芽胞杆菌B13解养分效果研究

目的评价芽胞杆菌B13的功能。方法通过培养基及土壤培养分析其解钾、解磷的效果。结果芽胞杆菌B13培养7d后液体培养基中的有效磷含量（0．94μg／mL）比对照组（0．75μg/mL）增加25．33％,有效钾含量（0．54μg/mL）比对照组（0．31μg／mL）增加74．19％,解磷和解钾差异均有统计学意义（P〈0．01）。芽胞杆菌B13添加在灭菌土壤和不灭菌土壤中都具有明显的解磷、解钾功效,说明芽胞杆菌B13有较好的土壤定植能力。平板解磷试验证明芽胞杆菌B13同时具有解无机磷和有机磷的功效。结论芽胞杆菌B13具有很大的研究与开发价值。     

Survival of gfp-tagged antagonistic bacteria in the rhizosphere of tomato plants and their effects on the indigenous bacterial community

The survival and colonization patterns of Pseudomonas putida PRD16 and Enterobacter cowanii PRF116 in the rhizosphere of greenhouse-grown tomato plants and the effects of their inoculation on the indigenous bacterial community were followed by selective plating, molecular fingerprinting, and confocal laser scanning microscopy (CLSM) over 3 weeks. Both strains, which showed in vitro antagonistic activity against Ralstonia solanacearum, were previously tagged with gfp. Seed and root inoculation were compared. Although plate counts decreased for both gfp-tagged antagonists, PRD16 showed a better survival in the rhizosphere of tomato roots independent of the inoculation method. Analysis of 16S rRNA gene fragments amplified from total community DNA by denaturing gradient gel electrophoresis and CLSM confirmed the decrease in the relative abundance of the inoculant strains. Pronounced differences in the Pseudomonas community patterns for plants inoculated with PRD16 compared to the control were detected 3 weeks after root inoculation, indicating a longer-lasting effect. Analysis by CLSM showed rather heterogeneous colonization patterns for both inoculant strains. In comparison with seed inoculation, root inoculation led to a much better colonization as evidenced by all three methods. The colonization patterns observed by CLSM provide important information on the sampling strategy required for monitoring inoculant strains in the rhizosphere.     

荧光假单胞菌CLW17菌株pqqE和GDH基因的功能

[背景]磷是植物生长所必需的大量元素,但绝大多数不能被植物吸收利用。然而溶磷微生物能够分泌有机酸来溶解土壤中难溶性磷,提高土壤中磷的利用率,促进植物生长,提高作物的产量和品质。[目的]探究高效解磷荧光假单胞菌CLW17菌株的pqqE和GDH基因的生理学功能。[方法]利用生物在线软件对2个基因编码蛋白进行生物信息学分析。利用同源重组技术分别获得pqqE和GDH基因缺失突变株(CLW17ΔpqqE,CLW17ΔGDH),并使用接合转移的方式获得回补菌株(ΔpqqE/pqqE,ΔGDH/GDH)。分别采用NBRIP培养基、钼锑抗比色法及高压液相色谱法(HPLC)对野生型、突变株及互补株的溶磷及产有机酸能力进行检测。[结果]pqqE和GDH基因编码氨基酸数目分别为390和803,均无信号肽。pqqE无跨膜结构域,而GDH预测有5个跨膜结构域。pqqE和GDH基因是CLW17菌株的溶磷相关基因,2个基因的缺失均使该菌株的溶磷能力显著下降,而回补株可以恢复溶磷能力。CLW17野生株能分泌多种有机酸,其中葡萄糖酸(gluconic acid,GA)含量最多,其次是乙酸;但敲除株产有机酸的能力明显降低...     

Effect of inoculation method and plant growth medium on endophytic colonization of sorghum by the entomopathogenic fungus <Emphasis Type="Italic">Beauveria bassiana</Emphasis>

A study was conducted to determine the effect of inoculation method and plant growth medium on colonization of sorghum by an endophytic Beauveria bassiana. Colonization of leaves, stems, and roots by B. bassiana was assessed 20-days after application of the fungus. Although B. bassiana established as an endophyte in sorghum leaves, stems, and roots regardless of inoculation method (leaf, seed, or soil inoculation), plant growth medium (sterile soil, non-sterile soil, or vermiculite) apparently influenced colonization rates. Seed inoculation with conidia caused no stem or leaf colonization by the fungus in non-sterile soil but did result in substantial endophytic colonization in vermiculite and sterile soil. Leaf inoculation did not result in root colonization, regardless of plant growth medium. Endophytic colonization was greater in leaves and stems than roots. Endophytic colonization by B. bassiana had no adverse effects on the growth of sorghum plants. Leaf inoculation with a conidial suspension proved to be the best method to introduce B. bassiana into sorghum leaves for plants growing in either sterile or non-sterile soil. Further research should focus on the virulence of endophytic B. bassiana against sorghum stem borers.     

Importance of pfkA for rapid growth of Enterobacter cloacae during colonization of crop seeds

Enterobacter cloacae A-11 is a prototrophic, glycolytic mutant of strain 501R3 with a single transposon insertion in pfkA. The populations of strain A-11 on cucumber and radish seeds were smaller than the populations of strain 501R3 in natural soil, but the populations of these two strains on pea, soybean, sunflower, and sweet corn seeds were similar (D. P. Roberts, P. D. Dery, I. Yucel, J. Buyer, M. A. Holtman, and D. Y. Kobayashi, Appl. Environ. Microbiol. 65:2513-2519, 1999). The net effect of the mutation in pfkA in vitro was a shift from rapid growth on certain carbohydrates detected in seed exudates to much slower growth on other carbohydrates, amino acids, and organic acids. The impact of the mutation in pfkA was greatest on the growth rate of E. cloacae on the seeds that released the smallest quantities of fructose, other carbohydrates, and amino acids. Corn, pea, soybean, and sunflower seeds released total amounts of carbohydrates and amino acids at rates that were approximately 10- to 100-fold greater than the rates observed with cucumber and radish seeds for the first 24 h after inhibition began. The growth rate of strain A-11 was significantly less (50% less) than the growth rate of strain 501R3 on radish seeds, and the growth rate of strain A-11 was too low to estimate on cucumber seeds in sterile sand for the first 24 h after inhibition began. The growth rate of strain A-11 was also significantly lower on soybean seeds, but it was only 17% lower than the growth rate of strain 501R3. The growth rates of strains 501R3 and A-11 were similar on pea, sunflower, and corn seeds in sterile sand for the first 30 h after imbibition began. Large reductions in the growth rates of strain A-11 on seeds were correlated with subsequent decreased levels of colonization of seeds compared to the levels of colonization of strain 501R3. The strain A-11 populations were significantly smaller than the strain 501R3 populations only on radish and cucumber seeds. The mutation in pfkA appears to decrease the level of colonization by E. cloacae for seeds that release small quantities of reduced carbon compounds by decreasing the size of the pool of compounds that support rapid growth by this bacterium.     

Importance of pfkA for Rapid Growth of Enterobacter cloacae during Colonization of Crop Seeds

Enterobacter cloacae A-11 is a prototrophic, glycolytic mutant of strain 501R3 with a single transposon insertion in pfkA. The populations of strain A-11 on cucumber and radish seeds were smaller than the populations of strain 501R3 in natural soil, but the populations of these two strains on pea, soybean, sunflower, and sweet corn seeds were similar (D. P. Roberts, P. D. Dery, I. Yucel, J. Buyer, M. A. Holtman, and D. Y. Kobayashi, Appl. Environ. Microbiol. 65:2513–2519, 1999). The net effect of the mutation in pfkA in vitro was a shift from rapid growth on certain carbohydrates detected in seed exudates to much slower growth on other carbohydrates, amino acids, and organic acids. The impact of the mutation in pfkA was greatest on the growth rate of E. cloacae on the seeds that released the smallest quantities of fructose, other carbohydrates, and amino acids. Corn, pea, soybean, and sunflower seeds released total amounts of carbohydrates and amino acids at rates that were approximately 10- to 100-fold greater than the rates observed with cucumber and radish seeds for the first 24 h after inhibition began. The growth rate of strain A-11 was significantly less (50% less) than the growth rate of strain 501R3 on radish seeds, and the growth rate of strain A-11 was too low to estimate on cucumber seeds in sterile sand for the first 24 h after inhibition began. The growth rate of strain A-11 was also significantly lower on soybean seeds, but it was only 17% lower than the growth rate of strain 501R3. The growth rates of strains 501R3 and A-11 were similar on pea, sunflower, and corn seeds in sterile sand for the first 30 h after imbibition began. Large reductions in the growth rates of strain A-11 on seeds were correlated with subsequent decreased levels of colonization of seeds compared to the levels of colonization of strain 501R3. The strain A-11 populations were significantly smaller than the strain 501R3 populations only on radish and cucumber seeds. The mutation in pfkA appears to decrease the level of colonization by E. cloacae for seeds that release small quantities of reduced carbon compounds by decreasing the size of the pool of compounds that support rapid growth by this bacterium.     

丛枝菌根真菌对枳根净离子流及锌污染下枳苗矿质营养的影响

盆栽实验研究了不同施Zn水平(0、300 mg/kg和600 mg/kg)下,接种丛枝菌根真菌Glomus intraradices对枳苗生长、Zn、Cu、P、K、Ca、Mg分布的影响,并采用非损伤微测技术测定分析了菌根化与非菌根化枳根净Ca²⁺、H⁺、NO₃^-离子流动态。结果表明:(1)在不同施Zn水平下,接种菌根真菌显著提高了枳苗地上部及根部鲜重;随着施Zn水平的提高,菌根侵染率呈降低趋势,枳苗地上部与根部Zn含量呈增加趋势,且接种株根部Zn含量显著高于未接种株。(2)接种株未施Zn处理的地上部Cu、P、K、Mg和根部Cu含量、施600 mg/kg Zn处理的根部Cu及施300 mg/kg Zn处理的根部P含量均显著高于对照,而菌根真菌侵染对枳苗Ca含量并无显著性影响。(3)接种株未施Zn处理的根部距根尖端0 μm和600 μm处净Ca²⁺流出速率、600 μm处净H⁺流入速率、2400 μm处净NO₃^-流入速率均显著高于未接种株。     

消除共生质粒的费氏中华根瘤菌HND29SR在大豆根圈的定殖动态

比较研究了费氏中华根瘤菌（Sinorhizobium fredii)HN01(出发菌）、发光酶基因标记菌HNO1L（参照菌）、消除HN01共生质粒的菌株HND29SR在无菌砂培条件下的大豆根圈定殖动态。供试菌单独接种时：HN01、HN01L和HND29SR的定殖动态基本一致,其早期定殖密度下降较快,播种后第16天时HN01和HN01L分别达到较高的定殖水平6.49logcfu/g鲜根和6.78logcfu/g鲜根,然后维持相对稳定的定殖水平。但HND29SR的定殖密度持续下降到播种后第16天时才开始上升,至第35天时仍维持相对稳定的定殖密度6.94logcfu/g鲜根。等量混合接种时供试菌在根圈定殖群体中各自定殖密度在测定过程中基本相等。结果表明消除HN01的共生质粒对其在大豆根圈中定殖能力无显著影响。     

Role of pfkA and general carbohydrate catabolism in seed colonization by Enterobacter cloacae.

Enterobacter cloacae A-11 is a transposon mutant of strain 501R3 that was deficient in cucumber spermosphere colonization and in the utilization of certain carbohydrates (D. P. Roberts, C. J. Sheets, and J. S. Hartung, Can. J. Microbiol. 38:1128-1134, 1992). In vitro growth of strain A-11 was reduced or deficient on most carbohydrates that supported growth of strain 501R3 but was unaffected on fructose, glycerol, and all amino acids and organic acids tested. Colonization by strain A-11 was significantly reduced (P 相似文献   

Detect of Escherichia coli O157:H7 in ground beef samples using piezoelectric excited millimeter-sized cantilever (PEMC) sensors

Piezoelectric-excited millimeter-sized cantilever (PEMC) sensors consisting of a piezoelectric and a borosilicate glass layer with a sensing area of 4 mm2 were fabricated. An antibody specific to Escherichia coli (anti-E. coli) O157:H7 was immobilized on PEMC sensors, and exposed to samples containing E. coli O157:H7 (EC) prepared in various matrices: (1) broth, broth plus raw ground beef, and broth plus sterile ground beef without inoculation of E. coli O157:H7 served as controls, (2) 100 mL of broth inoculated with 25 EC cells, (3) 100 mL of broth containing 25 g of raw ground beef and (4) 100 mL of broth with 25 g of sterile ground beef inoculated with 25 EC cells. The total resonant frequency change obtained for the broth plus EC samples were 16+/-2 Hz (n=2), 30 Hz (n=1), and 54+/-2 Hz (n=2) corresponding to 2, 4, and 6h growth at 37 degrees C, respectively. The response to the broth plus 25 g of sterile ground beef plus EC cells were 21+/-2 Hz (n=2), 37 Hz (n=1), and 70+/-2 Hz (n=2) corresponding to 2, 4, and 6 h, respectively. In all cases, the three different control samples yielded a frequency change of 0+/-2 Hz (n=6). The E. coli O157:H7 concentration in each broth and beef samples was determined by both plating and by pathogen modeling program. The results indicate that the PEMC sensor detects E. coli O157:H7 reliably at 50-100 cells/mL with a 3 mL sample.     

Thiols of Cu-treated maize plants inoculated with the arbuscular-mycorrhizal fungus Glomus intraradices

Mycorrhizal colonization of roots, fresh weight, content of cysteine, γ-glutamylcysteine (γEC). glutathione (GSH), thiol groups in Cu-binding peptides (CuBP), and the uptake of Cu were measured in roots and shoots of maize ( Zea mays L., cv. Honeycomb F-1) grown in quartz sand, with Cu at 0, 4.5, 9, 15 and 30 μg g⁻¹ added with or without inoculum of the arbuscular-mycorrhizal fungus (AMF) Glomus intraradices . In control plants (no Cu added) AMF significantly reduced shoot growth, but did not affect root growth. At an external Cu supply of 9 μg (g quartz sand)⁻¹ or higher, both mycorrhizal colonization and growth of roots and shoots of mycorrhizal and non-mycorrhizal plants were significantly reduced.
With up to 9 μg Cu g⁻¹, mycorrhizal colonization increased the content of cysteine, γEC and GSH in the roots. However, the amount of thiols in CuBPs was not increased by mycorrhizal colonization in Cu-treated plants and no differences in Cu uptake were detected between non-mycorrhizal and mycorrhizal plants. A CuBP-complex with a relative molecular mass of 7300 and a SH:Cu ratio of 1.77:1 was separated on a Sephadex G-50 column from both non-inoculated and inoculated roots of Cu-treated plants. HPLC chromatography of the CuBPs of both non-inoculated and inoculated roots resulted in a similar peak pattern, indicating that no additional CuBPs were formed by the fungus. In conclusion, our results do not support the idea that AMF protects maize from Cu-toxicity.     

矿区分离丛枝菌根真菌对万寿菊吸Cd潜力影响

盆栽试验研究土壤不同施Cd水平(0、5、20、50μg/g)下,接种矿区污染土壤中丛枝菌根真菌对万寿菊根系侵染率、植株生物量及Cd吸收与分配的影响。结果表明:接种丛枝菌根真菌显著提高Cd胁迫下万寿菊的根系侵染率和植株生物量;随着施Cd水平提高,各处理植株Cd浓度显著增加。各施Cd水平下万寿菊地上部Cd吸收量远远高于根系Cd吸收量,在土壤施Cd量达到50μg/g时,接种处理地上部Cd吸收量是根系的3.48倍,对照处理地上部Cd吸收量是根系的1.67倍;同一施Cd水平下接种处理植株Cd吸收量要显著高于对照。总体上,试验条件下污染土壤中分离的丛枝菌根真菌促进了万寿菊对土壤中Cd的吸收,并在一定程度上增加Cd向地上部分的运转,表现出植物提取的应用潜力。     

Tricalcium phosphate solubilizing abilities of Trichoderma spp. in relation to P uptake and growth and yield parameters of chickpea (Cicer arietinum L.)

Nine isolates of Trichoderma spp. were investigated for their ability to solubilize insoluble phosphate in Pikovskaya's broth and were compared with an efficient phosphate-solubilizing bacterium Bacillus megaterium subsp. phospaticum PB that was used as the reference strain. All 9 Trichoderma isolates were found to solubilize insoluble tricalcium phosphate to various extents. Trichoderma viride (TV 97) (9.03 microg x mL(-1)), Trichoderma virens (PDBCTVs 12) (9.0 microg x mL(-1)), and Trichoderma virens (PDBCTVs 13) (8.83 microg x mL(-1)) solubilized 70% of that solubilized by the reference strain Bacillus megaterium (12.43 microg x mL(-1)). Pot culture and field evaluations with Trichoderma harzianum (PDBCTH 10), Trichoderma viride (TV 97), and Trichoderma virens (PDBCTVs 12) using chickpea (Cicer arietinum L.) 'Annegeri-1' as the test plant and rock phosphate as the phosphorus source showed significantly increased P uptake in plants treated with Trichoderma harzianum (PDBCTH 10) followed by Trichoderma virens (PDBCTVs 12) and Trichoderma viride (TV 97). Inoculation of Trichoderma spp. also showed increased growth and yield parameters of chickpea compared with the uninoculated controls under both glasshouse and field conditions.     

VA-mycorrhiza mediated P effect on growth and yield of sunflower (Helianthus annuus L.) at different P levels

A field trial was conducted to study the response of sunflower (Helianthus annuus L.) to different phosphorus levels (16, 24 or 32 kg P ha^–1) and inoculation with vesicular-arbuscular mycorrhizal fungus, Glomus fasciculatum on vertisol during summer 1993. At the vegetative stage of sunflower, percent mycorrhizal root colonization, spore count, dry biomass and P uptake did not differ significantly between inoculated and uninoculated control plants. However, at later stages (flowering and maturity) percent root colonization, spore count, total dry biomass and total P uptake were significantly higher in inoculated plants than in uninoculated control plants. The total dry biomass, P content and seed yield increased with increasing P level in uninoculated plants, whereas no significant difference was observed between 16 and 32 kg P ha^–1 in inoculated plants. The positive effect of mycorrhizal inoculation decreased with increasing P level above 16 kg P ha^–1, due to decreased percent root colonization and spore count at higher P levels.     

Role of pfkA and General Carbohydrate Catabolism in Seed Colonization by Enterobacter cloacae

Enterobacter cloacae A-11 is a transposon mutant of strain 501R3 that was deficient in cucumber spermosphere colonization and in the utilization of certain carbohydrates (D. P. Roberts, C. J. Sheets, and J. S. Hartung, Can. J. Microbiol. 38:1128–1134, 1992). In vitro growth of strain A-11 was reduced or deficient on most carbohydrates that supported growth of strain 501R3 but was unaffected on fructose, glycerol, and all amino acids and organic acids tested. Colonization by strain A-11 was significantly reduced (P ≤ 0.05) for cucumber and radish seeds compared to that of strain 501R3, but colonization of pea, soybean, sunflower, and sweet corn seeds was not reduced. Pea seeds released several orders of magnitude more total carbohydrates and amino acids than cucumber and radish seeds and approximately 4,000-fold more fructose. Fructose was the only carbohydrate detected in the seed exudates which supported wild-type levels of in vitro growth of strain A-11. Soybean, sunflower, and sweet corn seeds also released significantly greater amounts of fructose and total carbohydrates and amino acids than cucumber or radish seeds. The exogenous addition of fructose to cucumber and radish seeds at quantities similar to the total quantity of carbohydrates released from pea seeds over 96 h increased the populations of strain A-11 to levels comparable to those of strain 501R3 in sterile sand. Molecular characterization of strain A-11 indicated that the mini-Tn5 kanamycin transposon was inserted in a region of the genome with significant homology to pfkA, which encodes phosphofructo kinase. A comparison of strain A-11 with Escherichia coli DF456, a known pfkA mutant, indicated that the nutritional loss phenotypes were identical. Furthermore, the pfkA homolog cloned from E. cloacae 501R3 complemented the nutritional loss phenotypes of both E. coli DF456 and E. cloacae A-11 and restored colonization by strain A-11 to near wild-type levels. These genetic and biochemical restoration experiments provide strong evidence that the quantities of reduced carbon sources found in seed exudates and the ability of microbes to use these compounds play important roles in the colonization of the spermosphere.     

Effects of arbuscular mycorrhizal fungi on the growth,nutrient uptake and glycyrrhizin production of licorice (<Emphasis Type="Italic">Glycyrrhiza uralensis</Emphasis> Fisch)

The growth of licorice in arid areas faces nutritional and environmental stresses. Arbuscular mycorrhizal (AM) fungi have been shown to increase the abilities of plants to develop. However, little is known regarding the role of AM fungi in licorice (Glycyrrhiza uralensis) growth. In the present study, by inoculation with two AM fungi, Glomus mosseae (Nicolson & Gerdemann) Gerd. & Trappe and Glomus veriforme (P. Karst.), the effects on licorice growth in sand were examined by measuring plant height, number of leaves, shoot and root fresh weight, and by analyzing morphological parameters of the root system in sand. The influence of the two microorganisms on the accumulation of mineral nutritions and bioactive components in licorice were also investigated. The results showed that mycorrhyzae were of the Arum-type and their colonization frequency (F %), colonization intensity (M %) and colonization intensity (m %) of AM fungi inoculation were found to be 80.0–84.6%, 49.4–60.0% and 58.4–71.9%, respectively. The inoculation significantly improved plant growth during early and late growth stages in comparison with the control. Moreover, inoculation of G. mosseae and G. versiforme, alone or in combination, improved plant phosphorus acquisition in the leaf over non-inoculation plants. In addition, mycorrhiza formation enhanced the glycyrrhizin concentration in roots, but resulted in a considerable reduction of the root oxidase activity. The results indicate that the inoculation with AM fungi could be a useful approach to increase the licorice pharmic quality.     

棘白菌素对氟康唑耐药的念珠菌体外药物敏感性的研究

目的评价3种棘白菌素类药物（卡泊芬净、米卡芬净、阿尼多芬净）体外对氟康唑耐药念珠菌的药物敏感性。方法采用微量液体稀释法和琼脂稀释法测定最小抑制浓度（MIC）。结果微量液体稀释法：59株耐药白念珠菌3种药物MIC50均为0.06μg/mL,米卡芬净、阿尼多芬净的MIC范围均为0.015～0.125μg/mL,卡泊芬净为0.015～0.25μg/mL;8株耐药光滑念珠菌MIC值均为0.063μg/mL。琼脂稀释法：59株耐药白念珠菌和8株耐药光滑念珠菌3种药物MIC值均为0.063μg/mL。结论3种棘白菌素类药物可能具有治疗氟康唑耐药的念珠菌感染的临床价值。     

红谷霉素生料发酵工艺的研究

红谷霉素是链霉菌702发酵所产的一种生物活性物质,具有较强抗细菌活性。在摇瓶条件下,对链霉菌702生料发酵生产红谷霉素进行研究。采用单因素实验探索生料发酵工艺,筛选出摇瓶生料发酵条件为7.5%的种子接种量和添加抗菌剂1.5ml。在此条件下进行生料发酵,使红谷霉素摇瓶发酵产量达到1.46g/L,比常规灭菌发酵产量提高25.7%,实验结果表明红谷霉素生料发酵是一种潜在可行的发酵方法,不但可以很好的节约发酵成本,还可以提高发酵产量。     

Overexpression of hns in the plant growth‐promoting bacterium Enterobacter cloacae UW5 increases root colonization

Aims: Plant growth‐promoting rhizobacteria (PGPR) introduced into soil often do not compete effectively with indigenous micro‐organisms for plant colonization. The aim of this study was to identify novel genes that are important for root colonization by the PGPR Enterobacter cloacae UW5. Methods and Results: A library of transposon mutants of Ent. cloacae UW5 was screened for mutants with altered ability to colonize canola roots using a thermal asymmetric interlaced (TAIL)‐PCR‐based approach. A PCR fragment from one mutant was reproducibly amplified at greater levels from genomic DNA extracted from mutant pools recovered from seedling roots 6 days after seed inoculation compared to that from the cognate inoculum cultures. Competition assays confirmed that the purified mutant designated Ent. cloacae J28 outcompetes the wild‐type strain on roots but not in liquid cultures. In Ent. cloacae J28, the transposon is inserted upstream of the hns gene. Quantitative RT‐PCR showed that transposon insertion increased expression of hns on roots. Conclusions: These results indicate that increased expression of hns in Ent. cloacae enhances competitive colonization of roots. Significance and Impact of the Study: A better understanding of the genes involved in plant colonization will contribute to the development of PGPR that can compete more effectively in agricultural soils.