling of these enrichment cultures recovered a SRB community composition congruent with the cultivation-independent profiling of the sediment. Pure culture representatives of the predominant Desulfohalobium-related lineage and of one of the Desulfobacteraceae-affilated lineages were successfully obtained. The growth performance of these isolates and of the enrichment cultures suggests that the sediment SRB community of the northern arm of Great Salt Lake consists of moderate halophiles, which are salt-stressed at the in situ salinity of 27%.     

河套平原不同深度高砷地下水硫酸盐还原菌群落分布特征及环境意义

【目的】探究不同深度的高砷含水层中硫酸盐还原菌的丰度、群落组成和多样性的差异,并结合硫酸盐硫同位素等多种水化参数,揭示不同深度高砷地下水中硫酸盐还原菌群落分布特征及其环境意义。【方法】以我国典型高砷地下水分布区河套平原为研究区,采集不同深度含水层中的高砷地下水样品,测定水化参数,采用qPCR对样品16S rRNA基因和dsrB基因进行定量;通过dsrB基因高通量测序对硫酸盐还原菌群落进行分析,并将dsrB基因相对丰度、群落组成及多样性与水化因子结合,进行统计学分析。【结果】基于dsrB基因的定量结果表明,浅层地下水中dsrB基因相对丰度高于深层地下水。浅层地下水中,dsrB基因相对丰度与CH₄浓度呈显著正相关,且δ³⁴S-SO₄^2–与CH₄浓度显著正相关。而深层高砷地下水中,dsrB基因相对丰度与SO₄^2–浓度、DOC浓度存在显著正相关性。高通量测序结果表明,深层地下水中硫酸盐还原菌的α多样性显著高于浅层地下水。研究区内硫酸盐还原菌可...