养殖黄颡鱼(Pelteobagrus fulvidraco)爱德华氏菌(Edwardsiella ictaluri)的分离鉴定与生物学特性研究

从四川眉山与新津两地患红头病的养殖黄颡鱼体内分离到2株优势菌(CHNYC001与CHNYC002),以腹腔注射与浸泡的方式进行人工感染试验,证实其为养殖黄颡鱼红头病的病原菌。根据分离菌株的形态、生理生化特性,结合16SrDNA序列测定(GenBank登录号分别为FJ766524、FJ766525)与系统发育分析,将其鉴定为鮰爱德华氏菌(Edwardsie lla ictaluri)。分离菌株最适生长温度为20—30℃,最适生长pH为7.0,在含盐量2%以上的培养基上不生长,对氟苯尼考、强力霉素、洛美沙星等敏感,对乙酰螺旋霉素、磺胺甲基异噁唑和麦迪霉素等不敏感。临床病理特征分析发现,养殖黄颡鱼感染爱德华氏菌临床上分为急性与慢性两种类型,急性型主要表现为败血症的病变特征,慢性型表现为头顶出血,发红与溃疡的病变特征。     

养殖黄颡鱼(Pelteobagrus fulvidraco) 鮰爱德华氏菌(Edwardsiella ictaluri)的分离鉴定与生物学特性研究

从四川眉山与新津两地患红头病的养殖黄颡鱼体内分离到2株优势菌(CHNYC001与CHNYC002),以腹腔注射与浸泡的方式进行人工感染试验,证实其为养殖黄颡鱼红头病的病原菌。根据分离菌株的形态、生理生化特性,结合16SrDNA序列测定(GenBank登录号分别为FJ766524、FJ766525)与系统发育分析,将其鉴定为鮰爱德华氏菌(Edwardsie lla ictaluri)。分离菌株最适生长温度为20—30℃,最适生长pH为7.0,在含盐量2%以上的培养基上不生长,对氟苯尼考、强力霉素、洛美沙星等敏感,对乙酰螺旋霉素、磺胺甲基异噁唑和麦迪霉素等不敏感。临床病理特征分析发现,养殖黄颡鱼感染爱德华氏菌临床上分为急性与慢性两种类型,急性型主要表现为败血症的病变特征,慢性型表现为头顶出血,发红与溃疡的病变特征。     

瓦氏黄颡鱼(Pelteobagrus vachellii)Toll样受体2(TLR2)基因克隆及免疫功能

为探究Toll样受体2(Toll-like receptor 2,TLR2)及下游免疫分子对瓦氏黄颡鱼(Pelteobagrus vachellii)机体的保护作用,本研究采用RT-PCR及RACE法获得瓦氏黄颡鱼TLR2全长c DNA(2611bp),编码789个氨基酸残基,含有10个富含亮氨酸的重复序列(Leucine Rich Repeat,LRR)和Toll/IL-1R(Toll/IL-1 Receptor Domain,TIR)同源区结构域,属I型跨膜受体。序列同源性比对发现,瓦氏黄颡鱼TLR2 c DNA与斑点叉尾、鲤及虹鳟的同源性分别为78%、62%及49%。系统进化树分析表明,瓦氏黄颡鱼TLR2与斑点叉尾聚为一支。q RT-PCR分析表明,TLR2 m RNA在检测的组织中均有表达,且在头肾和脾脏中表达水平显著高于其他组织(P0.05)。嗜水气单胞菌感染能显著上调瓦氏黄颡鱼肝脏、头肾及脾脏中TLR2 m RNA表达(P0.05),分别在24h、48h及12h达到最大值。头肾中的TLR 2信号通路下游的髓样分化因子、半胱氨酸蛋白酶8、核转录因子kappa B、肿瘤坏死因子α、白细胞介素1βm RNA均显著上升(P0.05),分别在24h、12h、48h,48h和48h达到最大值。结果表明,嗜水气单胞菌感染激活了TLR2信号通路,通过上调表达,肿瘤坏死因子α,白细胞介素1β等。本研究表明,TLR2在瓦氏黄颡鱼抵御嗜水气单胞菌侵染的过程中发挥了重要的免疫作用。     

牙鲆(Paralichthys olivaceus)TLR21 基因在迟缓爱德华氏菌(Edwardsiella tarda)感染后的表达特征

应用荧光定量PCR技术, 检测了TLR21 基因在牙鲆(Paralichthys olivaceus)感染迟缓爱德华 氏菌(Edwardsiella tarda)后, 在0 h、1 h、3 h、6 h、12 h、1 d、3 d、6 d 后, 在心脏、肝脏、脾脏、 头肾、鳃、小肠、肌肉和血的时空表达特征, 并探讨了它们与牙鲆先天免疫反应的关系。结果表明, 大 多组织在感染病原6 h 后TLR21 基因表达明显上调, 尤其是头肾和小肠。头肾6 h 的表达量达到了 对照组的59.3 倍, 小肠6 h 的表达量为对照组的38.6 倍。迟缓爱德华氏菌感染引起牙鲆体内各组织 中TLR21 的上调表达和变化, 为研究牙鲆对迟缓爱德华氏菌的防御机制提供了理论依据。     

黄颡鱼(Pelteobagrus fulvidraco)“红头病”病原菌迟钝爱德华氏菌(Edwardsiella tarda)的分离及鉴定

用ATB微生物自动鉴定系统对分离自湖南湘潭地区人工养殖的患“红头病”的黄颡鱼体内的2株细菌（即HN004和HN005）进行了鉴定,发现它们的生理生化特征完全相同,均为革兰氏阴性短杆菌、接触酶阳性、吲哚阳性、H2S阳性;氧化酶阴性、V．P测定为阴性,与迟钝爱德华氏菌的表型特征非常相似。为进一步确定2株菌的分类学地位,测定了其16S rRNA基因序列,分析了相关细菌相应序列的同源性,构建分子系统发育树。结果表明,2菌株的序列完全一致,与迟钝爱德华氏菌的亲缘关系最近,相似性为99．0％。综合上述结果,2菌株可鉴定为迟钝爱德华氏菌（Edwardsiella tarda）。     

瓦氏黄颡鱼(Pelteobagrus vachelli)卵黄蛋白原的纯化、性质鉴定及ELISA检测方法的建立

腹腔注射17β-雌二醇(E2),使瓦氏黄颡鱼雄鱼在7天内产生卵黄蛋白原(Vtg)。采用凝胶过滤和离子交换两种层析技术,从E2诱导的雄性瓦氏黄颡鱼血浆中分离、纯化出Vtg,采用糖、磷、脂蛋白染色技术证明分离、纯化的蛋白为Vtg,该Vtg在非变性条件下分子量约为240kDa,在SDS变性条件下分子量约为143kDa。纯化的瓦氏黄颡鱼Vtg经检测显示可能含有类胡萝卜素,但没有二硫键,对热相对稳定。利用纯化的瓦氏黄颡鱼Vtg,制备了兔抗瓦氏黄颡鱼Vtg多克隆抗血清。用双向免疫扩散法测得抗血清的纯度较高,效价为1︰32;Western blotting检测显示抗血清的特异性较好。以瓦氏黄颡鱼Vtg多克隆抗血清为抗体,以纯化的瓦氏黄颡鱼Vtg为抗原,建立了间接酶联免疫吸附反应(ELISA)方法检测瓦氏黄颡鱼体内Vtg的含量,标准曲线线性部分的线性方程为y=0.099x+0.4529(R2=0.9327),该方法检测的灵敏度为15.6ng/ml,工作范围为31.2—4000ng/ml,在此范围内,标准曲线具有良好的线性。     

美人鱼发光杆菌杀鱼亚种感染卵形鲳鲹的病理学观察

病原菌美人鱼发光杆菌杀鱼亚种(Photobacterium damselae subsp. piscicida)分离自发病的卵形鲳鲹(Trachinotus ovatus), 本实验利用腹腔注射和浸泡的感染途径, 观察卵形鲳鲹发光杆菌病的病理变化。感染发病鱼呈现急性和慢性临床症状, 主要急性症状为鳃盖周围轻微出血, 腹腔积水和内脏器官多灶性坏死; 主要慢性症状为脾脏、肾脏和心脏内能观察到直径为0.5~1.0 mm 的白色粟米样结节; 利用光学显微镜和透射电镜观察的组织病理显示: 急性病理症状主要表现为鳃、肝和肾发生变性及凝固性坏死, 肾管微绒毛紊乱, 线粒体的嵴脱落, 脾淋巴细胞增生, 核染色质边集, 心肌细胞发生多灶性坏死,线粒体增生, 肠道的病变较轻微。慢性病理症状主要表现为鳃丝上皮细胞坏死, 脾淋巴细胞线粒体、高尔基体和内质网溶解, 肾小管上皮细胞的微绒毛脱落, 心肌纤维Z 带排列紊乱, 线粒体变性, 肝脏、肾脏、脾脏、心脏和肠道出现典型的肉芽肿病变。相比之下, 脾脏、肾脏和心脏的病变是所有器官中最严重的。     

GFP标记的迟缓爱德华菌感染诸氏鲻虾虎鱼后组织分布研究

为探讨迟缓爱德华菌(Edwarsiellatarda)入侵途径,建立感染模型,作者通过电转化法构建GFP标记的迟缓爱德华菌EtMc1512(质粒PMDpp-EGFP),实验设立浸泡组、腹腔注射组和肌肉注射组,感染后采集各组实验诸氏鲻虾虎鱼(Mugilogobius chulae)血液、鳃、肝脏、肠、肌肉,培养法统计分析各组织中的荧光细菌数;浸泡组取样时间为0、2、4、6、8、12、24 h,腹腔注射组和肌肉注射组取样时间为6、12、24、48、72、96h。结果显示,构建的EtMc1512-GFP具有较强荧光,GFP标记前后菌株毒力基因(citC、mukF、esrB、katB、fimA、gadB)检测结果均为阳性。浸泡感染后实验鱼各组织内的荧光菌随时间表现为先升后降的趋势,最高菌量出现在肠道(2.51×106CFU/g),其次为鳃(4.19×104CFU/g)、血液(1.65×104CFU/g),肠道荧光菌显著高于其他组织(P0.05);腹腔注射感染后肝脏(4.55×106CFU/g)和血液(4.65×106CFU/g)菌量最高;肌肉注射感染后肌肉在48h首先检出荧光菌,血液(2.93×104 CFU/g)菌量最高。结果表明,肠道、肝脏和肌肉分别是迟缓爱德华菌浸泡感染、腹腔注射感染和肌肉注射感染诸氏鲻虾虎鱼的主要组织器官,在自然条件下迟缓爱德华菌经口感染诸氏鲻虾虎鱼风险较高。     

不同水温下儿茶素EGCG在黄颡鱼(Pelteobagrus vachelli)体内代谢动力学和肝消除研究

采用反相高效液相色谱-紫外检测法测定了黄颡鱼血浆、肝脏中儿茶素EGCG含量,并研究了不同水温(10℃和20℃)下EGCG在黄颡鱼体内代谢动力学和肝组织消除特征。结果表明,高温下黄颡鱼腹腔注射EGCG后体内吸收较快,T1/2ka为0.21h,T1/2α为2.07h,T1/2β为22.89h,AUC为5195.41μg·h/ml,Tp为0.95h,Cmax为257.75μg/ml;低温下分别为5.67h、9.53h、207.49h、17283.03μg·h/ml、12.56h和117.16μg/ml。同时在0.5、1、2、4、8、12、24、48、96h各时间点检测了肝脏组织中EGCG浓度,并计算得出高温下EGCG在肝组织中的T1/2β为24.15h,低温下肝组织中T1/2β为64.17h。对不同水温下代谢动力学和肝组织消除规律的研究表明,EGCG在黄颡鱼体内吸收、分布和消除速度与水温关系密切,高温下均较低温下迅速。     

南方鲇(Silurus meridionalis)豚鼠气单胞菌溶血素的分离、纯化与致病性研究

采用硫酸铵盐析、DEAE-Sephadex A50凝胶层析法对豚鼠气单胞菌的溶血素进行分离、纯化,获得分子量为32.2kDa的单一多肽溶血素,测定溶血价为25。将该溶血素倍比稀释后梯度接种南方鲇,研究其对南方鲇的致病性和组织病理损伤情况。结果表明:豚鼠气单胞菌溶血素具有较强的毒力,对南方鲇的半数致死剂量(LD50)为0.29mg蛋白/kg体重;病鱼表现为体表褪色,腹部膨大,肝、脾、肾肿大,肝、脾淤血、出血明显;胃肠道内充有大量粘液,黏膜充血、出血;组织病理学表现为骨骼肌水肿、变性,肝细胞变性、坏死,血窦扩张淤血,肾小管上皮细胞变性、坏死,脾脏淋巴细胞减少,胃肠道黏膜上皮变性、坏死,脱落。     

Microbial Biomass and Organic Nutrients in the Deep-Sea Sediments of the Central Indian Ocean Basin

In order to assess the impact of deep-sea mining on the in situ benthic life, we measured the microbial standing stock and concentration of organic nutrients in the deep-sea sediments of the Central Indian Ocean Basin in the Indian pioneer area. Sediments were collected using box core and grab samples during September 1996. The total bacterial numbers ranged from 10 10 -10 11 cells per g -1 dry weight sediment. There was a marginal decrease in the number of bacteria from surface to 30 cm depth, though the subsurface section registered a higher number than did the surface. The highest numbers were encountered at depths of 4-8 cm. The retrievable number of bacteria were two orders less in comparison with the direct total counts of bacteria. An almost homogeneous distribution of bacteria, total organic carbon, living biomass, and lipids throughout the depth of cores indicates active microbial and benthic processes in the deep sea sediments. On the other hand, a uniform distribution of total counts of bacteria, carbohydrates, and total organic carbon in all the cores indicates their stable nature and suggests that they can serve as useful parameters for long-term monitoring of the area after the benthic disturbance. Further studies on temporal variability in this region would not only verify the observed norms of distribution of these variables but would also help to understand restabilization processes after the simulated benthic disturbance.     

海上大直径钢管桩打桩过程中桩周土强度弱化研究

海上大直径钢管桩打桩过程中,桩周土体受到强烈扰动而发生强度弱化,掌握桩周土体强度弱化规律对于准确预测打桩过程、保证工程安全具有重要意义。为研究土体强度弱化规律,开展了环剪试验模拟打桩对桩周土体的扰动,测试土体强度随剪切速率的变化规律,建立了描述土体强度弱化规律的拟合公式,引入到打桩分析软件中。研究结果表明：土体的强度折减程度不仅与土体本身的性质有关还受到土体的埋深和剪切速率的影响,埋深越深土体强度折减程度越低,剪切速率越高土体强度折减越高,在打桩分析中可采用这里推荐的线性折减方法来模拟不同深度处土体强度的折减规律。     

Geoacoustic Inversion via Genetic Algorithm and Its Application to Manganese Sediment Identification

An acoustic inversion method using a wide-band signal and two near field receivers is proposed and applied to multiple layered seabed models including a manganese sediment. The inversion problem can be formulated into a probabilistic model comprised of signals, a forward model, and additive noise. The forward model simulates wide-band signals, such as chirp signals, and is chosen to be the source-waveletconvolution plane wave modeling method. The wavelet matching technique, using weighted least-squares fitting, estimates the sediment sound-speed and thickness on which determination of the possible numerical ranges for a priori uniform distribution is based. The genetic algorithm is applied to a global optimization problem to find a maximum a posteriori solution for determined a priori search space. Here the object function is defined by an L 2 norm of the difference between measured and modeled signals. Not only the marginal pdf but also its statistics are calculated by numerical evaluation of integrals using the samples selected during importance sampling process of the genetic algorithm.     

Phylogenetic diversity and phenotypic characterization of cultivable bacterioplankton isolated from polar oceans

A set of 27 marine planktonic bacteria isolated from the polar regions was characterized by 16S rDNA sequencing and physiological and biochemical testing. More than half of these bacteria were positive for caseinase, gelatinase and 13-glucosidase, and could utilize glucose, maltose or malic acid as carbon source for cell growth. Twelve isolates expressed nitrate reduction activities. Except for one antarctic isolate BSwlO175 belonging to Actinobacteria phylum, these isolates were classified as γ-Proteobacteria, suggesting that γ-Proteobacteria dominated in cultivable marine bacterioplankton at both poles. Genus Pseudoalteromonas was the predominant group in the Chukchi Sea and the Bering Sea, and genus ShewaneUa dominated in cultivable bacterioplankton in the Prydz Bay. With sequence similarities above 97%, genus Psychrobacter was found at both poles. These 27 isolates were psychrotolerant, and significant 16S rDNA sequence similarities were found not only between arctic and antarctic marine bacteria （ 〉 99% ）, but also between polar marine bacteria and bacteria from other aquatic environments （ ≥ 98.8% ）, including temperate ocean, deep sea, pond and lake, suggesting that in the polar oceans less temperature-sensitive bacteria may be cosmopolitan and have a bipolar, even global, distribution at the species level.     

Deep-Sea Sediment Compression Curves: Some Controlling Factors, Spurious Overconsolidation, Predictions, and Geophysical Reproduction

Oedometer tests have been carried out on 70 undisturbed surficial clays (at approximately 250 mm below the mudline), mostly collected by free-fall corers from sites widely scattered throughout the deep-sea North Atlantic. Acoustic measurements were also made, initially on contiguous samples and ultimately on the same sample using a geophysically instrumented oedometer which also collected electrical resistivity data. Apart from those quiescent areas below the carbonate compensation depth, such as north of the West Indies where very fine clays exist, most of the samples are silty clays whose geotechnical-geophysical properties are dependent on the type of clay minerals present (and their ability to take in moisture), the sand-size fraction, and the quantity of carbonate present. Thus the pure clays have high compressibilities which decrease on the addition of coarse particles, while the converse is true for the acoustic parameters, these increasing with the sand fraction. Using the notion of the intrinsic compression line for all samples, and comparison to it of the measured compression curves, it is clear that, contrary to some previously held ideas, most deep-sea clays are normally consolidated; the addition of carbonate has the effect of creating an open, stronger sediment skeleton. Interestingly, where information is available, the variation with depth of a sample's acoustic velocity follows the void ratio pressure relationship of the compression curve. This allows the construction of an in-situ sediment compression curve using the in-situ geophysical observations.     

INFORMATION FOR AUTHORS

<正>Acta Oceanologica Sinica(AOS)is a comprehensive academic journal edited by the Editorial Committee of Acta Oceanologica Sinica and is designed to provide a forum for important research papers of the marine scientific community which reflect the information on a worldwide basis.The journal publishes scholarly papers on marine science and technology,including physics,chemistry,biology,     

Estimation of Sediment Properties during Benthic Impact Experiments

Sediment properties, such as water content and density, have been used to estimate the dry and wet weights, as well as the volume of sediment recovered and discharged, during benthic impact experiments conducted in the Pacific and Indian Oceans. The estimates show that the weights of dry (355-1332 t) and wet sediment (1651-4888 t) recovered during the experiments, are not only a function of the total duration and distance covered during the experiment, but also depend on water content and density of the sediment. Estimation of dry sediment and its volumetric ratio in the discharge, are key parameters for calculating the discharged volume. The estimated volume of wet sediment recovered (1427-4049 m 3 ) as well as discharged (2693-6951 m 3 ) during the experiments, provide important inputs to evaluate the sediment resettlement and migration. Using these estimates, the average depth of excavation on the seafloor, can also been calculated. However, the stages of sediment recovery and discharge are expressed differently in some of the experiments. In order to standardize these, different stages have been identified, and definitions of certain terms have been suggested, for use in the future. The methods of calculating different properties, as well as weight and volume of discharged sediment are described in the paper for use in other applications concerning deep-sea discharges.