一株牙鲆源黏质沙雷氏菌YP1的分离鉴定及致病性分析

黏质沙雷氏菌(Serratia marcescens)是引起人类、动物及植物感染的重要条件致病菌,但其作为鱼类致病菌却鲜有报道。【目的】本研究以从患病牙鲆(Paralichthys olivaceus)病灶处分离的一株黏质沙雷氏菌YP1为研究对象,分析黏质沙雷氏菌对鱼类的致病性及对疾控的影响。【方法】利用形态学、分子生物学及生理生化实验综合鉴定菌株YP1;利用菌株YP1进行人工感染实验、组织病理实验及药敏试验,研究其感染症状、组织病理学、毒力和药物敏感性。【结果】分离自患病牙鲆体表溃疡病灶处的菌株YP1鉴定为黏质沙雷氏菌。感染实验结果显示,牙鲆和斑马鱼的半数致死量(LD₅₀)分别为3.44×10⁷CFU/g和6.28×10⁵CFU/g,除牙鲆外菌株YP1对其他鱼类也具有高致病性;菌株YP1主要导致牙鲆腹水,同时伴有呼吸急促、摄食减弱、脱肛、白便、鳃缺血及多脏器膨大出血等症状,并随着感染时间的延长对脏器损伤呈加重趋势。病理组织切片结果显示,菌株YP1对牙鲆鳃、肠、肝、脾、肾、心均造成损伤。药敏试验结果表明,YP1对左氧氟沙星、诺氟沙星等14种药物敏感;但对氨苄西林、头孢拉定等19种药物具有耐药性。【结论】本研究结果证实了黏质沙雷氏菌是能导致牙鲆腹水病的一种病原菌,同时对其他鱼类也具高致病性,为该菌感染鱼类导致疾病的检测、鉴别和防治提供科学依据。     

4种烟碱受体激动剂对松材线虫的毒杀活性

【目的】探究4种烟碱受体激动剂对松材线虫的杀虫作用。【方法】采用浸渍法以及马尾松松枝水培试验分别进行杀线虫活性测定,并测定4种烟碱受体激动剂水溶液对松材线虫体内三磷酸腺苷酶（ATP）、过氧化氢酶（CAT）、乙酰胆碱酯酶（AChE）、超氧化物歧化酶（SOD）的影响。【结果】盐酸左旋咪唑、盐酸四咪唑、烟碱、酒石酸噻吩嘧啶4种烟碱受体激动剂对松材线虫有良好的毒杀活性,4种药物水溶液浸渍处理松材线虫48 h,半致死浓度LC₅₀值分别为0.09、0.31、0.08、0.14 mg·mL^-1;马尾松水培试验处理12 d后结果显示,盐酸左旋咪唑、盐酸四咪唑、烟碱、CK处理组松枝均不发病,病情指数为0,酒石酸噻吩嘧啶处理组部分枝叶变黄,病情指数为31%,“CK+线虫”松枝枝叶完全变黄萎蔫,病情指数为100%;4种烟碱受体激动剂水溶液浸渍处理48 h后松材线虫体内ATP、CAT、AChE、SOD酶活性均低于对照组（P<0.05）。【结论】盐酸左旋咪唑、盐酸四咪唑、烟碱、酒石酸噻吩嘧啶对松材线虫有良好的防治潜力,具有进一步深入研究和开发利用的前景。     

松材线虫和拟松材线虫体内细菌的透射电镜观察及分离鉴定

【目的】松材线虫是松树萎蔫病的病原,拟松材线虫在形态等方面与松材线虫极其相似。关于两种线虫与细菌的研究多集中于体表伴生细菌。本文要揭示松材线虫和拟松材线虫体内是否存在细菌。【方法】对松材线虫和拟松材线虫进行透射电镜观察;并采用1%升汞和抗菌素混合液对两种线虫虫体进行体表消毒后研磨,制备悬浮液涂布NA平板;通过生理生化测定和16S rDNA序列分析鉴定细菌种类。【结果】松材线虫和拟松材线虫透射电镜照片显示在两种线虫肠道内均发现细菌;体表无菌的松材线虫和拟松材线虫共分离到3株体内细菌;这3株细菌分别属于寡养单胞菌属(Stenotrophomonas)和爱文氏菌属(Ewingella)。【结论】松材线虫和拟松材线虫体内均存在细菌;这些细菌对这两种线虫可能具有一定的生理生态作用。本文是松材线虫和拟松材线虫体内存在细菌的首次报导。     

一株耐碱变形假单胞菌ZY-3的鉴定及其脱氮特性

【背景】目前缺少具有高效脱氮能力、较高生物安全性、能处理高碱含氮污水的好氧反硝化菌株,难以使用生物方法处理高碱性的工业、养殖废水。【目的】对前期于佛山市一水产养殖池塘底泥中分离得到的耐碱高效好氧反硝化细菌ZY-3进行研究,期望获得一株能用于不同酸碱环境脱氮的高效、安全的好氧反硝化细菌。【方法】通过形态学、生理生化试验及16S rRNA基因序列分析方法对菌株种属进行鉴定,采用抗生素试验及斑马鱼攻毒试验进行菌株的环境生物安全性评估,利用3种含不同氮素的含氮模拟废水进行脱氮能力的测定。【结果】确定ZY-3为假单胞菌属变形假单胞菌（Pseudomonas plecoglossicida）,其对多种临床常用抗生素敏感,对水生生物的毒性低,该菌株在高浓度含氮模拟废水中以28℃、180 r/min振荡培养时,其对数期出现在4—12 h,在12 h时NH₄⁺-N、NO₃^--N和NO₂^--N的去除率分别达到94.87%、81.44%和98.02%,其pH耐受范围为6.0—10.0。【结论】得到一株安全、高效、具有广泛pH适应范围的耐碱好氧反硝化细菌P. plecoglossicida ZY-3,其在有氧条件下对3种氮素（NH₄⁺-N、NO₃^–-N、NO₂^–-N）具有快速去除能力。     

拟松材线虫纤维素酶活性与其致病性关系

【目的】研究拟松材线虫是否分泌纤维素酶以及纤维素酶与其致病力的关系。【方法】对拟松材线虫的不同致病性种群以及松材线虫的虫体蛋白提取液、分泌液的纤维素酶活性进行定量测定,比较群体间纤维素同工酶谱型差异。【结果】拟松材线虫也含有纤维素酶,并向体外分泌;且不同致病性种群的纤维素酶活性与其致病性有一定相关性,致病性越强纤维素酶活性越强。【结论】纤维素酶活性大小是导致拟松材线虫不同种群间致病性差异的重要原因,这对于全面认识松树线虫萎蔫病的致病机理有重要意义。     

大熊猫源肺炎克雷伯菌生物学特性

【背景】肺炎克雷伯菌是仅次于大肠杆菌的常见条件致病菌之一,严重时可导致大熊猫发生出血性肠炎、全身性败血症等。【目的】明确大熊猫源肺炎克雷伯菌的生物学特性,对防控该病作出科学指导。【方法】分别采用结晶紫染色法、拉丝实验、K-B纸片法和PCR技术对46株大熊猫源肺炎克雷伯菌的生物被膜形成能力、高黏性表型、耐药表型和15种常见毒力基因等生物学特性进行研究,并根据以上生物学特性选择一株可能具有致病性的分离菌pneumoniae-X-5,研究其对小鼠的致病性。【结果】46株肺炎克雷伯菌均可形成荚膜;12株为高黏性表型肺炎克雷伯菌;能形成生物被膜的菌株占比为65%(30/46);分离出的46株菌中多重耐药菌株占58%(27/46),对氨苄西林、苯唑西林、青霉素、万古霉素呈100%耐药;毒力基因检出率最高的为ureA(91.30%,42/46)。pneumoniae-X-5菌株对小鼠的LD₅₀为8.9×10⁴CFU/mL;该菌株攻毒小鼠肺泡间隔增厚,炎性细胞浸润,肝细胞变性坏死,脾充血,十二指肠黏膜上皮和固有层分离,固有层部分细胞坏死。死亡小鼠脾脏含细菌量最多,其次为肝脏。【结论】本试验阐明了部分大熊猫源肺炎克雷伯菌的多重耐药性、能形成生物被膜、具有高黏表型等病原生物学特性,为大熊猫肺炎克雷伯杆菌病的防控及临床治疗提供了科学依据。     

植物根际益生细菌代表性菌株贝莱斯芽孢杆菌FZB42对松材线虫的抑杀性

[目的]由松材线虫导致的松树萎蔫病是松树的毁灭性病害,也是我国最主要的林业病害之一.本研究测评了在农业上广泛使用的、我国微生物肥料行业主要菌种资源之一——贝莱斯芽孢杆菌,对松材线虫的潜在抑杀性能.[方法]选用贝莱斯芽孢杆菌的代表性菌株FZB42为材料,测定对不同条件下的菌液上清、不同菌株的菌液上清、细菌素plantaz...     

一株海洋来源铜绿假单胞菌Gxun-7角蛋白酶基因的克隆、表达及重组酶酶学性质

【背景】角蛋白酶是一类特异性降解角蛋白的水解酶,在动物饲料、生物肥料、医学、洗涤、制革及环境治理等方面具有重要的应用潜力。【目的】对前期从海洋环境筛选出的一株铜绿假单胞菌Gxun-7的角蛋白酶基因进行克隆、表达,并探究重组酶酶学性质,为角蛋白酶在工业生产中的应用奠定基础。【方法】以铜绿假单胞菌Gxun-7基因组推定的角蛋白酶基因为基础,设计引物克隆获得角蛋白酶基因kp2,构建重组表达质粒pET22b-kp2,并转化到E. coliRosettagamiB (DE3)中进行诱导表达,同时对重组表达菌株的表达条件进行优化。利用镍柱分离纯化重组角蛋白酶并研究其酶学性质。【结果】重组角蛋白酶的分子量约为33 kDa,最适温度和pH值分别为40 ℃和8.0,在温度30-60 ℃和pH 6.5-8.0具有较好的稳定性。金属离子Co²⁺、Cu²⁺和化学试剂十二烷基磺酸钠(sodium dodecyl sulfonate,SDS)、乙二胺四乙酸(ethylenediaminetetraacetic acid,EDTA)、苯甲基磺酰氟(phenylmethylsulfonyl fluoride,PMSF)对酶活力有抑制作用,而Mg²⁺、K⁺、巯基乙醇和二硫苏糖醇(dithiothreitol,DTT)对酶活力有促进作用。重组角蛋白酶具有良好的耐盐性,在12.5%的NaCl作用下相对酶活为87.55%。以酪蛋白为底物时,酶的K_m值为60.92 mg/mL、V_max值为9.70 U/mL。【结论】海洋来源铜绿假单胞菌Gxun-7的重组角蛋白酶具有良好的温度、碱、盐稳定性,可应用于工业生产中。     

一株兼具秸秆腐解能力玉米促生菌的筛选、鉴定及发酵优化

【背景】砂姜黑土地区存在秸秆腐解缓慢、秸秆还田后作物幼苗生长不良等问题。【目的】从砂姜黑土区农田筛选一株兼具秸秆腐解能力的玉米促生菌MC29,以促进秸秆腐解和玉米作物生长。【方法】通过16S rRNA基因序列分析对该菌株进行鉴定;采用液态摇瓶及盆栽试验验证菌株实际促腐、促生能力及土壤养分的提升效果,并且探究菌株的最佳生长及产吲哚乙酸(indole-3-acetic acid, IAA)条件;采用电击转化法将绿色荧光蛋白(green fluorescent protein, GFP)基因导入菌株细胞,并通过PCR琼脂糖凝胶电泳进行验证。【结果】分离筛选的玉米促生菌鉴定为纤维化纤维微细菌(Cellulosimicrobium cellulans)。该菌株MC29羧甲基纤维素(carboxymethyl cellulose, CMC)酶活可达13.32 U/mL,产IAA量为8.63 mg/L。与对照相比,施用菌株MC29后秸秆腐解率显著提高24.8%;玉米soil and plant analyzer development (SPAD)值、植株总重、根表面积和根长分别提高7.6%、21.3%、30.9%和18.3%;土壤碱解氮含量显著提高68.1%,土壤速效磷及土壤速效钾含量分别提高5.8%及6.0%。菌株MC29最佳生长条件为pH 7.0、装液量为25/250 mL、碳源为麦芽糖、氮源为酵母粉;最佳产IAA条件为pH 7.0、装液量为50/250 mL、碳源为果糖、氮源为硝酸钾;成功构建荧光标记菌株MC29-GFP,并据此追踪到其接入砂姜黑土15 d后定殖量为2.8×10⁵-9.5×10⁵ copies/g。【结论】所筛选的纤维化纤维微细菌MC29对于指导砂姜黑土区多功能秸秆促腐菌剂、微生物菌肥的研制及提升作物产量有一定的积极意义,并为探究其在砂姜黑土中的实际应用奠定基础。     

罗氏沼虾(Macrobrachium rosenbergii)养殖池塘沉积物中细菌、硫酸盐还原菌和硫氧化菌的垂直分布特征

沉积物中微生物介导的硫循环在有机物分解和养分循环中发挥重要作用,但目前我们对水产养殖生态系统中的参与硫酸盐还原和硫氧化过程的微生物多样性及其调控机制仍知之甚少。【目的】探究硫酸盐还原菌(sulfate-reducing bacteria,SRB)和硫氧化菌(sulfur-oxidizing bacteria,SOB)的垂直分布特征及其主要的环境驱动因素。【方法】本研究利用高通量测序和荧光定量PCR (qPCR)分析了罗氏沼虾(Macrobrachium rosenbergii)养殖池塘沉积物表层(0–1 cm)、中层(10–11 cm)和底层(20–21 cm)中的细菌、SRB和SOB的丰度、多样性和群落组成。【结果】细菌(16S rRNA)、硫酸盐还原菌(dsrB)和硫氧化菌(soxB)的基因拷贝数呈现着从表层到中层急剧骤降的趋势(ANOVA,P<0.05),但中层和底层样品之间的差异却并不显著(P>0.05),以及α多样性分析显示3个群体的物种丰富度和均匀度都随深度而逐步降低,这都说明硫循环过程主要发生于沉积物的表层。γ-、δ-和β-变形菌分别是细菌、SRB和SOB的优势类群;其中SRB以Desulfobacca属和脱硫八叠球菌属(Desulfosarcina)为主,前者在表层有着最低的比重,而后者却与之相反;硫杆菌(Thiobacillus)作为细菌和SOB的优势属,更广泛地分布于中层沉积物中。RDA分析和Mantel检验揭示了影响细菌群落的主要环境因子是NO₃^–、SO₄^2–、TOC和TON,而SRB的群落变异主要是由As、TON、NO₃^–和Pb所驱动,以及SOB的群落变化则主要响应了TC、NO₂^–、NH₄⁺和TON浓度。【结论】养殖池塘底栖的细菌、SRB和SOB的丰度、多样性和群落结构的垂直分布特征可能受到多种环境因素共同的影响。     

Systematic significance of mature embryo of bamboos

The mature embryo of seven species belonging to five genera of Indian bamboos is described. In all these the basic pattern of embryo organisation is same: the scutellar and coleoptilar bundles are not separated by an internode, the epiblast is absent, the lower portion of the scutellum and the coleorhiza are separated by a cleft and the margins of embryonic leaves overlap. The features unique to fleshy fruited bamboos are: presence of a massive scutellum, the juxtaposition of plumule and radicle and the occurrence of a bud in the axil of the coleoptile. The fleshy fruit bearing bamboos should be classified into one group, the tribeMelocanneae. Evidence is provided to recognise additional groups in the subfamilyBambusoideae.     

Taxonomic status of the species of the green algal genusNeochloris

Komárek has recently reviewed the various species assigned to the green algal genusNeochloris Starr (Chlorococcales, Chlorococcaceae) and removed those with uninucleate vegetative cells to a new genus,Ettlia. Watanabe & Floyd, unaware ofKomárek's work, also reviewed the species ofNeochloris and distributed them among three genera—Neochloris, Chlorococcopsis gen. nov., andParietochloris gen. nov.—on the basis of details of the covering of the zoospore and the arrangement of the basal bodies of the flagellar apparatus. This paper reconciles these two treatments and makes additional recommendations at the ranks of genus, family, order, and class.     

Phylogeny of cardinalfishes (Teleostei: Gobiiformes: Apogonidae) and the evolution of visceral bioluminescence

The cardinalfishes (Apogonidae) are a diverse clade of small, mostly reef-dwelling fishes, for which a variety of morphological data have not yielded a consistent phylogeny. We use DNA sequence to hypothesize phylogenetic relationships within Apogonidae and among apogonids and other acanthomorph families, to examine patterns of evolution including the distribution of a visceral bioluminescence system. In conformance with previous studies, Apogonidae is placed in a clade with Pempheridae, Kurtidae, Leiognathidae, and Gobioidei. The apogonid genus Pseudamia is recovered outside the remainder of the family, not as sister to the superficially similar genus Gymnapogon. Species sampled from the Caribbean and Western Atlantic (Phaeoptyx, Astrapogon, and some Apogon species) form a clade, as do the larger-bodied Glossamia and Cheilodipterus. Incidence of visceral bioluminescence is found scattered throughout the phylogeny, independently for each group in which it is present. Examination of the fine structure of the visceral bioluminescence system through histology shows that light organs exhibit a range of morphologies, with some composed of complex masses of tubules (Siphamia, Pempheris, Parapriacanthus) and others lacking tubules but containing chambers formed by folds of the visceral epithelium (Acropoma, Archamia, Jaydia, and Rhabdamia). Light organs in Siphamia, Acropoma, Pempheris and Parapriacanthus are distinct from but connected to the gut; those in Archamia, Jaydia, and Rhabdamia are simply portions of the intestinal tract, and are little differentiated from the surrounding tissues. The presence or absence of symbiotic luminescent bacteria does not correlate with light organ structure; the tubular light organs of Siphamia and chambered tubes of Acropoma house bacteria, those in Pempheridae and the other Apogonidae do not.     

Host plants of someRhinanthoideae (Scrophulariaceae) of eastern North America

Twenty three species in 11 genera were examined in the field to determine hosts. OnlyStriga asiatica andSeymeria cassioides have a narrow host range being restricted to grasses and pines, respectively. These are the only species which cause pronounced and sometimes serious host damage. The other species attach to a great diversity of hosts.     

Apoptotic Regulation of Caspase Activity

Intracellular cysteine aspartate-specific proteases (caspases) play both signaling and effector roles in realizing the program of cell death. Caspases function as proteolytic cascades unique for each cell type and signal triggering apoptosis. All parts of the proteolytic cascades are duplicated and controlled by feedback signals. Amplification cycles between pairs of caspases (the third and the sixth, the ninth and the third, the twelfth and the sixth, and others) help multiply the initial apoptotic signal. The presence of physiological inhibitors of apoptosis that directly interact with caspases creates a multilevel regulatory network of apoptosis in cell. The caspase proteolytic cascades are also regulated by sphingolipid secondary messengers, among them ceramide, sphingosine, and their phosphates. Moreover, an association of the caspase signaling with ubiquitin-dependent proteolysis is shown in cells. In particular, the use of extracellular activators and inhibitors of caspases allows irreversible activation of apoptosis in tumor cells or the prevention of apoptosis in cortical neurons under neurodegenerative diseases.     

Analysis of twin of eyeless regulation during early embryogenesis in Drosophila melanogaster

The Drosophila Pax6 homolog twin of eyeless (toy) is so far the first zygotically expressed gene involved in eye morphogenesis in Drosophila. The study of its expression during embryogenesis is therefore informative of the initial events of eye development in Drosophila. We have analyzed how the initial expression domain of toy at cellular blastoderm is regulated. We show that the three maternal patterning systems active in the cephalic region (the anterior, terminal and dorsal-ventral systems) cooperate with zygotically activated gap genes to shape the initial expression domain of toy. Whereas Bicoid, Dorsal and Torso signaling synergistically act as activators, Hunchback, Knirps and Decapentaplegic act as repressors.     

A molecular phylogeny of Hebeloma species from Europe

In order to widen the scope of existing phylogenies of the ectomycorrhizal agaric genus Hebeloma a total of 53 new rDNA ITS sequences from that genus was generated, augmented by sequences retrieved from GenBank, and analysed using Bayesian, strict consensus and neighbour joining methods. The lignicolous Hebelomina neerlandica, Gymnopilus penetrans, and two species of Galerina served as outgroup taxa. Anamika indica, as well as representatives of the genera Hymenogaster and Naucoria, were included to test the monophyly of Hebeloma, which is confirmed by the results. Hebeloma, Naucoria, Hymenogaster and Anamika indica cluster in a strongly supported monophyletic hebelomatoid clade. All trees largely reflect the current infrageneric classification within Hebeloma, and divide the genus into mostly well-supported monophyletic groups surrounding H. crustuliniforme, H. velutipes, H. sacchariolens, H. sinapizans, and H. radicosum, with H. sarcophyllum being shown at an independent position; however this is not well supported. The section Indusiata divides with strong support into three groups, the position of the pleurocystidiate Hebeloma cistophilum suggests the possible existence of a third subsection within sect. Indusiata. Subsection Sacchariolentia is raised to the rank of section.     

New species of Peruvian Orchidaceae III

Sixteen new species are proposed in the generaAckermania, Dressleria, Epidendrum, Maxillaria, Oncidium, Rodriguezia, Sigmatostalix, andTrigonidium. All new species are illustrated.Maxillaria vittariifolia L. O. Williams is newly recorded for Peru. A key is provided forTrigonidium of Peru.Trigonidium loretoense Schltr. andT. peruvianum Schltr. are lectotypified.     

Interaction of Synthetic Analogues of Natural Chalcones and Flavones with Guanidine

The reaction of chalcones and flavones with guanidine in ethanol results in 2-(2-aminopyrimidinyl-4)-phenols.