冰核活性基因及其作报告基因的应用研究进展

国内外已经克隆了8个冰核活性基因,可用作报告基因,主要应用在植物－微生物互作研究、营养元素利用研究、病原微生物高敏检测和各种用途的细菌细胞表面展示等领域。     

Large-scale Production and Purification of an Erwinia ananas Ice Nucleation Protein and Evaluation of Its Ice Nucleation Activity

The ice nucleation-active protein of Erwinia ananas IN-10 (inaA protein) was over-expressed as inclusion bodies in Escherichia coli in a yield of 15.3 mg of inaA protein from 60 mg of bacterial cells on a dry-matter basis. The inaA protein was purified from the inclusion bodies by solubilization with detergents to obtain a protein preparation free from sugar and lipid. This preparation had a distinct ice nucleation activity, indicating that the inaA protein per se is able to act as a nucleus.     

Detection of Variation of the R-Domain Structure of Ice Nucleation Genes in Erwinia herbicola-Group Bacteria by PCR-RFLP Analysis

The structure of ice nucleation (IN) genes was compared among 20 strains of Erwinia herbicola-group bacterium of plant- and insect-origin including E. herbicola M1 (IceE) and E. ananas IN10 (inaA) that had been previously reported. When the DNAs of N-domain or C-domain were amplified, PCR products with similar size were obtained in all strains, while the size of the PCR products from the whole genes containing the R domain varied remarkably within a range of 3.8 kb to 4.4 kb. RFLP analysis of the IN genes revealed that the size of the R-domains were varied within the region from the PvuII site to DraI site, and 20 IN genes were classified into 12 groups. Furthermore, all the strains identified as E. ananas based on six bacteriological properties were different from those of E. herbicola. These results suggest that the IN genes may be distributed only in E. ananas strains among “herbicola group bacteria.” Received: 18 March 1998 / Accepted: 28 April 1998     

Ice Nucleation Activity in Fusarium acuminatum and Fusarium avenaceum

Twenty fungal genera, including 14 Fusarium species, were examined for ice nucleation activity at −5.0°C, and this activity was found only in Fusarium acuminatum and Fusarium avenaceum. This characteristic is unique to these two species. Ice nucleation activity of F. avenaceum was compared with ice nucleation activity of a Pseudomonas sp. strain. Cumulative nucleus spectra are similar for both microorganisms, while the maximum temperatures of ice nucleation were −2.5°C for F. avenaceum and −1.0°C for the bacteria. Ice nucleation activity of F. avenaceum was stable at pH levels from 1 to 13 and tolerated temperature treatments up to 60°C, suggesting that these ice nuclei are more similar to lichen ice nuclei than to bacterial ones. Ice nuclei of F. avenaceum, unlike bacterial ice nuclei, pass through a 0.22-μm-pore-size filter. Fusarial nuclei share some characteristics with the so-called leaf-derived nuclei with which they might be identified: they are cell free and stable up to 60°C, and they are found in the same kinds of environment. Highly stable ice nuclei produced by fast-growing microorganisms have potential applications in biotechnology. This is the first report of ice nucleation activity in free-living fungi.     

肠道正常菌群与人体的关系

人体肠道正常菌群与人体有着非常密切的利害关系。正常情况下,人体内、外环境与肠道正常菌群保持着相对平衡状态,它确保着人体的健康,但一旦在某些因素的作用下,平衡被打破,将导致某些疾病的发生。主要介绍肠道正常菌群的概况、与人体的利害关系、以及菌群失调的调整等。     

Plants as Sources of Airborne Bacteria, Including Ice Nucleation-Active Bacteria

Vertical wind shear and concentration gradients of viable, airborne bacteria were used to calculate the upward flux of viable cells above bare soil and canopies of several crops. Concentrations at soil or canopy height varied from 46 colony-forming units per m³ over young corn and wet soil to 663 colony-forming units per m³ over dry soil and 6,500 colony-forming units per m³ over a closed wheat canopy. In simultaneous samples, concentrations of viable bacteria in the air 10 m inside an alfalfa field were fourfold higher than those over a field with dry, bare soil immediately upwind. The upward flux of viable bacteria over alfalfa was three- to fourfold greater than over dry soil. Concentrations of ice nucleation-active bacteria were higher over plants than over soil. Thus, plant canopies may constitute a major source of bacteria, including ice nucleation-active bacteria, in the air.     

冰晶核蛋白及其在细菌表面展示技术中的应用

冰晶核蛋白(ice nucleation protein,INP)是一种分泌型外膜蛋白,广泛分布于丁香假单胞菌,荧光假单胞菌和其他革兰氏阴性菌中。由于其在相对高温下(-2~-4℃)形成冰核的特性,INP最早应用于生物制冷领域。在细菌表面展示技术中,冰晶核蛋白作为运载蛋白得到广泛的应用。与其他的表面技术载体蛋白相比较,冰晶核蛋白具有稳定表达外源蛋白及展示分子量较大的外源蛋白的优点。INP细胞表面展示技术已被应用于全细胞生物催化剂、全细胞吸附剂和环境污染物降解剂等的开发,本文将简述INP表面展示技术的研究进展。     

Substrate Concentration and Plasmid Transfer Frequency between Bacteria in a Model Rhizosphere

Abstract The influence of substrate concentration on plasmid transfer frequency in the rhizosphere was investigated using a physical model employing a hollow fiber membrane instead of a real root. The absolute number of transconjugants produced increased with increasing substrate (glucose) concentration, but the plasmid transfer frequency decreased exponentially with increasing substrate concentration from 4.4 × 10^-3 at 90 μg ml^-1 glucose to 1.35 × 10^-5 at 3600 μg ml^-1 glucose. These results were found to be heavily dependant on donor to recipient ratio and distribution of strains, but independent of initial donor and recipient inoculum density and regime. These observations also show that plasmid transfer frequency is reduced at high substrate concentrations, which suggests that plasmid transfer is either stimulated when growth limiting nutrient is scarce or inhibited when it is abundant. Received: 30 November 1999; Accepted: 4 April 2000; Online Publication: 18 July 2000     

Injury of Bacteria by Sanitizers

Injury of test cultures was quantitated by differences in colony counts obtained with a complete medium and those obtained on conventional selective media. Staphylococcus aureus, Streptococcus faecalis, and several strains of Escherichia coli were injured when exposed to the quaternary ammonium compound methylalkyltrimethyl ammonium chloride. Representative hypochlorite sanitizers also caused injury of E. coli ML30. Sanitizer concentration appeared to be the main factor in the cause of death and injury, a higher concentration being needed to cause death. Increases in temperature did not result in substantial increases in injury; however, the lethal effect was greater at higher temperatures. Varying the cell concentration from 10(7) to 10(9) cells per ml did not change the fraction of cell population killed or injured. The inability or failure of common selective media to detect injured bacteria in food could have serious public health consequences.     

蛇场病死蛇与感染腐败谢瓦纳拉菌的探讨

目的探讨蛇场蛇死亡与细菌感染的关系。方法从８５条死蛇中随机抽样２０条,取其心脏积血进行微生物检验。结果从其中的１４条死蛇中分离出腐败谢瓦纳拉菌,分离阳性率为７０％。结论认为该菌可能是蛇病死的原因之一。     

Relationship between Cell Surface Properties and Transport of Bacteria through Soil

A study was conducted to relate the properties of Enterobacter, Pseudomonas, Bacillus, Achromobacter, Flavobacterium, and Arthrobacter strains to their transport with water moving through soil. The bacteria differed markedly in their extent of transport; their hydrophobicity, as measured by adherence to n-octane and by hydrophobic-interaction chromatography; and their net surface electrostatic charge, as determined by electrostatic interaction chromatography and by measurements of the zeta potential. Transport of the 19 strains through Kendaia loam or their retention by this soil was not correlated with hydrophobicities or net surface charges of the cells or the presence of capsules. Among 10 strains tested, the presence of flagella was also not correlated with transport. Retention was statistically related to cell size, with bacteria shorter than 1.0 μm usually showing higher percentages of cells being transported through the soil. We suggest that more than one characteristic of bacterial cells determines whether the organisms are transported through soil with moving water.     

A Comparative Analysis of the Ice Nucleation Activity of Pseudomonad Cells and Lipopolysaccharides

The paper deals with the study of the ice nucleation activity of the cells, extracellular lipopolysaccharides (ELPSs), lipopolysaccharides (LPSs), and LPS structural components (lipid A, core oligosaccharide, and O-specific polysaccharide) of Pseudomonas fluorescens, P. syringae, P. fragi, and P. pseudoalcaligenes. Aqueous suspensions of intact cells of P. syringae IMV 1951 and IMV 185 began to freeze at –1 and –4°C, respectively. This suggests that these cells possess ice nucleation activity. Aqueous cell suspensions of two other strains, P. fluorescens IMV 1433 and IMV 2125, began to freeze at lower temperatures than did distilled water (–9°C), which suggests that the cells of these strains possess antifreeze activity. The ice nucleation activity of the bacterial strains studied did not show any correlation with their taxonomic status. The ice nucleation activity of ELPSs depended little on their concentration (within a concentration range of 0.2–0.4%). In most cases, the ice nucleation activity of ELPSs, LPSs, and LPS structural components differed from that of the intact cells from which these biopolymers were obtained. This may indicate that the biopolymers under study play a role in ice nucleation but this role is not crucial. The relationship between the structure of LPSs and their effect on ice nucleation is discussed.     

Frost Damage Resulting from Ice Crystal Formation in Seedlings of Spruce and Pine

The influence of ice crystal formation in tissues of unhardened and hardened seedlings of Pinus silvestris L. and Picea abies (L.) Karst. was determined. An apparatus for this purpose was constructed. Unhardened seedlings of spruce collapsed completely as a result of ice crystal formation in their shoots, while unhardened seedlings of pine survived ice crystal formation with or without injury to the needles. After a hardening period of 4–5 weeks, seedlings of spruce and pine survived ice formation but injuries occurred in spruce. For pine, on the contrary, the temperature after ice formation had a decisive effect on the extent of the injuries. Observed injuries are discussed in terms of inter- and intracellular ice crystal formation.     

Toxicity of Smoke to Epiphytic Ice Nucleation-Active Bacteria

Wheat straw smoke aerosols and liquid smoke condensates reduced significantly both the viability and the ice-nucleating activity of Pseudomonas syringae pv. syringae and Erwinia herbicola in vitro and on leaf surfaces in vivo. Highly significant reductions in numbers of bacterial ice nuclei on the surface of both corn and almond were observed after exposure to smoke aerosols. At −5°C, frost injury to corn seedlings colonized by ice nucleation-active bacteria was reduced after exposure to smoke aerosols. Effects on −9°C ice nuclei, although significant, were less than on ice nuclei active at −5°C. These results suggest that smoke from wildfires or smudge pots may reduce plant frost susceptibility and sources of ice nuclei important in other natural processes under some conditions.     

草莓开花期发生霜害的温度

用人工霜箱对草莓(Fragaria ananassa)叶片和花托进行模拟春霜实验.结果表明草莓叶片有忍耐胞间结冰的能力,最低叶温-6.4℃以上,结冰持续90分钟以内的叶片解冻后还能存活.花托不能忍耐胞间结冰,是通过保持过冷却状态以回避结冰伤害.花托温度越低,发生霜害的累积百分率越大,半开花的花托温度降到-5.4℃时累计有50%发生结冰而造成霜害.盛开的花及鲜重大的花发生霜害的温度较高.     

草莓开花期发生霜害的温度

用人工霜箱对草莓(Fragaria ananassa)叶片和花托进行模拟春霜实验。结果表明草莓叶片有忍耐胞间结冰的能力, 最低叶温-6.4 ℃以上, 结冰持续90分钟以内的叶片解冻后还能存活。花托不能忍耐胞间结冰, 是通过保持过冷却状态以回避结冰伤害。花托温度越低, 发生霜害的累积百分率越大, 半开花的花托温度降到-5.4 ℃时累计有50%发生结冰而造成霜害。盛开的花及鲜重大的花发生霜害的温度较高。     

砂岩型铀矿床硫酸盐还原菌与铀地球化学分布关系研究

利用生物学方法对十红滩砂岩型铀矿床各亚带硫酸盐还原菌分布特征和各亚带铀含量与硫酸盐还原菌之间关系进行了分析探讨.结果表明:硫酸盐还原菌在矿石带种类、数量较多,硫酸盐还原菌这种分布特征与铀的地球化学分布呈相关性,这为砂岩型铀矿中铀的地球化学分布研究提供生物学检测手段.