显微观察中国荷斯坦牛瘤胃纤毛虫的物种多样性北大核心CSCD

瘤胃纤毛虫是反刍动物瘤胃微生物的重要组成部分,在维持瘤胃微生态和对宿主动物供能等方面起重要作用。本研究采集5头中国荷斯坦牛(Bos taurus)瘤胃液样品固定染色,而后通过光学显微镜计数瘤胃纤毛虫的密度和属的组成及组成比,并对其中常见属进行观察和特征描述,旨在了解中国荷斯坦牛瘤胃纤毛虫的物种多样性,为反刍动物的瘤胃内纤毛虫生物多样性研究提供资料。结果发现,中国荷斯坦牛瘤胃纤毛虫在瘤胃液中的平均密度为(3.2±3.4)×10^(8)个/L,共检出13属瘤胃纤毛虫,其中检出率最高的3个属依次为内毛属(Entodinium)、单甲属(Eremoplastron)和双毛属(Diplodinium),检出率最低的3个属依次为鞘甲属(Elytroplastron)、后毛属(Metadinium)和多甲属(Polyplastron)。内毛属是中国荷斯坦牛瘤胃液中最主要的属,其平均组成比为84.0%±10.2%,而其他属的组成比都低于5%。本研究表明,中国荷斯坦牛瘤胃液中瘤胃纤毛虫具有丰富的物种多样性。     

Specificity of Rumen Ciliate Protozoa in Cattle and Sheep*

SYNOPSIS. Six protozoa-free sheep, 3 fed alfalfa hay and 3 fed a concentrate diet, were inoculated with rumen contents from a steer fed the same alfalfa hay. All 24 species of protozoa in the inoculum became established in the sheep fed alfalfa hay, while only 9 species established in the sheep fed concentrate. Percentage species composition in the alfalfa-fed sheep was fairly similar to that of the inoculum. Rumen volumes of the alfalfa hay-fed sheep were significantly higher than those of the concentrate-fed sheep; however, fluid turnover rates were similar. Total protozoan numbers per ml of rumen contents were significantly higher in the concentrate-fed sheep, but after adjustment for rumen volume, there was no significant difference in the total number of protozoa in the rumen.     

Electron microprobe analysis of intracellular electrolytes in resting and isoproterenol-stimulated exocrine glands of frog skin

Summary In the intact, in vitro frog skin, isoproterenol (ISO) stimulates and amiloride-insensitive increase in short-circuit current (SCC) that can be localized to the exocrine glands and is associated with secretion of chloride. To determine which cells in the glands respond to stimulation we measured the intracellular electrolyte concentrations of the various cell types of the mucous and seromucous glands of the skin using freeze-dried cryosections and electron microprobe analysis. In the resting state, the various cell types of the glands have intracellular electrolyte concentrations similar to the epithelial cells of the skin. Exposure to amiloride (10^–4 m) has little effect on the concentration of Na and Cl in the cells of the glands. The effect of isoproterenol has two distinct phases. Analysis of glands in tissues frozen at the peak of the SCC response (13 min after addition of isoproterenol) shows that the only significant change is an increase in Na and Ca in a group of cells at the ductal pole of the acini of both gland types. These are termed gland cells. The duct cells and cells that secrete macromolecules did not show any significant changes at this timepoint. In the gland cells, after a one-hour exposure to isoproterenol the Na concentration is at prestimulation levels while Cl drops. There is also a smaller drop in Cl in the duct and skin epithelial cells. Ouabain, which can completely block the isoproterenol SCC response, has little short-term effect on Na and Cl in the control gland but accentuates the gain of Na and drop in Cl in the isoproterenol-treated condition. Bumetanide and, to a lesser extent, furosemide, also blocks the isoproterenol SCC response and causes a further drop in Cl. The results provide indirect evidence that a major portion of the ionic component of the gland secretion is produced by a distinct group of cells separate from those producing the macromolecular component and that the mechanism of secretion involves a Na:Cl coupled transport system linked to the activity of the basolateral Na pump.     

Distribution of Elements in Rat Peripheral Axons and Nerve Cell Bodies Determined by X-Ray Microprobe Analysis

X-ray microprobe analysis was used to determine concentrations (millimoles of element per kilogram dry weight) of Na, P, Cl, K, and Ca in cellular compartments of frozen, unfixed sections of rat sciatic and tibial nerves and dorsal root ganglion (DRG). Five compartments were examined in peripheral nerve (axoplasm, mitochondria, myelin, extraaxonal space, and Schwann cell cytoplasm), and four were analyzed in DRG nerve cell bodies (cytoplasm, mitochondria, nucleus, and nucleolus). Each morphological compartment exhibited characteristic concentrations of elements. The extraaxonal space contained high concentrations of Na, Cl, and Ca, whereas intraaxonal compartments exhibited lower concentrations of these elements but relatively high K contents. Nerve axoplasm and axonal mitochondria had similar elemental profiles, and both compartments displayed proximodistal gradients of decreasing levels of K, Cl, and, to some extent, Na. Myelin had a selectively high P concentration with low levels of other elements. The elemental concentrations of Schwann cell cytoplasm and DRG were similar, but both were different from that of axoplasm, in that K and Cl were markedly lower whereas P was higher. DRG cell nuclei contained substantially higher K levels than cytoplasm. The subcellular distribution of elements was clearly shown by color-coded images generated by computer-directed digital x-ray imaging. The results of this study demonstrate characteristic elemental distributions for each anatomical compartment, which doubtless reflect nerve cell structure and function.     

采用未培养技术对荷斯坦奶牛瘤胃细菌多样性进行初步分析

采用未培养(Culture independent)技术直接从荷斯坦奶牛瘤胃液中提取瘤胃细菌微生物混合DNA(也叫元基因组DNA),利用细菌16SrDNA通用引物27F与1492R,扩增瘤胃混合微生物的16SrDNA,根据16SrDNA序列对瘤胃细菌多样性进行初步分析。通过16SrDNA序列同源性分析,发现有多于一半以上的序列与可培养的菌株的同源性小于90%,属于不可培养的菌株。选用45条测得序列与已知序列构建系统发育树,分析结果表明,它们分属于两大类LGCGPB(the lowG CGram positivebac teria)和CFB(Cytophaga_Flexibacter $CBacteroides group),剩下的克隆尚难确定其分类地位,可能是代表新属和种的序列,这些序列已向GenBank提交并得到序列号(AY986777_AY986791)。     

Zn对细胞保护作用机理的研究

应用扫描质子微探针和同步辐射ｘ荧光分析技术测定了细胞中元素的分布和组成,为确定Zn是细胞结构成分提供了直接的实验依据.用上述核技术结合有关生化指标,分析测定了正常和损伤细胞（脂质过氧化损伤）中Fe,Zn和丙二醛、SH基含量变化的相互关系.实验结果表明,当细胞发生脂质过氧化损伤时,Fe含量和丙二醛含量同步增高,而Zn含量和SH基量则降低.给细胞补充Zn后,提高了细胞质膜中的Zn含量,SH基量也随之增加,同时丙二醛量降低.提示Zn保护细胞完整性的作用机理之一是控制脂质过氧化作用.Zn可保护膜蛋白的SH基,减少和阻止被Fe所催化的过氧化反应.     

Ciliate Protozoa in the Rumen of Kafue Lechwe, Kobus leche kafuensis, in Zambia, with the Description of Four New Species

ABSTRACT The composition of the rumen ciliate fauna in 76 Kafue lechwe inhabiting a limited area in Zambia was surveyed and five genera containing 24 species with 16 formae belonging to the family Ophryoscolecidae were identified. Four new species belonging to Diplodiniinae were recognized and described as Diplodinium lochinvarense n. sp., Diplodinium leche n. sp., Diplodinium zambiense n. sp., and Metadinium ossiculi n. sp. In addition, Ostracodinium gracile form fissilaminatum Dogiel, 1932 was found for the second time and described as Metadinium fissilaminatum n. comb. The species composition was fairly unusual. Seven of the species have been found only in African wild antelopes and these species were found more frequently than cosmopolitan species. There was no evidence of isotrichid species. The average density of ciliates per 1 ml of the rumen fluid was 25.7 times 10⁴, and the number of ciliate species per head of host was 10.8.     

Electron microscopy study of intrahepatic ultrasmall superparamagnetic iron oxide kinetics in the rat. Relation with magnetic resonance imaging.

Ultrasmall superparamagnetic iron oxide (USPIO) contrast agents for use in magnetic resonance imaging (MRI) are currently undergoing clinical evaluation. However, the images observed and the kinetic profiles obtained differ from one agent to another. In this study, BD IX rats received an intravenous penis injection of the USPIO contrast agents AMI-HS and AMI-227. A cytologic study of the liver was performed, and the data obtained were compared with those of MRI. Images acquired in light microscopy, transmission electron microscopy, microanalysis and electron diffraction provided data on the cell categories involved in the processing of these contrast agents, the importance and modalities of each category relative to this processing, and the modalities of agent elimination. AMI-HS was rapidly removed from the bloodstream by Kupffer's cells and hepatocytes and then eliminated through bile ducts. AMI-227 remained much longer in the blood compartment since it was processed very slowly by endothelial and Kuppfer's cells in the near absence of hepatocytic participation and thus of elimination by the bile ducts. These results allowed us to base our interpretation of MRI sequences on cytologic observations.     

Ciliate Protozoa in the Rumen of Brazilian Water Buffalo,Bubalus bubalis Linnaeus*

SYNOPSIS Total numbers and distribution of genera, subgenera and species were determined for the ciliate protozoa in rumen contents of 4 Brazilian water buffalo Bubalus bubalis Linnaeus. The fauna of one animal, housed in close proximity to European and zebu-type cattle, differed considerably from that of the remaining animals, which were somewhat isolated on a large ranch. Several of the protozoan species observed in the semi-isolated animals were first described in rumen contents from humped Indian cattle, and their subsequent occurrence in other hosts and geographic locations has been limited or absent. In all, 49 different species of protozoa were found, 8 of which have not been previously described. Three of the new species belong to the genus Entodinium: E. ciculum sp. n., E. spinonucleatum sp. n. and E. triangulum sp. n.: 4 to Diplodinium (Ostracodinium): D. (O.) brazili sp. n., D. (O.) esalqum sp. n., D. (O.) nucleolobum sp. n., and D. (O.) tiete sp. n.; and one to Diplodinium (Eudiplodinium): D. (E.) bubalus sp. n.     

锌对盐藻生长与物质积累的调控作用

实验研究了不同浓度的锌对盐藻细胞生长与物质积累的调控作用。结果表明,培养液中供给锌过多或过少都不利于盐藻细胞的生长与物质积累。以培养基中6 mg/L的锌浓度对盐藻细胞生长、蛋白质合成与β-胡萝卜素积累的促进作用最大。这一锌浓度可用于盐藻的生产性培养。当培养液中锌浓度较高(8 mg/L)或较低(2 mg/L)时,单个盐藻细胞中的蛋白质与β-胡萝卜素含量较高。但此时,因培养液中细胞密度较低,盐藻细胞积累的物质总量仍然较少。在锌浓度较高或较低的逆境条件下,盐藻可能通过适应性反应形成了逆境蛋白质与胡萝卜素等。     

锌金属蛋白酶家族介绍及结构机理研究进展

锌金属蛋白酶是一类分布广泛,种类繁多的水解酶家族,是近年来人们研究的热点．一般来说,HEXXH保守序列一直都作为锌金属蛋白酶家族分类的依据．除此之外,不同种类的酶还有一些其它的序列特征．谷氨酸锌蛋白（gluzincin）在锌离子配体处共有2组保守区域;甲硫氨酸锌蛋白（metzincin）则拥有1个延长的保守序列HEBXHXBGBXH,里面包含了第3个His配体．大量的金属蛋白酶晶体结构被解析出来,从中可以发现该类酶的活性中心都包含有1个锌离子．锌金属蛋白酶的催化机理通常认为是锌离子与1个水分子结合活化而成．但是,最近的发现证实了并不是所有的锌金属蛋白酶催化都需要水分子参与活化．在本文中,综述了这些已发表的锌金属蛋白酶家族的分类、结构特征、底物特异性识别和催化机理．     

锌胁迫对铜锈环棱螺致死率、富集性、CAT和SOD活性的影响

以浙江省温州市瓯海茶山某河内的铜锈环棱螺Bellamya aeruginosa为受试生物,研究了不同浓度Zn2+胁迫下,铜锈环棱螺的致死效应、铜锈环棱螺对锌的富集性以及锌对铜锈环棱螺体内的过氧化氢酶(CAT)和超氧化物歧化酶(SOD)活性的影响.结果表明Zn2+对铜锈环棱螺具有较大的毒害影响.当Zn2+浓度达到3.0 mg/L时,就会引起个别铜锈环棱螺死亡,死亡率为2.66％.随着Zn2+浓度的增大(0.3 mg/L到2.4 mg/L)和处理时间的增加(2d到8d),除个别组外,铜锈环棱螺对锌的累积量均随之上升,具有一定的时间效应和剂量效应.铜锈环棱螺在锌胁迫下,肝胰腺的CAT和SOD活性受到较大的影响.当Zn2+浓度较低时,CAT和SOD活性均有较大的升高,但当Zn2+浓度较高时,CAT和SOD活性则受到一定的抑制.     

Electron tomography of biological samples

Electron tomography allows computing three-dimensional (3D) reconstructions of objects from their projections recorded at several angles. Combined with transmission electron microscopy, electron tomography has contributed greatly to the understanding of subcellular structures and organelles. Performed on frozen-hydrated samples, electron tomography has yielded useful information about complex biological structures. Combined with energy filtered transmission electron microscopy (EFTEM) it can be used to analyze the spatial distribution of chemical elements in biological or material sciences samples. In the present review, we present an overview of the requirements, applications, and perspectives of electron tomography in structural biology.Translated from Biokhimiya, Vol. 69, No. 11, 2004, pp. 1497–1505.Original Russian Text Copyright © 2004 by Marco, Boudier, Messaoudi, Rigaud.     

Electron microscopy,electron diffraction,and element analysis of wet biological specimens

Temperature controlled differentially pumped environmental chambers now allow more routine examination of wet specimens in the electron microscope. A sensitive test of their efficiency is the ability to provide high resolution electron diffraction patterns from wet, unfixed protein microcrystals. Fortunately, wet specimens can be prepared with only a few tens of nanometers thickness of remaining water, so extraneous electron scattering by liquid water can be kept to a minimum. It still remains to be determined whether microprobe analysis (X-ray or electron energy-loss spectroscopy) using wet specimens gives better element localization in cells than the current freezing methods. More extensive comparisons are also required of the ultrastructural preservation and visibility of macromolecules immersed in a thin layer of water vs immersion in a thin layer of amorphous ice. However, the recent introduction of commercial forms of the necessary equipment now make these comparisons more feasible.     

Zinc tolerance and accumulation in metallicolous and nonmetallicolous populations of Arabidopsis halleri (Brassicaceae)

Zinc tolerance was investigated in five populations of Arabidopsis halleri (syn.: Cardaminopsis halleri ) raised from seeds collected from contaminated and uncontaminated sites. Tolerance was measured by determining the concentration which inhibited root growth (EC₁₀₀). A. halleri populations from contaminated and uncontaminated sites were found to be Zn-tolerant compared with the Zn-nontolerant species Arabidopsis thaliana and A. lyrata subsp. petraea . At very high Zn concentrations, populations of A. halleri from uncontaminated sites were slightly less Zn-tolerant than those from contaminated sites. These observations support the hypothesis that in A. halleri , Zn tolerance is largely a constitutive property. One population from an uncontaminated site and one population from a contaminated site were studied for Zn uptake. Zinc content was measured in shoots and roots using a colorimetric test under laboratory conditions. The results showed that whatever their origin, individuals from both populations are Zn accumulators compared with the nonaccumulator species A. thaliana . Moreover, the population from the uncontaminated area accumulated Zn in its shoots and roots more quickly than the population from the contaminated site. These results suggest that, in A. halleri , Zn accumulation to very high concentration is a constitutive property.     

Crystallographic and Biochemical Analysis of the Ran-binding Zinc Finger Domain

The nuclear pore complex (NPC) resides in circular openings within the nuclear envelope and serves as the sole conduit to facilitate nucleocytoplasmic transport in eukaryotes. The asymmetric distribution of the small G protein Ran across the nuclear envelope regulates directionality of protein transport. Ran interacts with the NPC of metazoa via two asymmetrically localized components, Nup153 at the nuclear face and Nup358 at the cytoplasmic face. Both nucleoporins contain a stretch of distinct, Ran-binding zinc finger domains. Here, we present six crystal structures of Nup153-zinc fingers in complex with Ran and a 1.48 Å crystal structure of RanGDP. Crystal engineering allowed us to obtain well diffracting crystals so that all ZnF-Ran complex structures are refined to high resolution. Each of the four zinc finger modules of Nup153 binds one Ran molecule in apparently non-allosteric fashion. The affinity is measurably higher for RanGDP than for RanGTP and varies modestly between the individual zinc fingers. By microcalorimetric and mutational analysis, we determined that one specific hydrogen bond accounts for most of the differences in the binding affinity of individual zinc fingers. Genomic analysis reveals that only in animals do NPCs contain Ran-binding zinc fingers. We speculate that these organisms evolved a mechanism to maintain a high local concentration of Ran at the vicinity of the NPC, using this zinc finger domain as a sink.     

Foliar Spraying and Seed Soaking of Zinc Fertilizers Decreased Cadmium Accumulation in Cucumbers Grown in Cd-Contaminated Soils

It is important to use proper agronomic management to reduce cadmium (Cd) accumulation in plants, ensuring food safety. To find the most effective agronomic approach, the effect of foliar spraying and seed soaking of zinc (Zn) fertilizers on Cd accumulation in cucumbers (Cucumis sativus L.) grown in two soil Cd levels (2 and 5 mg kg^?1 Cd) with and without an immobilizing amendment (red mud, RM) was investigated in the present study. The results showed that the treatment of foliar Zn or seed Zn significantly decreased the Cd concentration in cucumber shoots by about 12–36% in Cd-contaminated soils without amendment. Combined with RM treatment, the foliar Zn treatment further decreased the Cd concentration in cucumber shoots by up to 48–66% in Cd-contaminated soils. There were significant negative correlations between Cd and Zn concentrations in shoots of cucumbers grown in soils treated with RM and foliar Zn. The results revealed that the cucumber seedlings treated with RM and foliar Zn had a higher capacity for limiting the transfer of Cd to aboveground tissues. The results also suggested that increasing seed Zn concentrations sufficiently might act as an efficient, economic, and practical method for decreasing Cd uptake in crops grown in mildly Cd-contaminated and Zn-deficient soils.     

小麦锌指蛋白基因的克隆、序列与表达分析

根据R基因及其调控基因保守序列设计简并性引物,对白粉菌接种和未接种处理的一对抗病和感病的小麦—黑麦等位突变易位系TAM104R和TAM104S总RNA进行RT-PCR扩增,得到一个诱导表达的cDNA片段。序列分析表明,该片段全长2474bp,其中含有一个822bp的完整开放阅读框,推测其编码一个有273个氨基酸残基、分子量约31kD的蛋白质分子。蛋白质的氨基酸序列比对显示,该蛋白质分子具有C2HC锌结合motif CX2CX4HX4C结构和锌指domain,可见克隆的cDNA是一个锌指蛋白基因,命名为TaZF。Southern杂交表明,TaZF在抗病易位系TAM104R的基因组中是多拷贝的。半定量RT-PCR分析显示,TaZF基因属组成型表达、但受白粉菌诱导表达上调的基因,推测其与白粉病菌的侵染过程相关。基因组DNA专化扩增、克隆和测序揭示TaZF基因无内含子。