根际微域研究中土样采集方法的研究进展

综述了６０年代以来植物根际微域研究中土壤样品的采集方法。认为根际土样采集的发愤趋势是从田间的各采集到室内模拟实验的精细划分，再把模拟装置运用到田间，最终达到指导生产的目的。同时，随着根际概念的拓（生态界面Ｅｃｏ－ｂｏｕｎｄａｒｙ ｌａｙｅｒ），根际土样的采集朝着保持原位和更精细的方向发展。     

根系活动对湿地植物根际铁异化还原的影响及机制研究进展

根系活动是影响湿地植物根际铁异化还原速率的关键因素之一。以往国内外湿地铁异化还原的研究多为分析和比较各类中宏观生境中铁异化还原能力的差异。近年来,湿地植物根际微域铁的生物地球化学行为也日益成为该领域的研究热点。综述了根际铁异化还原研究概况,梳理了根系活动对根际铁异化还原关键因子的作用机制,分析了根际铁异化还原和其他有机质代谢途径的竞争关系,探讨了根际铁异化还原对根系活动动态变化和异质性的响应,提出了根际铁异化还原的概念模型,并指出了未来我国湿地植物根际铁异化还原研究应加强的工作。     

Microscale fumigation-extraction and substrate-induced respiration methods for measuring microbial biomass in barley rhizosphere

Changes in microbial biomass in the rhizosphere of young barley seedlings was studied. A fumigation-extraction (FE) method with measurement of ninhydrin-reactive nitrogen (NR-N) and a substrate-induced respiration (SIR) method were applied on a microscale to rhizosphere soil samples of approximately 0.1 g. Rhizosphere soil was defined as the soil adhering to the roots when they were carefully separated from the bulk soil. The rhizosphere soil was gently washed off the roots with either distilled water (FE) or with glucose solution (SIR). Shaking and mild sonication was used to disperse the soil without disrupting the roots. Fumigation was carried out by direct addition of liquid chloroform to the isolated soil. These techniques were proven to give reliable results under the experimental conditions of this investigation. Rhizosphere soil was isolated from segments of the roots representing different distances to the seed different root ages. In the rhizosphere of young barley seedlings, biomass NR-N increased significantly compared to the bulk soil from day 6 after sowing (average increases of 33–97%), especially where adventitious roots had developed. From this time, SIR rates were also significantly higher in the rhizosphere than in bulk soil (average increases 72–170%). The average ratio of SIR rate to biomass NR-N was found to be approximately 50% higher in the rhizosphere than in the bulk soil, which may indicate that a larger fraction of the microbial community is potentially active in the rhizosphere as compared to the bulk soil.     

Rhizosphere soil nitrogenase (C2H2 reduction) as influenced by plant density in intermediate deep water rice

Summary Nitrogenase activity in the rhizosphere soil of intermediate deep water rice was investigated employing gas chromatographic acetylene reduction assay. A raise in the plant density decreased the rhizosphere nitrogenase. Moreover, nitrogen fixation in the rhizosphere soil varied among the three rice varieties under intermediate deep water situations. Results indicate that nitrogen fixation is affected by plant density and the rice variety.     

Effects of freezing on rhizosphere and root nutrient content using two soil sampling methods

This study investigated how deep freezing affects the mineral nutrient content of roots and rhizosphere soil. Two different methods of separating the rhizosphere from the roots were used: i) a brushing method, where the rhizosphere soil was brushed off and ii) a washing method, where the rhizosphere was extracted together with the roots.When unfrozen material was used, the concentrations of K and Fe were significantly higher in the washing method as compared with the brushing method. When the material had been deep frozen, significantly higher concentrations of K, Fe, Mg, and Al were found in the extract from the washing method, indicating a considerable leakage from the roots. No significant differences were found between frozen and unfrozen material with the brushing method. In bulk soil, freezing resulted in increased concentrations of Mn, Al and Fe, even when no roots were present. The brushing method can be used for both frozen and fresh material, although fresh material is preferable. Extraction of soil plus roots cannot be recommended for deep frozen soil.     

大棚蔬菜轮/连作系统土壤肥力与微生物因子综合评价

从土壤肥力与微生物因子探索连茬障碍机理,以期为其提供科学依据。研究草莓番茄轮作(RST)、番茄连作4年(CT4)和番茄连作10年(CT10)3种蔬菜种植模式根际与非根际土壤微生物区系及生理菌群,并对土壤肥力与微生物生物因子进行主成分分析。结果表明根际土壤微生物三大类群和生理菌群数量均高于非根际,根际效应显著。番茄连作根际与非根际土壤细菌和放线菌数量呈现先增加后减少趋势;真菌数量呈线型增加趋势,CT4和CT10在根际与非根际较RST分别增加9.09%和2.11%、75.48%和57.72%。番茄连作根际土壤硝化细菌和好气性自生固氮菌数量的减少,氨化细菌与好气性纤维素分解菌在短期连作表现为增加长期减少的变化趋势;解钾菌、无机磷和有机磷细菌数量在根际与非根际土壤均减少。在6种研究的种植模式中,RST根际土壤状况最好,其次为CT4的根际与RST非根际土壤,CT10的根际土壤、CT4与CT10非根际土壤状况最差。结论是蔬菜连作造成土壤质量下降,连作年限越长下降越显著。     

Manganese reduction in the rhizosphere of mycorrhizal and nonmycorrhizal maize

The influence of rhizosphere microorganisms and vesicular-arbuscular (VA) mycorrhiza on manganese (Mn) uptake in maize (Zea mays L. cv. Tau) plants was studied in pot experiments under controlled environmental conditions. The plants were grown for 7 weeks in sterilized calcareous soil in pots having separate compartments for growth of roots and of VA mycorrhizal fungal hyphae. The soil was left either uninoculated (control) or prior to planting was inoculated with rhizosphere microorganisms only (MO-VA) or with rhizosphere microorganisms together with a VA mycorrhizal fungus [Glomus mosseae (Nicol and Gerd.) Gerdemann and Trappe] (MO+VA). Mycorrhiza treatment did not affect shoot dry weight, but root dry weight was slightly inhibited in the MO+VA and MO-VA treatments compared with the uninoculated control. Concentrations of Mn in shoots decreased in the order MO-VA > MO+VA > control. In the rhizosphere soil, the total microbial population was higher in mycorrhizal (MO+VA) than nonmycorrhizal (MO-VA) treatments, but the proportion of Mn-reducing microbial populations was fivefold higher in the nonmycorrhizal treatment, suggesting substantial qualitative changes in rhizosphere microbial populations upon root infection with the mycorrhizal fungi. The most important microbial group taking part in the reduction of Mn was fluorescent Pseudomonas. Mycorrhizal treatment decreased not only the number of Mn reducers but also the release of Mn-solubilizing root exudates, which were collected by percolation from maize plants cultivated in plastic tubes filled with gravel quartz sand. Compared with mycorrhizal plants, the root exudates of nonmycorrhizal plants had two fold higher capacity for reduction of Mn. Therefore, changes in both rhizosphere microbial population and root exudation are probably responsible for the lower acquisition of Mn in mycorrhizal plants.     

Functional Potential of Soil Microbial Communities in the Maize Rhizosphere

Microbial communities in the rhizosphere make significant contributions to crop health and nutrient cycling. However, their ability to perform important biogeochemical processes remains uncharacterized. Here, we identified important functional genes that characterize the rhizosphere microbial community to understand metabolic capabilities in the maize rhizosphere using the GeoChip-based functional gene array method. Significant differences in functional gene structure were apparent between rhizosphere and bulk soil microbial communities. Approximately half of the detected gene families were significantly (p<0.05) increased in the rhizosphere. Based on the detected gyrB genes, Gammaproteobacteria, Betaproteobacteria, Firmicutes, Bacteroidetes and Cyanobacteria were most enriched in the rhizosphere compared to those in the bulk soil. The rhizosphere niche also supported greater functional diversity in catabolic pathways. The maize rhizosphere had significantly enriched genes involved in carbon fixation and degradation (especially for hemicelluloses, aromatics and lignin), nitrogen fixation, ammonification, denitrification, polyphosphate biosynthesis and degradation, sulfur reduction and oxidation. This research demonstrates that the maize rhizosphere is a hotspot of genes, mostly originating from dominant soil microbial groups such as Proteobacteria, providing functional capacity for the transformation of labile and recalcitrant organic C, N, P and S compounds.     

Effects of organic amendments to soil on the rhizosphere microflora of antirrhinum infected withVerticillium dahliae Kleb.

Summary Organic (e.g. chitin, green manure, cellulose) amendments to soil induced quantitative and qualitative changes in the rhizosphere microflora of antirrhinum plants infected withVerticillium dahliae Kleb. Whereas reduction in disease severity occurred with chitin and green manure amendments, an increase in disease severity was observed with the application of cellulose. The reduction of the disease severity with chitin and green manure may be correlated with the increased population of actinomycetes in the antirrhinum rhizosphere.     

Rhizosphere bacterial community composition responds to arbuscular mycorrhiza, but not to reductions in microbial activity induced by foliar cutting

Differences in bacterial community composition (BCC) between bulk and rhizosphere soil and between rhizospheres of different plant species are assumed to be strongly governed by quantitative and qualitative rhizodeposit differences. However, data on the relationship between rhizodeposit amounts and BCC are lacking. Other soil microorganisms, e.g. arbuscular mycorrhizal fungi (AMF), may also influence BCC. We simulated foliar herbivory (cutting) to reduce belowground carbon allocation and rhizodeposition of pea plants grown either with or without AMF. This reduced soil respiration, rhizosphere microbial biomass and bacteriovorous protozoan abundance, whereas none of these were affected by AMF. After labelling plants with (13)CO(2), root and rhizosphere soil (13)C enrichment of cut plants were reduced to a higher extent (24-46%) than shoot (13)C enrichment (10-24%). AMF did not affect (13)C enrichment. Despite these clear indications of reduced rhizosphere carbon-input, denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes PCR-amplified targeting DNA and RNA from rhizosphere soil did not reveal any effects of cutting on banding patterns. In contrast, AMF induced consistent differences in both DNA- and RNA-based DGGE profiles. These results show that a reduction in rhizosphere microbial activity is not necessarily accompanied by changes in BCC, whereas AMF presence inhibits proliferation of some bacterial taxa while stimulating others.     

不同主伐方式对兴安落叶松(Larix gmelinii)根际土壤理化性质及微生物群落的影响

对内蒙古根河大兴安岭林区1987年（恢复后期）、2013年（恢复前期）的皆伐与渐伐样地以及未采伐对照样地兴安落叶松的根际土壤理化性质、微生物群落结构和多样性进行了分析,旨在揭示不同主伐方式对兴安落叶松根际土壤理化性质以及微生物群落的影响。结果表明,主伐后兴安落叶松根际土壤的理化性质以及微生物群落的变化特征与非根际土壤存在区别,且不同主伐方式在不同恢复时期会对兴安落叶松根际土壤理化性质以及微生物群落产生不同的影响：（1）根际与非根际土壤微生物群落中真菌均比细菌更容易受到土壤理化性质的影响,但是单一种理化性质的改变对根际与非根际土壤微生物群落均不能造成显著影响。（2）相较于未采伐对照样地,皆伐样地恢复前期兴安落叶松根际土壤理化性质、微生物群落结构和多样性没有显著变化。皆伐样地恢复后期,兴安落叶松根际土壤理化性质（总碳、总氮、速效氮、pH）发生了显著变化,导致了微生物量碳氮、真菌磷脂脂肪酸（PLFA）含量显著降低、细菌/真菌显著升高,辛普森多样性指数显著降低。（3）渐伐样地恢复前期兴安落叶松根际土壤总碳、总氮、速效氮含量以及含水量均显著降低,总钾、速效磷含量显著上升,根际土壤微生物量碳含量显著降低。恢复后期,兴安落叶松根际土壤总磷含量显著升高,根际土壤微生物量碳的含量已恢复到渐伐前水平。渐伐干扰对根际土壤各微生物类群PLFA含量、微生物群落结构以及多样性没有显著影响。     

Nitrate and nitrite microgradients in barley rhizosphere as detected by a highly sensitive denitrification bioassay.

A highly sensitive denitrification bioassay was developed for detection of NO3- and NO2- in rhizosphere soil samples. Denitrifying Pseudomonas aeruginosa ON12 was grown anaerobically in citrate (30 mM) minimal medium with KClO3 (10 mM) and NaNO2 (3 mM), which gave cells capable of NO2- reduction to N2O but incapable of NO3- reduction to NO2-. Growth on citrate minimal medium further resulted in the absence of N2O reduction. When added to small soil samples in O2-free vials, such cells could be used to convert the indigenous NO2- pool to N2O, which was subsequently quantified by gas chromatography. Cells grown in KClO3-free citrate medium with 10 mM NaNO3 as the electron acceptor were capable of reducing both NO3- and NO2-, and these cells could subsequently be added to the sample to convert the indigenous NO3- pool to N2O. Concentrations of both NO3- and NO2- were thus determined as N2O, with a detection limit of approximately 10 pmol of N. The bioassay could be used to determine NO3- and NO2- pools in 10-mg soil samples taken along a microgradient in the rhizosphere of field-grown barley plants. At both low (10%, wt/wt) and high (18%, wt/wt) water content, relatively high levels of NO2- were found in the rhizosphere compared with bulk soil. Under dry conditions, NO3- was also more abundant in the rhizosphere than in the bulk soil, whereas such a difference was not observed at the high water content. The roles of plant metabolism and bacterial nitrification and denitrification processes for NO3- and NO2- availability in the rhizosphere are discussed.     

Nitrate and nitrite microgradients in barley rhizosphere as detected by a highly sensitive denitrification bioassay.

A highly sensitive denitrification bioassay was developed for detection of NO3- and NO2- in rhizosphere soil samples. Denitrifying Pseudomonas aeruginosa ON12 was grown anaerobically in citrate (30 mM) minimal medium with KClO3 (10 mM) and NaNO2 (3 mM), which gave cells capable of NO2- reduction to N2O but incapable of NO3- reduction to NO2-. Growth on citrate minimal medium further resulted in the absence of N2O reduction. When added to small soil samples in O2-free vials, such cells could be used to convert the indigenous NO2- pool to N2O, which was subsequently quantified by gas chromatography. Cells grown in KClO3-free citrate medium with 10 mM NaNO3 as the electron acceptor were capable of reducing both NO3- and NO2-, and these cells could subsequently be added to the sample to convert the indigenous NO3- pool to N2O. Concentrations of both NO3- and NO2- were thus determined as N2O, with a detection limit of approximately 10 pmol of N. The bioassay could be used to determine NO3- and NO2- pools in 10-mg soil samples taken along a microgradient in the rhizosphere of field-grown barley plants. At both low (10%, wt/wt) and high (18%, wt/wt) water content, relatively high levels of NO2- were found in the rhizosphere compared with bulk soil. Under dry conditions, NO3- was also more abundant in the rhizosphere than in the bulk soil, whereas such a difference was not observed at the high water content. The roles of plant metabolism and bacterial nitrification and denitrification processes for NO3- and NO2- availability in the rhizosphere are discussed.     

Colonization of root tissues and protection against Fusarium wilt of rye (Secale cereale) by nonpathogenic rhizosphere strains of Fusarium culmorum

Colonization of rye (Secale cereale) tissues by nonpathogenic rhizosphere Fusarium culmorum isolates DEMFc2 and DEMFc5 and a pathogenic strain DEMFc37, and their effect on plant fresh weight were studied in pot experiments. Both rhizosphere isolates colonized the epidermis and the cortex but were not found in vessels, while the pathogen colonized all three layers of root cells. The numbers of pathogen CFU isolated from plant tissues were much higher than those of the rhizosphere isolates in spite of the same number of macroconidia used as inoculum (1 × 10⁵ g⁻¹ of soil). Inoculation of seedlings with DEMFc2 resulted in a 20% increase, with DEMFc5 in more than a 20% reduction, and with DEMFc37 in a 38% reduction of shoot fresh weight of 14-day-old plants. Pre-colonization of plants with (either of) the rhizosphere isolates and subsequent inoculation with the pathogen resulted in plant weights the same as those observed in plants inoculated with the rhizosphere strain alone. The disease severity index for shoots of plants pre-colonized with DEMFc2 was reduced from class 4 (86% diseased plants) observed for plants inoculated with the pathogen alone to class 2 (average of 8% diseased plants) when pre-treated with the rhizosphere strain. The CFU number of the pathogen isolated from the interior of roots of plants pre-colonized with the rhizosphere isolates was as low as 10% of the number isolated from plants inoculated with the pathogen alone. A study of in vitro interactions between the rhizosphere isolates and the pathogen suggests that changes in plant colonization by the pathogen and its effect on fresh weight of plants pre-colonized with the rhizosphere isolates were not connected with inhibition of its growth by a direct action of the rhizosphere isolates. The results suggest that strain DEMFc2 can be considered as a potential biocontrol agent.     

Effects of inorganic amendments (N,P and K) to soil on the rhizosphere microflora of antirrhinum plants infected withVerticillium dahliae Kleb

Summary A study of the inorganic amendments (N, P and K) to soil, and their effect on the rhizosphere microflora, as well as their relation to the control of wilt of antirrhinum plants caused byVerticillium dahliae Kleb. was done. Ammonium sulphate was the only chemical found to be significantly inhibitory toV. dahliae in vitro. Soil amendments (NPK) affected the rhizosphere microorganisms of the antirrhinum plants. Higher concentration of the chemicals were phytotoxic. It was further observed that ammonium sulphate, and the combined chemicals (NPK 25%) in soil delayed the senescence in healthy plants, suggests that chemical fertilisers affected the host plants directly. Addition of ammonium sulphate (0.25%), calcium nitrate (0.25%, 0.5%) combined NPK (0.25%) to soil caused considerable reduction in disease severity. It is assumed that this reduction may be caused by the (1) fungitoxic nature of the chemicali.e. ammonium sulphate, (2) antagonistic environment for the pathogen in the rhizosphere was boostedi.e. where calcium nitrate was added as soil amendments and (3) reduction in disease severity in soil-amended with combined NPK, may be due to the fact that antagonistic actinomycete population was boosted in the rhizosphere.     

Acetylene reduction in the rhizosphere of rice: Methods of assay

Summary A method is described for thein situ assay of acetylene reduction activity in the rhizosphere of rice involving the use of inexpensive, lightweight equipment. The assay is quick to set up and thus many replicates can be run simultaneously. Control of any acetylene-reducing blue-green algae in the assay chamber is achieved with a photosynthetically active herbicide which has no measurable effect on rhizosphere activity. Results using this method are compared with those from an excised root assay of the same plants. Only a very weak correlation between the two methods was found.     

Regulation of Soybean Nitrogen Fixation in Response to Rhizosphere Oxygen: II. Quantification of Nodule Gas Permeability

Nodule nitrogen fixation rates are regulated by a mechanism which is responsive to the rhizosphere oxygen concentration. In some legumes, this oxygen-sensitive mechanism appears to involve changes in the gas permeability of a diffusion barrier in the nodule cortex. In soybean evidence for such a mechanism has not been found. The purpose of this research was to make quantitative measurements of soybean nodule gas permeability to test the hypothesis that soybean nodule gas permeability is under physiological control and responsive to the rhizosphere oxygen concentration. Intact hydroponically grown soybean plants were exposed to altered rhizosphere oxygen concentrations, and the nodule gas permeability, acetylene reduction and nodule respiration rates were repeatedly assayed. After a change in the external oxygen concentration, nitrogenase activity and nodule respiration rates displayed a short-term transient response after which the values returned to rates similar to those observed under ambient oxygen conditions. In contrast to steady-state nitrogenase activity and nodule respiration, nodule gas permeability was dramatically affected by the change in oxygen concentration. Decreasing the external oxygen concentration to 0.1 cubic millimeter per cubic millimeter resulted in a mean increase in nodule gas permeability of 63%. Increasing the rhizosphere oxygen concentration resulted in decreased nodule gas permeability. These data are consistent with the hypothesis that soybean nodules are capable of regulating nitrogen fixation and nodule respiration rates in response to changes in the rhizosphere oxygen concentration and indicate that the regulatory mechanism involves physiological control of the nodule gas permeability.     

根系分泌物研究方法(综述)

根系分泌物是植物与土壤进行物质、能量与信息交流的重要载体物质.研究根系分泌物的种类、数量对了解植物与土壤及根际微生物相互作用机理具有重要指导意义,根系分泌物研究方法的选择对研究结果有重要的影响.本文着重讨论根系分泌物收集、分离纯化及鉴定的常用方法,并列举特定根系分泌物的研究方法,旨在为根系分泌物的研究提供借鉴和参考.     

Biological control of anthracnose of chilli with rhizosphere and rhizoplane fungal isolates from grasses

The fungal species from rhizosphere and rhizoplane of perennial grasses of the Western Ghats of India were studied for their pathogenicity, antagonism in vitro, substrate and root colonization abilities, rhizosphere competence, growth in different soil pH and inoculum shelf-life. Out of 138 non-pathogenic fungal isolates tested, 85 were antagonistic in vitro to chilli anthracnose pathogen Colletotrichum capsici. Fifteen isolates with >60% inhibition zone to pathogen culture had saprophytic and root and rhizosphere colonization abilities. The sorghum grain inocula of test antagonistic fungi- Fusarium oxysporum, Chaetomium globosum and Trichoderma harzianum had the shelf-life of 90 days at 20?±?2?°C and required optimum soil pH of 6.5. The above fungal isolates when tested for biocontrol of anthracnose disease in greenhouse and field caused reduction in seedling mortality and decreased disease incidence and severity at various plant growth stages and significant reduction in chilli fruit and seed infection. The test antagonistic fungi promoted seedling and mature plant growth and increased fruit and seed yield. Populations of these antagonistic fungi were fairly high in chilli rhizosphere at harvest. The present study indicated that antagonistic fungi from grass rhizosphere and rhizoplane could be used to control anthracnose and promote plant growth, and increase yield of chilli in field.     

Combined use of colorimetric and microelectrode methods for evaluating rhizosphere pH

Plant control of rhizosphere pH is important for nutrient mobilization and uptake, and also affects microbial activity and pathogens in the vicinity of the root. Limited information is available on the ability of plant species and genotypes within a species to induce pH changes in the rhizosphere. A growth chamber study was conducted to characterize patterns of pH change within the rhizosphere of selected genotypes in an alkaline environment with a balanced nutrient supply. After germination in incubators, seedlings of 32 genotypes of maize (Zea mays L.), soybean (Glycine max. L.), sorghum (Sorghum bicolor L.), sordan [sorghum (Sorghum bicolor L.), sudangrass (Sorghum sudanese L.) hybrid], wheat (Triticum aestivum L.), oats (Avena sativa L.), and barley (Hordeum vulgare L.) were transferred into aseptic agar medium (pH 7.6) with bromocresol purple indicator. Ability of the embedded roots to induce rhizosphere pH change was followed by photographing the color change of the bromocresol purple indicator. The pH for selected genotypes at different root zones (maturation, elongation, meristematic) was also monitored by a microelectrode at 1-, 2-, 3- and 4-mm distances from the root surface. Rhizosphere acidification for selected genotypes within a species were in the order: soybean, Hawkeye>PI-54169; maize, Pioneer-3737>Pioneer-3732>CM-37; sordan, S-757>S-333; sorghum, SC-33-8-9EYSC-118-15E; barley, Bowman>Primus II; oats, Hytest>SD-84104. The pH patterns within the root system varied from species to species. The highest amount of acidification was found at the elongation and meristematic zones for soybean, while the highest amount of acidification was found at the maturation zone for barley under the same experimental conditions. The agar method allowed the determination of a genotype's capability to induce rhizosphere pH changes while the microelectrode method is necessary for quantifying the spatial variation of specific root developmental zones with high resolution.This work is a part of H.T. Gollany's dissertation in partial fulfillment of the requirements for the PhD degree.