甘蔗内生固氮菌研究

对甘蔗内生固氮的分布特点、生理学特性、生物学特性、遗传学特点及应用前景等方面的研究进展进行了全面的概述.     

接种固氮菌Klebsiella sp.120对甘蔗光合特性和主要矿质营养元素含量的影响

【目的】甘蔗是最重要的糖料和能源作物,生物量大,产量高,但大量氮肥的使用不仅增加了甘蔗的种植成本,而且对农田环境造成了污染。发挥与甘蔗联合固氮的生物固氮菌的作用,是降低甘蔗氮肥施用量的方法之一。Klebsiella sp.120是从广西种植的甘蔗体内分离到的具有固氮能力的菌株。为了探讨该菌对甘蔗的促生效应及其在甘蔗上的应用潜能,本文研究了接种该固氮菌对甘蔗光合特性及矿质元素含量的影响。【方法】采用桶栽方法,在甘蔗的分蘖期用接种菌株淋浇4个不同甘蔗品种的根际,在处理后的30 d和60 d取样分析该菌对甘蔗的光合生理和矿质营养元素含量的影响。【结果】接种菌株对4个甘蔗品种的叶绿素含量和净光合速率有一定的促进作用,但不同品种之间存在差异。处理30 d后,淋浇固氮菌处理的甘蔗品种ROC22的叶绿素含量明显提高,比对照增加23%。另外3个甘蔗品种(B8、GT21和GT28)处理组的叶绿素含量均有一定的提高,但与对照相比差异不显著。处理30 d后,只有ROC22和B8品种的净光合速率高于对照,且ROC22品种处理与对照间的差异达到了显著水平。处理60 d后,4个甘蔗品种的净光合速率均高于处理30 d,而且各品种接种处理均高于对照,其中以对ROC22和GT21的促进效果最为明显,达到了显著水平。接种处理对不同甘蔗品种的营养元素含量影响不同,接种处理对4个甘蔗品种叶片中的氮元素含量有促进作用,其中对ROC22的促进效果最为明显,接种处理比对照增加59.8%。接种固氮菌对甘蔗叶片磷和钾元素含量的影响不明显。接种固氮菌对4个甘蔗品种叶片的铁、锰元素含量的影响不明显;ROC22叶片中的锌含量及ROC22和B8叶片中的铜含量接种处理与对照相比显著提高。【结论】接种Klebsiella sp.120固氮菌对甘蔗光合特性及矿质元素含量的影响与甘蔗品种有关,其中对ROC22品种的促生效果最为明显。研究结果可为固氮菌株Klebsiella sp.120在甘蔗上的进一步应用提供参考依据。     

钼对甘蔗体内固氮菌的固氮酶活性的影响

以巴西固氮甘蔗品种B1和B8为材料,在温室桶栽砂培条件下,对甘蔗施以含不同钼水平的营养液,以了解钼对甘蔗体内固氮菌的固氮酶活性的效应。结果表明,在含氮条件下,较低浓度的钼处理能提高甘蔗根内固氮菌的固氮酶活性,高浓度钼处理则能提高B1茎、叶片和B8茎中固氮菌的固氮酶活性;在无氮条件下,钼处理提高了B1叶片和茎中固氮菌的固氮酶活性,而B1根及B8根、茎、叶中固氮菌的固氮酶活性有所降低。此外,甘蔗根、茎、叶中固氮菌的固氮酶活性之间关系较为密切,根中固氮菌的固氮酶活性与叶片和茎中的都呈负相关。上述结果说明:巴西甘蔗在本地也具有一定的固氮能力;在不缺氮条件下,钼处理较利于调节甘蔗体内固氮菌的固氮酶活性,促进其固氮;而在缺氮条件下,钼处理不利于调节甘蔗体内固氮菌的固氮酶活性,抑制其固氮作用。     

两种不同土壤条件下可培养甘蔗内生固氮菌的多样性

为明确不同土壤条件对甘蔗内生固氮菌多样性的影响,对分别种植在红壤土和水稻土的4个甘蔗品种的内生固氮菌进行分离和鉴定,并利用数学生态学方法分析不同来源内生固氮菌的多样性。结果表明,从红壤土的甘蔗组织中分离筛选获得23个内生固氮菌株,它们在基于16S r DNA序列分析结果上属于9个不同的菌属;而从水稻土的甘蔗组织中获得59个内生固氮菌株,分类上属于19个不同的菌属。群落生态学分析结果显示,来源于水稻土的甘蔗内生固氮菌的群落丰富度指数(dma)、多样性指数(H')以及均匀度指数(Jsw)均明显高于红壤土,表明水稻土条件下的甘蔗内生固氮菌群落结构相对复杂、稳定和均匀;而相同土壤条件下4个甘蔗品种间的内生固氮菌丰富度指数和多样性指数差异并不明显。本研究结果反映了土壤生态条件是影响甘蔗内生固氮菌多样性的重要因子。     

降香黄檀根际固氮菌分离鉴定及菌群活性研究

利用Ashby培养基从不同林龄的降香黄檀根际土壤中分离高效固氮菌,为专用生物复合肥的研制提供理论基础。固氮酶活在75.30~432.76 nmol h-1 ml-1之间的菌株有21株,其中菌株JT-N305的固氮酶活最高。其中15株菌株具有分泌IAA的能力,其分泌量在9.76~41.21μg ml-1。结合形态学、生理生化特性及16S r DNA序列分析,初步鉴定JT-N301属于Burkholderia anthina、JT-N304属于Enterobacter ludwigii、JT-N305属于Burkholderia seminalis。菌群的研究表明:3株菌株混菌培养,其固氮酶活高达498.66 nmol h-1 ml-1;相对于其它氮源而言,菌群更易于吸收铵盐;而菌群的最佳碳源为葡萄糖。     

水稻联合固氮菌质粒的鉴定及固氮(nif)基因定位

应用改进的Casse法提取质粒,鉴定了供试菌株的质粒存在情况:高NH_4~+和N_2培养的Alcaligenes faecalis A_15,Enterobacter cloacae E26,E.cloacaeEnSs和klebsiella planticola DWUL2分别携有1—2个大质粒,分子量在30—200Md之间;K.planficola和Pseudomonas saccharophila含有小质粒;K.oxytocaNG13不携质粒。供试菌株中DNA与nif探针R1 nif DH和Ec nif B-Y均具有同源性。A.faecalis A15,E.cloacae E26和K.oxytoca NG13的nif基因位于染色体上,而E.cloacae EnSs的nif基因则位于一个较大质粒上。     

我国甘蔗亲本遗传多样性的AFLP标记分析

采用15对多态性较好的AFLP引物对我国自育的78个甘蔗亲本材料遗传多样性进行分析,结果表明,每对引物的多态性位点平均为65．67,多态陆位点率为82．55％。78个甘蔗亲本的遗传相似系数在0．4535～0．8927之间,平均为0．6821。相同组合后代的遗传相似系数较高,平均为0．7656。以遗传相似系数阈值约为0．700划分,聚类分析把78个亲本分为7大类,系谱记录中亲缘关系密切的亲本品系,大多数都能归为同一类。遗传多样性指数分析显示,不同年代亲本多样性指数差异明显,20世纪80年代最高,为0-3185,70年代最低,为0．2645;不同省区的亲本遗传多样指数变化更显著,在0．1952～0．2999之间,广东最高,云南最低。     

不同大豆品种根瘤固氮酶活性与固氮量差异研究

大豆根瘤固氮酶活性与固氮量是衡量大豆固氮能力的重要指标,也是合理施用氮肥的依据.试验采用框栽方法,应用15N示踪技术,研究了不同大豆品种根瘤固氮酶活性与固氮量差异.结果表明:4个供试大豆品种根瘤固氮酶活性、根瘤干重及根瘤固氮潜力都呈现先升后降的变化趋势,但品种间有显著差异;生育期短的大豆品种根瘤固氮酶活性高于生育期长的品种,根瘤干重则相反,生育期越长,根瘤干重越大;根瘤固氮潜力随大豆品种生育期的延长而增加.大豆植株氮素构成品种间存在明显的差异性,随着品种生育期的延长,根瘤固氮所占的比例提高;在大豆品种黑河41氮素构成中,土壤氮和根瘤固氮所占比例相同,其他品种均以根瘤固氮为主,其次是土壤氮,肥料氮所占的比例很低.     

Biotoxic Effects of Herbicides on Growth,Nodulation, Nitrogenase Activity,and Seed Production in Chickpeas

Abstract

The effects of pre‐emergent (PRE) application of methabenzthiazuron (MBT), terbutryn, and linuron on plant vigor, number of nodules formed, nitrogenase activity (NA), chlorophyll contents, nitrogen (N) uptake, and seed yield were assessed in inoculated chickpeas, grown in sandy clay loam soil, in a controlled environment. The PRE application of MBT, terbutryn, and linuron at 2.5, 1.25 and 2 g a.i. kg^?1, respectively, had no significant adverse effect on plant vitality, yield, and NA of excised nodules in inoculated chickpea plants, but reduced these parameters at 7.5, 3.75, and 6 g a.i. kg^?1 of soil. Among the herbicides tested, linuron at each dose rates significantly (P≤0.05) decreased the grain yield, whereas terbutryn at 1.25 and 2.5 g a.i. kg^?1 improved the grain yield by 19.4% and 11%, respectively, relative to the nonherbicide control. While comparing the sum of mean values of toxicity of all dose rates of each herbicide on grain yield, the toxicity increased in the following order: control=linuron>methabenzthiazuron>terbutryn. Nodulation (nodule number per plant and their dry mass) and chlorophyll contents per plant decreased consistently with increased herbicide rates, except linuron, which at 2 g a.i. kg^?1 improved nodulation at both 45 and 90 days after seeding and corresponded with Mesorhizobium activity. The N content in grain and straw significantly decreased at the highest tested rates of each herbicide. Linuron at 2 and 4 g a.i. kg^?1 improved N content in straw, whereas at 4 g a.i. kg^?1 it significantly reduced N content in grain.     

甘蔗生产机械化系统的试验和分析

该文对广前公司引进的甘蔗生产机器系统3年来的机械化试验进行了评价及经济分析。机械化是一项复杂的系统工程,机械化系统需要合理的机具配置和高质量的现代管理,各个环节都影响整个系统的工作效率。甘蔗生产机械化系统虽然还有很多问题,但是其潜在的高工效、高经济效益是人工难以比拟的。如果建立和完善管理体系和社会化服务体系,并且能够很好地解决土地清理、机具配套、收获部署及人员培训等问题,我国实现甘蔗生产全面机械化将有广阔的前景     

乙烯利对甘蔗节间过氧化物酶活性的影响及酶细胞化学

酶细胞化学研究显示：甘蔗茎杆节间薄壁细胞中过氧化物酶主要分布在细胞的液泡膜内表面和液泡液中,线粒体、叶绿体和高尔基体等细胞器膜表面也有过氧化物酶存在,而细胞核、细胞壁和胞间隙中很少或末见过氧化物酶分布。在甘蔗工艺成熟过程中,节间过氧化物酶活性是波动性变化的,不同品种表现不一致,但存在一个基础酶活性。乙烯利处理对过氧化物酶活性有影响,能提高和（或）维持较高水平的过氧化物酶活性,影响酶活性时间在处理后１～３周,因品种不同而差异。３个参试品种中有２个品种（桂１５和新台糖１６号）在乙烯利处理后酶活性表现趋势一致。     

Nitrogenase activity associated with soil cores of grasses in Brazil

The N₂-ase activities of field-grown Brazilian grasses were measured with C₂H₂ reduction by soil cores containing the plants. C₂H₂ and C₂H₄ were observed to diffuse at similar rates through soil and equilibrated across the Brazilian soil in 3 h, but could take up to 30 h or more with some British soils. The diurnal fluctuation in the rates of N₂-ase activity by Brachiaria mutica and Sorghum vulgare were similar and the variation in rate was correlated with soil temperature. Estimates of N₂-fixation by measurement of C₂H₂ reduction by soil cores ranged from 14.7 to 51.4 g N ha^?1 day^?1 and were much lower than with “pre-incubated” excised roots from the cores or taken directly from the field. The merits of the soil core and the “pre-incubated” excised root assays are discussed. ft1|Present address: U.S. Department of Agriculture, Science and Education Administration, Agricultural Research, Northeastern Region, Room 309, Building 001, Beltsville, MD 20705, U.S.A.     

Sampling of Sugarcane Leaves in Field Experiments to Determine the Activity of Nitrate Reductase

Nitrogen (N) assimilation depends on nitrate reductase activity (NRA). Leaf sampling methods in field experimentation with sugarcane and analytical procedures are essential for the determination of the maximum activity (MA) of the enzyme. Thus, in five tests, it was possible to determine which period of the crop cycle should be used for leaf collection, the most adequate leaf for collection, the ideal number of leaves to be collected, the period of the day in which the enzyme has MA and which analytical procedures can be optimized. The evaluation of NRA in sugarcane should occur between 95 and 100 days after planting (DAP) and can be performed in any green leaf. In field experiments, it is recommended the collection of 15 leaves, which must occur between 12:00 and 12:30 h, with later incubation of the leaf tissues for 430 min, without vacuum application.     

Nitrogenase Activity and Nodule Biomass of Cowpea (Vigna unguiculata L. Walp.) Decrease in Cover Crop Mixtures

Legume cover crops are often used to build soil nitrogen (N) fertility and there is increasing interest in cover crop mixtures. The objective of this mechanistic greenhouse study was to determine the effect of cover crop community diversity and soil fertility on nitrogenase activity and nodule biomass of cowpea. Cover crops were grown for 42–53 days, aboveground biomass was harvested, and nitrogenase activity was estimated with the acetylene reduction assay. Roots were then excavated to determine nodule and root biomass. Nitrogenase activity and nodule biomass per plant were greatest in cowpea monoculture and reduced by 71–98 percent in four-species mixtures. Reduced capacity for N₂ fixation was partially driven by lower cowpea biomass in mixtures. The ratio of root nodule / shoot biomass increased by 81–297 percent in low-fertility relative to high-fertility soils, which contributed to increased nitrogenase activity. Results suggest cowpea monocultures in low-fertility soils have the greatest potential for N₂ fixation.     

Nitrogenase activity associated with pasture grasses in Northern New South Wales

Nitrogenase activity associated with roots of grasses was initially examined at 67 sites in New South Wales using an enriched malate medium. Twenty six of the 39 grass species examined produced at least 10 nmol C₂H₄cm^?1 root h^?1—a level accepted as positive presumptive evidence of N₂-fixation: 40 of the 288 samples exceeded 100nmol C₂H₄h^?1. The seasonal N₂ase activity of up to 4 grass species collected in soil cores at 6 sites was measured over 16 months.Activity at field moisture levels, but incubated at 30°C was greatest for cores collected in summer months. Activity was increased in 33.5% of samples by raising soil moisture to field capacity. No single species of grass consistently supported higher activity than any other.Nitrogenase activity was compared in cores of Kikuyu grass (Pennisetum clandestinum) watered to in excess of field capacity and allowed to drain for between 7 and 28 days before re-watering. Activity declined rapidly in the first 7 days and although recovery was also rapid, integration of N₂ase activity over time showed a loss of 20 and 61% for 7 and 28 days drainage respectively. N₂ase activity was greatest at 30°C.Maximum N₂ase activity in field samples was only 246 nmol C₂H₄ core^?1 h^?1 indicating that fixation of N₂ would not be of agronomic significance.     

Nitrogenase (C2H2) Activity in Roots of Non-Cultivated and Cereal Plants: Influence of Nitrogen Fertilizer on Populations and Activity of Nitrogen-Fixing Bacteria

Root samples of 11 non-cultivated monocotyledonous and 7 dicotyledonous species taken during a wet summer had low mean nitrogenase activities of 10.2 and 7.1 nmol C₂H₄·g^?1 DW·h^?1 after preincubation at pO₂ 0.02, respectively. Maxima of 139–169 nmol·g^?1·h^?1 were observed with Agrostis vulgaris and Agropyron repens on a sandy soil poor in C_org. Three of 6 early, but none of 4 late fodder maize cultivars had a very low activity up to 0.5 nmol·g^?1h^?1. Oat, rye and wheat roots from plots with organic or mineral N fertilizers had activities between 1.3 and 7.3 nmol·g^?1h^?1 at flowering, which were not correlated with their Azospirillum populations (10²-10⁷·g^?1 after preincubation). Winter wheat and barley roots given 0, 40, 80 and 120 kg. ha^?1 NH₄NO₃-N in 0–3 applications had mean activities of 0.08, 4.06, 0.09 and 0.08 nmol or 1.77, 2.67, 0.36 and 0.23 nmol C₂H₄g^?1·h^?1 after flowering, respectively. An appreciable part of this activity could be removed by root washing. In preincubated rhizosphere soil of wheat and barley populations of N₂-fixing, facultative anaerobic Klebsiella and Enterobacter spp. were 10–100 times higher than those of Azospirillum sp., both being higher in O N than in 80 kg N·ha^?1 trials.