Effects of Pseudomonas aeruginosa on the diversity of culturable microfungi and nematodes associated with tomato: impact on root-knot disease and plant growth

The available information on Pseudomonas biocontrol inoculants on the non-target fungal and nematode community is scant. The current paper addresses this issue and investigates the effects of biocontrol agents Pseudomonas aeruginosa IE-6 and IE-6S⁺ (previously shown to suppress several soil-borne plant pathogens) on soil microfungi and plant-parasitic nematodes as well as on the root-knot development and growth of tomato (Lycopersicon esculentum). Furadan, a granular nematicide was included as a treatment for comparative purposes. Treatments were applied to soil at the start of each 52-day-long tomato growth cycle, and their effects on the composition and diversity of rhizosphere and endophytic microfungi and plant-parasitic nematodes were examined at the end of first and fourth growth cycle. Several diversity indices were employed to assess community diversity. A total of 16 genera comprising 23 microfungal species were isolated from the tomato rhizosphere. The most abundant fungal species belonged to the genera Aspergillus, Fusarium, and Penicillium. With a few exceptions, fungi were neither exclusively inhibited nor specifically promoted by the application of treatments at any of the growth cycles studied. However, Paecilomyces lilacinus, an egg and female parasite of root-knot nematode, though exclusively absent in the controls was isolated from the treatments. Both general diversity and equitability of rhizosphere microfungi were greater at first compared to the fourth growth cycle while species richness remained uninfluenced across the growth cycles and treatments. However, Furadan and IE-6S⁺ treatments considerably abated general diversity and equitability. Of the microfungal species isolated from the rhizosphere seven were also recovered from surface-sterilized root tissue of tomato suggesting that all the endophytes are primarily rhizosphere organisms. Diversity of endophytic fungi was consistently lower compared with that of the rhizosphere. Both general diversity and equitability declined in all three treatments relative to controls in the root tissue but species richness remained unaltered. Diversity and equitability of plant-parasitic nematodes in soil were reduced by all three treatments over the controls at fourth growth cycle whilst species richness did not change at either growth cycle. The biocontrol agents significantly reduced root-knot development and enhanced shoot growth of tomato over the controls. The possible implications of fungal composition and abundance because of biocontrol by Pseudomonas application are discussed.     

粘土矿物对土壤腐殖质化学性质的影响

Chemical characteristics of humic substances in soils with different mineralogical characteristics and under different utilization paterns in Zhangpu,Fujian Province,together with two pairs of cultivated soils in North China Plain were studied by chemical analysis,visible and IR spectroscopy and ^13C NMR spectrometry.For soils in Zhanpu the HA/FA ratio and both the aromaticity and the degree of humification of HA were higher in soils with montmorillonite as the predominant clay mineral than in those with kaolinite as the predominant clay mineral,provided these soils were under the same utilization pattern.While for each pair of soils with similar mineralogical characteristics the HA/FA ratio was higher and the C/H ratio and the contnet of carboxyl group of HA were lower in paddy soil than in upland soil.Among the upland soils(or paddy soils)studied the Ha/FA ratio of soil in Zhangpu with kaolinite as the predominant clay mineral was the lowest,and that of soil in Zhangpu with montmorillonite as the predominant clay mineral was the highest .the lowest.and that of soil in Zhangpu with montmorillonite as the predominant clay mineral was the highest It was concluded that the presence of montmorillonite favored the fromation and maturation of humic acid.     

Ligninolytic fungi in bioremediation: extracellular enzyme production and degradation rate

Ligninolytic fungi can be used for remediation of pollutants in water and soil. Extracellular peroxidases and laccases have been shown to oxidize recalcitrant compounds in vitro but the likely significance of individual enzyme levels in vivo remains unclear. This study documents the amounts and activities of Mn-dependent peroxidase (MnP), lignin peroxidase and laccase (LAC) in various species of ligninolytic fungi grown in liquid medium and soil and their effect on degradation of polycyclic aromatic hydrocarbons (anthracene and pyrene), a polychlorinated biphenyl mixture (Delor 106) and a number of synthetic dyes. Stationary cultures of a highly degradative strain Irpex lacteus exhibited 380-fold and 2-fold increase in production of MnP and LAC, respectively, compared to submerged cultures. Addition of Tween 80 to the submerged culture increased MnP levels 260-fold. High levels of MnP correlated with efficient decolorization of Reactive Orange 16 azo dye but not of Remazol Brilliant Blue R anthraquinone dye. Degradation of anthracene and pyrene in spiked soil by straw-grown explorative mycelium of Phanerochaete chrysosporium, Trametes versicolor and Pleurotus ostreatus showed the importance of MnP and LAC levels secreted into the soil. The importance of high fungal enzyme levels for efficient degradation of recalcitrant compounds was better demonstrated in liquid media compared to the same strains growing in soil.     

土壤中钙结合和铁铝结合腐殖质的组成及结构特征

Calcium-bound and iron- and aluminium-bound humus extracted from different soils collected from north to south of China were characterized by chemical and spectroscopic methods. Meaningful differences in the composition and structure between them were revealed by ¹³C NMR, visible spectroscopy and elemental analysis. Results showed that the contents of carbon, hydrogen and nitrogen were higher in iron- and aluminium-bound humus than in calcium-bound humus while oxygen content in calcium-bound humus was shown to be higher. The calcium-bound humus had higher C/N and O/C ratios than iron- and aluminium-bound humus. The calcium-bound humic acid (HA1) showed higher E₄/E₆ ratios than iron- and aluminum-bound humic acid (HA2) while iron- and aluminum-bound fulvic acid (FA2) showed higher E₄/E₆ ratios than calcium-bound fulvic acid (FA1). An inverse relationship between E₄/E₆ ratios and aromaticity as determined by ¹³C NMR spectra was observed for HA and FA from black soil. The ¹³C NMR spectroscopy revealed that HA2 was more aromatic than HA1. On the other hand, FA1 exhibited a higher aromaticity than FA2.     

中国黑土上腐殖酸和腐殖物质的提取及其描述

Twenty-three progressive extractions were performed to study individual humic acids (HAs) and humin fractions from a typical black soil (Mollisol) in Heilongjiang Province, China using elemental analysis and spectroscopic techniques. After 23 HA extractions the residue was separated into high and low organic carbon humin fractions. HA yield was the highest for the first extraction and then gradually decreased with further extractions. Organic carbon (OC) of the humin fractions accounted for 58% of total OC even after 23 successive HA extractions. In addition, the atomic C/H ratio decreased during the course of extraction while C/O increased; the E4/E6 ratio from the UV analysis decreased with further extraction while E~/E3 increased; the band assigned to aliphatic carbon (2 930 cm-1) in the diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS) spectra gradually increased with progressive extraction; the calculated ratio of the sum of aromatic carbon peak heights to that of aliphatic carbon peak heights from DRIFTS spectra declined with extractions; and nuclear magnetic resonance (NMR) data suggested that HA aliphatic carbons increased with extractions while aromatic carbons decreased. Thus, hydrophobicity and aliphaticity of HAs increased with extractions while polarity and aromaticity decreased. These data showed substantial chemical, structural, and molecular differences among the 23 HAs and two humin fractions. Therefore, these results may help explain why soil and sediment humin fractions have high sorption capacity for organic contaminants.     

Comparison of soil microfungi in 40-year-old stands of pure alder,pure conifer,and alder-conifer mixtures

The species composition of soil microfungal populations in adjacent stands of red alder Alnus rubra, conifers Pseudotsuga menziesii, Tsuga heterophylla, and Picea sitchensis, and mixed alder conifer correlated strongly with the dominant vascular vegetation. A total of 92 species were isolated: 55 from the alder stand; 45 from the conifers; and 46 from the mixed alder-conifer, with few species (16.7, and 5 in the three plots, respectively) reaching average frequencies of 50 per cent or higher. Penicillium nigricans, Aureobasidiiim pullulans, Cephalosporium curtipes, and Cladosporium herbarium were present with high frequency at all sites. Penicillium daleae, which occurred with a frequency of 83 per cent in alder soil, appears to be a rare fungus elsewhere. There was little difference in species composition among soil horizons within a stand.     

Collembolan dietary specialisation on soil grown fungi

Preferences of six collembolans for eight species of microfungi grown in soil have been investigated. Collembolans (Folsomia fimetaria, Isotoma anglicana, Isotama notabilis, Heteromurus nitidus, Protaphorura armata, Pseudosinella alba) and microfungi where chosen for the experiment on the basis of their abundance and vertical distribution in an organically grown field where all organisms and soil were collected. Collembolans were tested with pairs of different species of microfungi and a control (sterile soil) in petri dishes and their position determined after 90 min. Based on the first two axes in a Principal Component Analysis we could identify two main feeding patterns in the collembolans: (1) their general acceptance of the fungi (the mean value of the preference indices for all eight fungi), and (2) their alternating preference for Cladosporium herbarum and Fusarium culmorum (high preference index for C. herbarum and low for F. culmorum or vice versa). The six collembolan species in our study combined these two feeding patterns with an intraspecific preference for the eight fungal species and seemed to minimise food competition among collembolans co-occurring in the same soil horizons. Our data suggest that differences in preference between collembolan species may help to explain the coexistence of many species in the same microhabitats of the soil.     

The microbial biomass and activity in soil with shrub (Caragana korshinskii K.) plantation in the semi-arid loess plateau in China

Caragana korshinskii K. is a shrub species which is adapted to arid and semi-arid environments and plays an important role in soil protection. The objective of this study was to determine the influence of this shrub plantation on the soil ecosystem functions driven by microorganisms in the long-term. The changes in the size and activity of soil microbial biomass and the relationship between soil microbial biomass and chemical properties were investigated under shrub plantations aged 6, 18 and 26 years. The results showed that the pH value in the soil decreased gradually, while soil organic carbon (OC) and total nitrogen (TN) significantly increased with the age of C. korshinskii. Although microbial biomass carbon (MBC) and MBC/OC ratio gradually increased, the ratio of basal respiration to MBC (qCO₂) decreased with the age of C. korshinskii. The microbial biomass nitrogen (MBN) and MBC had a positive relationship with soil TN and OC, respectively. The flux of CO₂ decreased with the age of C. korshinskii which had a significant negative relationship with soil OC, TN, MBN and MBC. The results indicate that C. korshinskii plantations may help to improve microbially driven ecosystem functioning through long-term creation of resource-island.     

Biotransformation of soil humic acids by blue laccase of Panus tigrinus 8/18: an in vitro study

We describe here the role of the polyphenoloxidases in the oxidation of recalcitrant soil organic compounds and consider what changes occur in their structure during experiments on the biotransformation of soil and peat-derived humic acids (HA). These transformations were carried out by laccase (EC 1.10.3.2.) of the white-rot basidiomycete Panus tigrinus 8/18. It was shown that purified laccase alone is capable both of polymerizing and depolymerizing HA in vitro. The direction of transformations depends on the nature and properties of HA. Those fractions of HA are affected by laccase, which cause the lowest inhibitory effect on the enzyme. Contrary to previous studies depolymerization of HA was not necessarily accompanied by decolorization: chernozem-derived HA showed increase in absorbance of its aqueous solution in the region of 240-500 nm during depolymerization, while peat-derived HA showed decrease in absorbance throughout the entire spectrum during polymerization. All studied HA were competitive inhibitors of laccase towards oxidation of synthetic substrate 2,2′-azino-bis-(3-ethylthiazoline-6-sulfonate) (ABTS). When studying the nature of the inhibitory effect, it was shown that more ‘hydrophobic’ HA as well as more ‘hydrophobic’ HA fragments were stronger inhibitors of blue laccase.     

Diversity of soil basidiomycete communities associated with Quercus suber L. in Portuguese montados

The montado is an agro-silvo pastoral system characterized by an open oak formation combined with shrubs or a rotation of cultures/pastures/fallow. A key, but frequently overlooked, component of these systems is the fungal community associated with the dominant oaks. We present a study of the basidiomycete community in montado areas of Quercus suber L. from Central-Western Portugal, based on fruiting body assessment and PCR-DGGE of soil mycelium. Soil available phosphorus and organic matter content were also determined. The most frequent fruiting body found in the cork oak montados belonged to Lactarius, Clitocybe, Russula and Cortinarius species. Lactarius chrysorrheus was the most widely distributed species. Soil management practices cause a temporal increase in soil available phosphorus, and harrowing and fertilizer application also result in an increase in soil organic matter content. Mechanical clearing with recent soil disturbance had a negative effect on the richness and diversity of the basidiomycete community assessed using fruiting body. The molecular analysis of the basidiomycete community separated the sites with recent shrub-removal from the others sites. Our results show that soil management techniques that avoid disturbance of the top-soil layers are the best way to preserve the structure and diversity of the soil fungal community in the montado.     

我国几种土壤中腐殖质性质的研究

研究了几种土壤腐殖酸的基本性质,其中包括土壤的腐殖质组成,胡敏酸以及富里酸的元素组成,含氧官能团含量,光密度值与芳化度等,计算了E₄与E₄/E₆比值与上述性质的相关系数。结果表明土壤腐殖质组成和性质与其形成条件有着密切的联系,并且有一定的地带规律性。水稻土中有机质含量增加,HA/FA比值变高,其活性HA显著降低,说明水稻土有利于有机质的积累并改变了腐殖质的组成。E₄、E₄/E₆比值与元素组成,C/H比值,含氧官能团及芳化度之间的相关系数表明E₄除了与C%,O%,酚羟基-O%,醌基-O%呈极显著或显著相关外,还与C/H比值及芳化度呈极显著相关,因此E₄值可作为腐殖物质芳化度的一个指标。     

Carbon and nitrogen in operationally defined soil organic matter pools

Humic substances [humic acid (HA), fulvic acid (FA), and insoluble humin], particulate organic matter (POM), and glomalin comprise the majority (ca 75%) of operationally defined extractable soil organic matter (SOM). The purpose of this work was to compare amounts of carbon (C) and nitrogen (N) in HA, FA, POM, and glomalin pools in six undisturbed soils. POM, glomalin, HA, and FA in POM, and glomalin, HA, and FA in POM-free soil were extracted in the following sequence: (1) POM fraction separation from the soil, (2) glomalin extraction from the POM fraction and POM-free soil, and (3) co-extraction of HA and FA from the POM fraction and POM-free soil. Only trace amounts of HA and FA were present in the POM fraction, while POM-associated glomalin (POM-glomalin) and POM alone contributed 2 and 12%, respectively, of the total C in the soil. Mean combined weights for chemically extracted pools from POM and from POM-free soil were 9.92 g glomalin, 1.12 g HA, and 0.88 g FA kg⁻¹ soil. Total protein and C, N, and H concentrations showed that glomalin and HA were, for the most part, separate pools, although protein was detected in HA extracts. Even though percentage carbon was higher in HA than in glomalin, glomalin was a larger (almost nine times) operationally defined pool of soil organic C. Glomalin was also the largest pool of soil N of all the pools isolated, but all pools combined only contained 31% of the total N in the soil.     

Fulvic acid in foliar spray is more effective than humic acid via soil in improving coffee seedlings growth

ABSTRACT

Humic (HA) and fulvic (FA) acids improve the nutrient availability and uptake by plants but some aspects of their agronomic use still need to be clarified. The effects of HA soil application and FA foliar application on the growth, Zn and B uptake by coffee seedlings were evaluated. HA was added to an Oxisol at concentrations 0, 10, 25, 50, 75 and 100 mg kg^?1 (C-HA), in both limed (pH 6.2) and overlimed (pH 7.2) conditions. FA (0, 0.2, 0.5 and 1 g L^?1 C-FA) was applied to coffee leaves in three different application modes (M): with 0.3% Zn and 0.6% B supplied via foliar (M₁), 0.6% B and 1.2% Zn supplied via foliar (M₂) and 1.2 mg kg^?1 B and 6 mg kg^?1 Zn supplied via soil (M₃). HA addition in soil significantly (p < 0.05) reduced leaf B and Zn accumulation and coffee growth in both pH conditions. In the M₁ and M₂, FA application significantly (p < 0.05) increased the shoot growth at 0.59 and 0.45 g L^?1 and B accumulation at 0.96 and 0.45 g L^?1 C-FA. Foliar application of C-FA, instead soil application of C-HA, is a suitable practice to improve coffee seedlings growth and nutrition on Oxisol.     

连续提取对土壤腐殖质组分数量与特性的影响

以暗棕壤为供试土壤,研究了NaOH和NaOH+Na4P2O7连续提取对土壤腐殖质组分数量和特性的影响。结果表明:用两种提取剂得到的土壤腐殖质组分总量相差不大,并且其中HA组分的数量均高于FA组分。随提取次数增加,胡敏酸(HA)、富里酸(FA)和胡敏素(HM)的绝对数量均下降,HA、FA的相对数量也下降,而HM的相对数量则增加。以NaOH为提取剂时,最后得到的腐殖质组分中FA的相对比例高于HA;而用NaOH+Na4P2O7为提取剂时,最后得到的腐殖质组分中HA的相对比例则高于FA。连续提取过程中,HA、FA的色调系数(Δlogk)和活化度(AD)通常呈增加趋势,而相对色度(RF)下降,说明后提取出的HA、FA的缩合度较低,脂族性较强。用RF来表征HA、FA特性变化更为合适。     

Clonal and seasonal shifts in communities of saprotrophic microfungi and soil enzyme activities in the mycorrhizosphere of Salix spp.

The species‐specific microbial root and rhizosphere colonization contributes essentially to the plant nutrient supply. The species number and colonization densities of cultivable saprotrophic microfungi and the activities of nutrient‐releasing soil enzymes (protease, acid and alkaline phosphatase, arylsulfatase) were investigated in the rhizosphere of one low mycorrhizal (Salix viminalis) and one higher mycorrhizal (S. × dasyclados) willow clone at a Eutric Cambisol in N Germany. After soil washing, in total 32 and 28 saprotrophic microfungal species were isolated and identified microscopically from the rhizosphere of S. viminalis and S. × dasyclados, respectively. The fungal species composition changed within the growing season but the species number was always lower under S. × dasyclados than under S. viminalis. Under both willow clones, the fungal colonization density was largest in spring, and the species number was largest in autumn. Acid‐phosphatase activity (p < 0.001) and protease activity (p < 0.003) were significantly affected by the Salix clone, whereas arylsulfatase and alkaline‐phosphatase activities did not show clone‐specific differences. All enzyme activities reached their maxima in the summer sampling. Rhizosphere colonization with Acremonium butyri, Cladosporium herbarum, and Penicillium janthinellum contributed significantly to explain the activities of acid phosphatase. Rhizosphere colonization with Cylindrocarpon destructans, Penicillium spinulosum, Plectosphaerella cucumerina, and Trichoderma polysporum contributed significantly to explain the arylsulfatase activities. Effects of the saprotrophic fungal colonization densities on the protease activities in the rhizosphere were low. Acid‐ and alkaline‐phosphatase and arylsulfatase activities in the rhizosphere soil were stronger affected by the composition of the saprotrophic fungal communities than by the Salix clone itself. In conclusion, the colonization density of some saprotrophic microfungi in the rhizosphere contributed to explain shifts in soil‐enzyme activities of the P and S cycles under different willow clones.     

红壤复合胶体腐殖质的特性

对闽北6个样品测定结果表明,红壤复合胶体中C、N含量较高,其腐殖质的C/N和HA/FA、胡敏酸的芳构化度和腐殖质氧化稳定性系数均较重组小,而松结合态/紧结合态腐殖质的比值则较大.这些特性有利于土壤养分保存与供应和改善土壤结构.     

不同土地利用方式对黑土胶散复合体的影响及与酸度的关系

以吉林省黑土玉米带区域内10对长期定位采样点的2种土地利用方式土壤(耕作黑土和防护林带黑土)为研究对象,采用胶散分组法提取土壤胶散复合体,探究长期不同利用方式对黑土胶散复合体和酸度的影响。结果表明:耕作黑土与防护林带黑土均表现为G_0与G_2组的绝对含量增加,G_1组的绝对含量下降,复合体总量分别下降了3.88%和0.28%。耕作黑土和防护林带黑土pH分别平均下降12.12%和3.77%,有机碳含量在耕作黑土和防护林带黑土中分别平均下降23.17%和9.00%,2种利用方式土壤的胡敏酸、富里酸、交换性钙离子和碳酸钙均出现下降;耕作黑土分别平均下降12.18%,3.53%,16.29%,31.53%;防护林带黑土分别平均下降20.07%,13.14%,2.81%,7.81%。G_1组与pH、HA、Ca~(2+)和CaCO_3呈极显著正相关,与FA呈显著负相关;G_2组与pH、HA、Ca~(2+)和CaCO_3呈极显著负相关。pH与HA、Ca~(2+)和CaCO_3呈极显著正相关,与FA呈显著负相关。经过11年的变化,黑土胶散复合体稳定性下降,相比非耕作状态下黑土,耕作模式下的黑土胶散复合体稳定性下降更为严重,胶散复合体总量下降的更多,酸度下降更为明显。Ca~(2+)和HA的变化是影响黑土土壤酸化和土壤团聚体稳定性的主要驱动因素。土壤酸化的结果使土壤胶散复合体数量减少,稳定性变差。     

Inhibitory action of dissolved humic substances on the growth of soil bacteria degrading DDT

Abstract

Dissolved humic acid (HA) and fulvic acid (FA) prepared from a Dando brown forest soil (Typic Dystrochrept) inhibited the growth of soil bacteria degrading DDT (1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane) in the culture. The population of DDT-degrading Gram-variable rod Bll6 decreased by the application of both HA and FA, suggesting the presence of bactericidal effect. Such inhibitory effect was stronger for HA and resulted in a lower degrading activity of DDT in the culture of Bll6. No inhibitory effect was observed on the growth of DDT-degrading Bacillus sp. B75. The electron spin resonance spectra showed the presence of organic free radicals in both HA and FA. The relative concentration of the radicals was higher in HA. Storage of HA solution for 3 months at 4°C decreased the concentration of the radicals as well as the inhibitory action. The addition of catalase decreased the inhibitory effect of humic acid. It is suggested that a hydroxy radical, which is derived from free radicals of humic substances, is involved in the inhibition of bacterial growth and degradation of DDT.     

Rhizodeposits of Trifolium pratense and Lolium perenne: their comparative effects on 2,4-D mineralization in two contrasting soils

Rhizosphere enhanced biodegradation of organic pollutants has been reported frequently and a stimulatory role for specific components of rhizodeposits postulated. As rhizodeposit composition is a function of plant species and soil type, we compared the effect of Lolium perenne and Trifolium pratense grown in two different soils (a sandy silt loam: pH 4, 2.8% OC, no previous 2,4-D exposure and a silt loam: pH 6.5, 4.3% OC, previous 2,4-D exposure) on the mineralization of the herbicide 2,4-D (2,4-dichlorophenoxyacetic acid). We investigated the relationship of mineralization kinetics to dehydrogenase activity, most probable number of 2,4-D degraders (MPN_2,4-D) and 2,4-D degrader composition (using sequence analysis of the gene encoding α-ketoglutarate/2,4-D dioxygenase (tfdA)). There were significant (P<0.01) plant-soil interaction effects on MPN_2,4-D and 2,4-D mineralization kinetics (e.g. T. pratense rhizodeposits enhanced the maximum mineralization rate by 30% in the acid sandy silt loam soil, but not in the neutral silt loam soil). Differences in mineralization kinetics could not be ascribed to 2,4-D degrader composition as both soils had tfdA sequences which clustered with tfdAs representative of two distinct classes of 2,4-D degrader: canonical R. eutropha JMP134-like and oligotrophic α-proteobacterial-like. Other explanations for the differential rhizodeposit effect between soils and plants (e.g. nutrient competition effects) are discussed. Our findings stress that complexity of soil-plant-microbe interactions in the rhizosphere make the occurrence and extent of rhizosphere-enhanced xenobiotic degradation difficult to predict.