Microbial synthesis of organic and condensed forms of phosphorus in acid and calcareous soils

The potential for microorganisms to affect the quantity and quality of organic and condensed forms of phosphorus (P) in soils was investigated by repeated addition of different carbon sources (glucose, starch, cellulose; 2.5 g C kg^?1) with or without inorganic P (50 mg P kg^?1) to acid and calcareous soils which were either natural soils or clay–sand mixtures free of organic matter. Forms of P after five amendments and subsequent incubation periods of 5 weeks each were analyzed by ³¹P solution nuclear magnetic resonance (NMR) spectroscopy, and the microbial community composition was assessed by selective plate counts and fatty acid methyl ester (FAME) analysis. All carbon additions induced a redistribution of P from inorganic to organic and condensed forms, which was only little affected by the addition of inorganic P. Compared to non-carbon-amended controls, the greatest increase (7–38 mg P kg^?1) in organic P was observed in the monoester region. In the acid clay–sand mixture, there was a large accumulation of pyrophosphate (101 mg P kg^?1) after glucose addition and smaller accumulations (6–25 mg P kg^?1) after addition of starch and cellulose. Carbon additions increased the microbial biomass in all cases and except in the natural calcareous soil also the proportion of fungi. Redundancy analysis with Monte Carlo permutation tests revealed that for carbon-amended soils, the microbial community composition was more strongly influenced by soil type than by carbon source. Pyrophosphate was positively related to fungi, and diester P was positively related to soil pH. A large proportion of organic and condensed forms of P may still have been in microbial cells at the time of extraction. We have shown that soil organic P consists of some discrete and simple compounds along with some more complex forms, and that organic P recently synthesized by microbes consists almost exclusively of and thus is a likely source for the simple compounds found in natural soils.     

Interrelationships between microbial and soil properties in young volcanic ash soils of Nicaragua

The activity and biomass of soil microorganisms were measured in soils from 25 different arable sites in the Pacific region of Nicaragua with the objective of elucidating their interrelationship with soil textural and soil chemical properties. All soils developed from recent volcanic deposits but differ in their particle size distribution. Short-term phosphorus fixation capacity varied widely and was, on average, 11% of added P. In contrast, long-term P fixation capacity varied within a small range of around 55%. Mean basal respiration was 8.6 μg CO₂–C d⁻¹ g⁻¹ soil, average contents of biomass C, biomass P, and ergosterol as an indicator of fungal biomass were 116, 1.95, and 0.34 μg g⁻¹ soil, respectively. They were all, except biomass P, significantly lower in the sandy than in the loamy soils. The mean biomass C-to-soil C ratio was 0.69%, the mean metabolic quotient 95 mg CO₂–C d⁻¹ g⁻¹ biomass C, the mean ergosterol-to-biomass C ratio 0.31% and the mean biomass C-to-P ratio 107. The very low ergosterol-to-biomass C ratio indicates that fungi contribute only a relatively small percentage to the microbial biomass. The biomass C-to-P ratio exceeded considerably the soil C-to-total P ratio. Metabolic quotient qCO₂ and ergosterol-to-biomass C were both negatively correlated with biomass C-to-soil C ratio and clay content, indicating positive correlations between qCO₂ and ergosterol-to-biomass C ratio and between biomass C-to-soil C ratio and clay content. Key problems of soil fertility and soil quality in Nicaragua are low availability of soil organic matter and phosphorus to soil microorganisms, which are magnified by a low percentage of fungi, probably reducing the ability of soil to provide nutrients for plant growth.     

Isotope ratio mass spectrometry identifies soil microbial biocontrol agents having trophic relations with the plant pathogen Armillaria mellea

An understanding of the types of interactions that take place between plant pathogens and other microorganisms in the natural environment is crucial in order to identify new potential biocontrol agents. The use of microorganisms labelled with stable isotopes is a potentially useful method for studying direct parasitisation of a given pathogen or assimilation of the pathogen's metabolites by microorganisms. A microorganism labelled with a stable isotope can be monitored in the environment and isotope ratio mass spectrometry can detect whether it is directly parasitised or its metabolites are used by other microorganisms. In this study, we isolated 158 different species of fungi and bacteria from soil and assayed their biocontrol potential against a plant pathogen (Armillaria mellea) by coupling a dual-culture test with mass spectrometry analysis of the ¹³C isotope in the microorganisms in presence of ¹³C-labelled A. mellea. The microorganisms affected the pathogen by means of antibiosis phenomena (total or partial inhibition of pathogen growth, alteration of its morphology) and by antagonism, probably resulting from competition for space and nutrients or from mycoparasitism. Isotope ratio mass spectrometry was used to identify direct trophic interactions between microorganisms and the pathogen as in dual cultures as in soil microcosms. Six fungi and one bacterium were found to display the best active trophic behaviour against the pathogenin dual cultures; three microorganisms were discarded due to their plant pathogen potential. Trichoderma harzianum, Pseudomonas fluorescens and Rhodosporidium babjevae were selected to carry out the experiments. T. harzianum inhibited pathogen development (rate of inhibition 80 ± 0.19%) and its δ ¹³C values increased (244.03 ± 36.70‰) in contact with ¹³C-labelled A. mellea. Lower levels of antagonism and correspondingly lower assimilation of ¹³C were detected in P. fluorescens and R. babjevae. Only T. harzianum maintained mycoparasitic activity in the soil microcosm, showing a δ ¹³C value of 1.97 ± 2.24‰ after one month in co-presence with the labelled pathogen. This study provides support for the use of isotope ratio mass spectrometry as an additional tool in screening for potential biocontrol agents.     

A genotype dependent-response to cadmium contamination in soil is displayed by Pinus pinaster in symbiosis with different mycorrhizal fungi

Soil contamination with Cd is of primary concern and beneficial soil restoration strategies urge. The aim of this work is to evaluate the response of two different genotypes of Pinus pinaster (wild and selected) to Cd contamination and to assess how inoculation with ectomycorrhizal fungi, Suillus bovinus and Rhizopogon roseolus, influenced each genotype. Seedlings were exposed to soil contaminated at 15 and 30 mg Cd kg⁻¹. Plant growth, mycorrhizal traits and Cd accumulation in different tissues of the plant were determined at harvest. The fungal community was assessed by denaturing gradient gel electrophoresis. At 15 mg Cd kg⁻¹ S. bovinus increased aboveground development in both genotypes. At 30 mg Cd kg⁻¹ non-inoculated wild genotype accumulated more Cd in the shoots (1.7-fold) than the selected genotype; inoculation with R. roseolus decreased Cd concentration in the roots of the selected genotype whereas the opposite occurred in the wild genotype. Cd concentration in the root system was the parameter most influenced by the interaction between the three studied variables. The fungal community established was affected by the Cd concentration in the soil. Results show that different genotypes of P. pinaster react differently to Cd exposure depending on the mycorrhizal association. The importance of considering the combination between plant genotype and its symbiotic partners when aiming at the forestation of degraded land is highlighted.     

Accumulation of As,Pb, Zn,Cd and Cu and arbuscular mycorrhizal status in populations of Cynodon dactylon grown on metal-contaminated soils

Metal(loid) accumulation and arbuscular mycorrhizal (AM) status of the dominant plant species, Cynodon dactylon, growing at four multi-metal(loid)s-contaminated sites and an uncontaminated site of China were investigated. Up to 94.7 As mg kg^?1, 417 Pb mg kg^?1, 498 Zn mg kg^?1, 5.8 Cd mg kg^?1 and 27.7 Cu mg kg^?1 in shoots of C. dactylon were recorded. The plant was colonized consistently by AM fungi (33.0–65.5%) at both uncontaminated site and metal-contaminated sites. Based on morphological characteristics, fourteen species of AM fungi were identified in the rhizosphere of C. dactylon, with one belonging to the genus of Acaulospora and the other thirteen belonging to the genus of Glomus. Glomus etunicatum was the most common species associated with C. dactylon growing at metal-contaminated sites. Spore abundance in the rhizosphere of C. dactylon growing at the metal-contaminated soils (22–82 spores per 25 g soil) was significantly lower than that of the uncontaminated soils (371 spores per 25 g soil). However, AM fungal species diversity in the metal-contaminated soils was significantly higher than that in the uncontaminated soils. This is the first report of AM status in the rhizosphere of C. dactylon, the dominant plant survival in metal-contaminated soils. The investigation also suggests that phytorestoration of metal-contaminated sites might be facilitated using the appropriate plant with the aid of tolerant AM fungi.     

Removal of benzo (a) pyrene from soil using an endogeic earthworm Pontoscolex corethrurus ()

The endogeic earthworm Pontoscolex corethrurus (Müller, 1857) was the most abundant species (75%) in soil contaminated with hydrocarbons, mostly benzo(a)pyrene (BaP), in the state of Tabasco (Mexico). The earthworm P. corethrurus was tested for its capacity to remove 100 mg BaP kg⁻¹ from an Anthrosol soil (sterilized or not) and amended with legume Mucuna pruriens (L.) DC. var. utilis (Wall. ex Wight) Baker ex Burck (3%) or the grass Brachiaria humidicola (L.) DC (3%) (recently renamed as Urochloa humidicola (Rendle) Morrone & Zuloaga) in an aerobic incubation experiment. P. corethrurus removed 26.6 mg BaP kg⁻¹ from the sterilized soil and application of B. humidicola as feed increased this to 35.7 mg BaP kg⁻¹ and M. pruriens to 34.2 mg BaP kg⁻¹ after 112 days. The autochthonous microorganisms removed 9.1 mg BaP kg⁻¹ from the unsterilized soil and application of B. humidicola increased this to 18.0 mg BaP kg⁻¹ and M. pruriens to 11.2 mg BaP kg⁻¹. Adding P. corethrurus to the unsterilized soil accelerated the removal of BaP and 36.1 mg kg⁻¹ was dissipated from soil. It was found that the autochthonous microorganisms removed BaP from soil, but addition of P. corethrurus increased the dissipation 4-fold. The endogeic earthworm P. corethrurus can thus be used to remediate hydrocarbon-contaminated soils in tropical regions.     

Interactions between plant species and mycorrhizal colonization on the bacterial community composition in the rhizosphere

This study assessed the effect of mycorrhizal colonization by Glomus intraradices (Gi) and G. versiforme (Gv) on the bacterial community composition in the rhizosphere of canola, clover and two tomato genotypes (wild type (76R) and its mutant with reduced mycorrhizal colonization (rmc)). Additionally, the effect of light intensity on the rhizosphere bacterial community composition of the tomato genotypes was studied. The bacterial community composition was assessed by denaturing gradient gel electrophoresis (DGGE). In canola, which is considered to be a non-mycorrhizal species, inoculation with Gi increased the shoot dw compared to Gv and the non-mycorrhizal control plants and also induced changes in the bacterial community composition in the rhizosphere. These fungal effects were observed although less than 8% of the root length of canola was colonized. On the other hand, about 50% of the root length of clover was colonized and inoculation with Gv resulted in a higher shoot dw compared to Gi or the control plants but the rhizosphere bacterial community composition was not affected by inoculation. Plant growth, mycorrhizal colonization and bacterial community composition of the two tomato genotypes were affected by a complex interaction between tomato genotype, AM fungal species and light intensity. Low light intensity (photosynthetic photon flux 200–250 μmol m⁻² s⁻¹) increased the shoot–root ratio in both genotypes and reduced colonization in the wild type. The differences in bacterial community composition between the two genotypes were more pronounced at low than at high light intensity (550–650 μmol m⁻² s⁻¹).     

Arbuscular mycorrhizal abundance in contaminated soils around a zinc and lead deposit

In order to study the variations in spore abundance and root colonization parameters of arbuscular mycorrhizal (AM) fungi in a naturally heavy metals polluted site and their relationships with soil properties, 35 plots in the Anguran Zn and Pb mining region were selected along a transect from the mine to 4500 m away. Within each plot, a composite sample of root and rhizospheric soil from a dominant indigenous plant was collected. The soil samples were analyzed for their physico-chemical characteristics. Spores were extracted, counted and identified at genus level. The roots were examined for colonization, arbuscular abundance, mycorrhizal frequency and intensity. Along the transect, the total and available (DTPA-extractable) concentration of Zn decreased from 6472 to 45 mg kg⁻¹ and 75 to 5 mg kg⁻¹, respectively. For Pb the values varied from 5203 to 0 mg kg⁻¹ and 32 to 0 mg kg⁻¹, respectively. In parallel, root colonization rate in the dominant native plants (except Alyssum sp.) varied from 35% to 85% and the spore numbers from 80 to 1306 per 200 g dry soil along the transect. Spores of Glomus were abundantly found in all plots as dominant, while Acaulospora spores were observed only in some moderately polluted and in control plots. AM fungal propagules never disappeared completely even in soils with the highest rates of both heavy metals. Spore numbers were more affected by Zn and Pb concentrations than root colonization. The variations of AM fungi propagules were better related to available than to total concentration of both metals. Spore numbers were positively correlated with mycorrhizal colonization parameters, particularly with arbuscular abundance.     

Biochar soil amendment increased bacterial but decreased fungal gene abundance with shifts in community structure in a slightly acid rice paddy from Southwest China

Biochar’s role on greenhouse gas emission and plant growth has been well addressed. However, there have been few studies on changes in soil microbial community and activities with biochar soil amendment (BSA) in croplands. In a field experiment, biochar was amended at rates of 0, 20 and 40 t ha⁻¹ (C0, C1 and C2, respectively) in May 2010 before rice transplantation in a rice paddy from Sichuan, China. Topsoil (0–15 cm) was collected from the rice paddy while rice harvest in late October 2011. Soil physico-chemical properties and microbial biomass carbon (MBC) and nitrogen (MBN) as well as selected soil enzyme activities were determined. Based on 16S rRNA and 18S rRNA gene, bacterial and fungal community structure and abundance were characterized using terminal-restriction fragment length polymorphism (T-RFLP) combined with clone library analysis, denaturing gradient gel electrophoresis (DGGE) and quantitative real-time PCR assay (qPCR). Contents of SOC and total N and soil pH were increased but bulk density decreased significantly. While no changes in MBC and MBN, gene copy numbers of bacterial 16S rRNA was shown significantly increased by 28% and 64% and that of fungal 18S rRNA significantly decreased by 35% and 46% under BSA at 20 and 40 t ha⁻¹ respectively over control. Moreover, there was a significant decrease by 70% in abundance of Methylophilaceae and of Hydrogenophilaceae with an increase by 45% in Anaerolineae abundance under BSA at 40 t ha⁻¹ over control. Whereas, using sequencing DGGE bands of fungal 18S rRNA gene, some bands affiliated with Ascomycota and Glomeromycota were shown inhibited by BSA at rate of 40 t ha⁻¹. Significant increases in activities of dehydrogenase, alkaline phosphatases while decreased β-glucosidase were also observed under BSA. The results here indicated a shift toward a bacterial dominated microbial community in the rice paddy with BSA.     

Accounting for variability in soil microbial communities of temperate upland grassland ecosystems

This study aimed to determine the factors which regulate soil microbial community organisation and function in temperate upland grassland ecosystems. Soil microbial biomass (C_mic), activity (respiration and potential carbon utilisation) and community structure (phospholipid fatty acid (PLFA) analysis, culturing and community level physiological profiles (CLPP) (Biolog^®)) were measured across a gradient of three upland grassland types; Festuca–Agrostis–Galium grassland (unimproved grassland, National Vegetation Classification (NVC) — U4a); Festuca–Agrostis–Galium grassland, Holcus–Trifolium sub-community (semi-improved grassland, NVC — U4b); Lolium–Cynosurus grassland (improved grassland, NVC — MG6) at three sites in different biogeographic areas of the UK over a period of 1 year. Variation in C_mic was mainly due to grassland type and site (accounting for 55% variance, v, in the data). C_mic was significantly (P<0.001) high in the unimproved grassland at Torridon (237.4 g C m⁻² cf. 81.2 g C m⁻² in semi- and 63.8 g C m⁻² in improved grasslands) and Sourhope (114.6 g C m⁻² cf. in 44.8 g C m⁻² semi- and 68.3 g C m⁻² in improved grasslands) and semi-improved grassland at Abergwyngregyn (76.0 g C m⁻² cf. 41.7 g C m⁻² in un- and 58.3 g C m⁻² in improved grasslands). C_mic showed little temporal variation (v=3.7%). Soil microbial activity, measured as basal respiration was also mainly affected by grassland type and site (n=32%). In contrast to C_mic, respiration was significantly (P<0.001) high in the improved grassland at Sourhope (263.4 l h⁻¹m⁻² cf. 79.6 l h⁻¹m⁻² in semi- and 203.9 l h⁻¹m⁻² unimproved grasslands) and Abergwyngregyn (198.8 l h⁻¹m⁻² cf. 173.7 l h⁻¹m⁻² in semi- and 88.2 l h⁻¹m⁻² unimproved grasslands). Microbial activity, measured as potential carbon utilisation, agreed with the respiration measurements and was significantly (P<0.001) high in the improved grassland at all three sites (A₅₉₀ 0.14 cf. 0.09 in semi- and 0.07 in unimproved grassland). However, date of sampling also had a significant (P<0.001) impact on C utilisation potential (v=24.7%) with samples from April 1997 having highest activity at all three sites. Variation in microbial community structure was due, predominantly, to grassland type (average v=23.6% for bacterial and fungal numbers and PLFA) and date of sampling (average v=39.7% for bacterial and fungal numbers and PLFA). Numbers of culturable bacteria and bacterial PLFA were significantly (P<0.001) high in the improved grassland at all three sites. Fungal populations were significantly (P<0.01) high in the unimproved grassland at Sourhope and Abergwyngregyn. The results demonstrate a shift in soil microbial community structure from one favouring fungi to one favouring bacteria as grassland improvement increased. Numbers of bacteria and fungi were also significantly (P<0.001) higher in August than any other sampling date. Canonical variate analysis (CVA) of the carbon utilisation data significantly (P<0.05) differentiated microbial communities from the three grassland types, mainly due to greater utilisation of sugars and citric acid in the improved grasslands compared to greater utilisation of carboxylic acids, phenolics and neutral amino acids in the unimproved grasslands, possibly reflecting substrate availability in these grasslands. Differences in C_mic, activity and community structure between grassland types were robust over time. In addition, broad scale measures of microbial growth and activity (C_mic and respiration) showed little temporal variation compared to measures of soil microbial community structure, which varied quantitatively with respect to environmental variables (temperature, moisture) and plant productivity, hence substrate supply.     

Effects of dazomet on soil organisms and recolonisation of fumigated soil

Fumigation is a common practice to control soil pathogens, but little is known about the impacts of fumigation on other soil biota groups. The purpose of this study was to investigate the effects of fumigation on soil biota, including microorganisms, nematodes, and microarthropods. Bacteria were the most resistant group and some survived following treatment with 2000 mg kg⁻¹ dazomet. Some soil fungi survived 100 mg kg⁻¹ dazomet, although they were mainly Trichoderma. The fungi pathogenic to ginseng were all killed at 100 mg kg⁻¹, and showed both inter- and intra-species variation with respect to dazomet susceptibility. Among the nematodes, Aphelenchus was relatively resistant. The results suggested that susceptibility of soil organisms to dazomet differs between species, and that tolerant organisms may engage in recolonisation. In microcosm experiments, the microbial biomass and community were assessed using phospholipid fatty acid (PLFA) analysis while recolonisation of soil organisms was controlled by mesh size. The bacterial PLFA levels were changed little after fumigation, whereas the fungal PLFA levels gradually increased after fumigation. Principal analysis of the PLFA levels and the ratio of gram-negative to gram-positive bacteria showed that fumigation altered the microbial community. The number of nematodes did not recover even at 12 weeks after fumigation. The increased Collembolan numbers suggest that fumigated soil could be recolonised by specific organisms that have adapted to the conditions. In field experiments, we tested the ability of organic materials to enhance the recolonisation of fumigated soil by soil organisms. Bean powder and rice bran increased the microbial PLFA levels and nematode numbers at 6 weeks and 12 weeks after treatment, and the abundance of nematodes continued to increase 42 weeks after fumigation. The abundance of microarthropods was only slightly affected by the presence of the organic materials. We suggest that treating fumigated soils with organic materials is an effective technique to promote soil organism numbers. In addition, Trichoderma was observed to be relatively resistant to fumigation, and therefore, we propose that the fumigation effect can be improved by using a combination of resistant Trichoderma and dazomet.     

Effects of ecological restoration on microbial activity,microbial functional diversity,and soil organic matter in mixed-oak forests of southern Ohio,USA

As a result of many decades of fire suppression and atmospheric deposition the deciduous forests of eastern North America have changed significantly in stem density, basal area, tree size-frequency distribution, and community structure. Consequently, soil organic matter quality and quantity, nutrient availability, and microbial activity have likely been altered. This study evaluated the effects of four alternative forest ecosystem restoration strategies on soil microbial activity, microbial functional diversity, soil organic C, and soil N status in two mixed-oak (Quercus spp.) forests in southern Ohio, USA. The soils of these forests were sampled during the fourth growing season after application of (1) prescribed fire, (2) thinning of the understory and midstory to pre-settlement characteristics, (3) the combination of fire and thinning, and (4) an untreated control. Prescribed fire, with or without thinning, resulted in increased bacterial but not fungal activity when assessed using Biolog^®. In contrast, assays of acid phosphatase and phenol oxidase activity indicated greater microbial activity in the thinning treatment than in the other three treatments. Functional diversity of both bacteria and fungi was affected by restoration treatment, with the bacterial and fungal assemblages present in the thin + burn sites and the fungal assemblage present in the thinned sites differing significantly from those of the control and burned sites. Treatments did not result in significant differences in soil organic C content among experimental sites; however, the soil C:N ratio was significantly greater in thinned sites than in sites given the other three treatments. Similarly, there were no significant differences in dissolve inorganic N, dissolved organic N, or microbial biomass N among treatments. Bacterial and fungal functional diversity was altered significantly. Based on Biolog^® utilization treatments the bacterial assemblage in the thin-only treatment appeared to be relatively N-limited and the fungal assemblage relatively C-limited, whereas in the thin + burn treatment this was reversed. Although effects of restoration treatments on soil organic matter and overall microbial activity may not persist through the fourth post-treatment year, effects on microbial functional diversity are persistent.     

Stimulatory effect of phosphate-solubilizing fungal strains (Aspergillus awamori and Penicillium citrinum) on the yield of chickpea (Cicer arietinum L. cv. GPF2)

The effect of six phosphate-solubilizing fungi (PSF, two strains of Aspergillus awamori, and four of Penicillium citrinum) isolated from rhizosphere of various crops, was observed on the growth and seed production of chickpea plants (Cicer arietinum L. cv. GPF2) in pot experiments. The phosphate (P) solubilizing activity of PSF in liquid varied from 38 to 760 μg ml^?1 for tricalcium phosphate (TCP) and 28–248 μg ml^?1 for mussoorie rock phosphate (MRP). All PSF isolates were biocompatible and produced growth-promoting hormone, Indole acetic acid (IAA), varying in concentration from 2.5 to 9.8 μg ml^?1. Of the various pot experiments carried out in green house, maximum stimulatory effect on chickpea plants growth was observed by inoculation of two A. awamori strains. This treatment resulted in 7–12% increase in shoot height, nearly three-fold increase in seed number and two-fold increase in seeds weight as compared to the control (un-inoculated) plants. Inoculation of four strains of P. citrinum exhibited lesser stimulatory effect. It showed 7% increase in shoot height, two-fold increase in seed number and 87% increase in seeds weight as compared to the control plants. However, a consortium of all the six fungal isolates showed no stimulatory effect on chickpea plants growth.     

Can P NMR spectroscopy be used to indicate the origins of soil organic phosphates?

The relationship between organic P status of 4 soils, 20 microorganisms isolated from these soils (2 bacteria and 3 fungi for each soil) and 13 dominant plant species of typical natural ecosystems of these soils was evaluated. The soils used were represented by two pairs with different ratios of monoester and diester P, and of DNA and other diester P. A Dystric Podzoluvisol and an alpine Umbric Leptosol were characterized by a relatively high proportion of diester P including much DNA P, while a Calcic Chernozem and subalpine Umbric Leptosol had lower proportion of diesters containing relatively less DNA P. The proportions of P compounds in bacteria and plants were very similar on average, based on the monoester to diester P ratio and on the proportions of different diesters in alkaline extract, whereas fungi contained considerably higher proportions of monoesters and polyphosphates, and a higher proportion of phospholipids in the diester fraction. The results showed that the P_org composition of NaOH extracts from different soils was more similar to the composition of extracts from different groups of microorganisms. There was no clear correspondence between soil and microbial diester P proportion and composition. A high proportion of polyphosphate P including pyrophosphate P in soil extracts indicates a significant contribution of fungal P compounds in the soil while the monoester to diester P ratio, and DNA to non-DNA P ratio should be used with caution to interpret the origins of soil P_org. The relative contributions of microorganisms and plants to monoester and diester P in soils is only partially understood.     

Assessing the tolerance to heavy metals of arbuscular mycorrhizal fungi isolated from sewage sludge-contaminated soils

Different fungal ecotypes were isolated from soils which had received long-term applications of metal-contaminated sewage sludge with the aim of studying the degree of tolerance and adaptation to heavy metals of arbuscular mycorrhizal (AM) fungi. The development and structural aspects of AM colonization produced by the different fungal isolates were studied using two host plants, Allium porrum and Sorghum bicolor, which were grown in either contaminated or non-contaminated soils. Four different AM fungi were successfully isolated from the experimental field plots: (i) Glomus claroideum, isolated from plots receiving only inorganic fertilizer; (ii) another apparently similar ecotype of Glomus claroideum, but isolated from plots with 300 m³ ha⁻¹ year⁻¹ of contaminated sludge added, (iii) an unidentified Glomus sp., present only in the less contaminated plots (100 m³ ha⁻¹ year⁻¹ of unamended sludge) and (iv) Glomus mosseae, isolated from plots receiving 100 or 300 m³ ha⁻¹ year⁻¹ of amended or unamended sludge (intermediate rates of contamination). There were consistent differences in behaviour among the four AM fungi tested with regard to the colonization levels they produced in non-contaminated and contaminated soils. Both total and arbuscular colonization were affected by heavy metal contamination. The main conclusions of this study are that Glomus sp. and G. mosseae isolates are strongly inhibited by heavy metals, which acted mainly by interfering with the growth of the external mycelium, and also by limiting the production of arbuscules. Our results suggest that G. claroideum isolates, particularly the ecotype which was isolated from the plots receiving the highest dose of metal-contaminated sludge, shows a potential adaptation to increased metal concentration in soil.     

Importance of DNA quality in comparative soil microbial community structure analyses

This study assessed how different in-situ lysis soil DNA extraction methods influence the DNA yield, quality and hence the results obtained by bacterial and fungal automated ribosomal intergenic spacer analysis (ARISA). Of the methods tested in three soils, a modified hexadecyltrimethylammonium bromide-dithiotreitol (CTAB-DTT)-based method produced ?3 times more DNA of higher quality than the other methods (260/230 nm ratios=1.64–1.82 and 260/280 nm ratios=1.82–1.89 and extracts were less inhibitory of PCR). DNA extracted by this method also yielded more reproducible ARISA ribotypes (89?119 for bacteria and 48?88 for fungi; P<0.05) than DNA extracted by other methods, and consequently produced more reliable estimates of bacterial and fungal diversity in all three test soils. The significant correlations observed between the numbers of reproducible ribotypes and DNA extract 260/230 nm ratios (r=0.88 and 0.72 for bacteria and fungi, respectively; P<0.001) reaffirmed the strong influence of DNA quality on the reliability of microbial diversity indices determined based on PCR-based DNA fingerprinting technique. Results of discriminant function analysis (DFA) and multivariate analysis of variances (MANOVA) performed on ARISA profiles (number and relative abundance of ribotype) revealed that the variability associated with DNA extraction methods did not exceed the biological variability among soil types; this supports the conclusion that high-quality DNA underpins DNA fingerprinting techniques.     

Land use influences soil fungal community composition across central Victoria,south-eastern Australia

Current theory expects that fungi, on the one hand, are spatially ubiquitous but, on the other, are more susceptible than bacteria to disturbance such as land use change due to dispersal limitations. This study examined the relative importance of location and land use effects in determining soil fungal community composition in south-eastern Australia. We use terminal restriction fragment length polymorphism (T-RFLP; primer pair ITS1-F–ITS4) and multivariate statistical methods (NMDS ordinations, ANOSIM tests) to compare relative similarities of soil fungal communities from nine sites encompassing three locations (ca 50–200 km apart) and four land uses (native eucalypt forest, Pinus radiata plantation, Eucalyptus globulus plantation, and unimproved pasture). Location effects were generally weak (e.g. ANOSIM test statistic R ≤ 0.49) and were, in part, attributed to minor differences in soil texture. By contrast, we found clear and consistent evidence of land use effects on soil fungal community composition (R ≤ 0.95). That is, soils from sites of the same land use grouped together in NMDS ordinations of fungal composition despite geographic separations of up to ca 175 km (native eucalypt forests) and 215 km (P. radiata plantations). In addition, different land uses from the same location were clearly separate in NMDS ordinations, despite, in one case, being just 180 m apart and having similar land use histories (i.e. P. radiata versus E. globulus plantation both established on pasture in the previous decade). Given negligible management of all sites beyond the early establishment phase, we attribute much of the land use effects to changes in dominant plant species based on consistent evidence elsewhere of strong specificity in pine and eucalypt mycorrhizal associations. In addition, weak to moderate correlations between soil fungal community composition and soil chemical variables (e.g. Spearman rank correlation coefficients for individual variables of 0.08–0.32), indicated a minor contributing role of vegetation-mediated changes in litter and soil chemistry. Our data provide evidence of considerable plasticity in soil fungal community composition over time spans as short as 6–11 years. This suggests that – at least within geographic zones characterised by more-or-less contiguous forest cover – soil fungal community composition depends most on availability of suitable habitat because dispersal propagules are readily available for colonisation after land use change.     

Detection and quantification of the nematophagous fungus Hirsutella minnesotensis in soil with real-time PCR

A real-time PCR assay was developed to quantify in soil the fungus Hirsutella minnesotensis, an important parasite of secondary-stage juvenile (J2) of the soybean cyst nematode. A primer pair 5′-GGGAGGCCCGGTGGA-3′ and 5′-TGATCCGAGGTCAACTTCTGAA-3′ and a TaqMan probe 5′-CGTCCGCCGTAAAACGCCCAAC-3′ were designed based on the sequence of the ITS region of the rRNA gene. The primers were highly species-specific. The PCR reaction system was very sensitive and able to detect as few as 4 conidia g^?1 soil. Regression analysis showed similar slopes and efficiency on DNA from pure culture (y = ?3.587x + 41.017, R² = 0.9971, E = 0.9055) and from Log conidia g^?1 soil (y = ?3.855x + 37.669, R² = 0.9139, E = 0.8172), indicating that the real-time PCR protocol can reliably quantify H. minnesotensis in the soil. The real-time PCR assay was applied to 20 soil samples from soybean fields, and compared with a parasitism assay. The real-time PCR assay detected H. minnesotensis in six of the soils, whereas the parasitism assay detected H. minnesotensis in the same six soils and three additional soils. The real-time PCR assay was weakly correlated (R² = 0.49) with the percentage of parasitized J2 in the six soils, indicating that different types of soil may interfere the efficiency of the real-time PCR assay, possibly due to the effect of soil types on efficacy of DNA extraction. The parasitism assay appeared to be more sensitive than real-time PCR in detecting presence of H. minnesotensis, but real-time PCR was much faster and less costly and provided a direct assessment of fungal biomass. Using the two assays in combination can obtain more complete information about the fungus in soil than either assay alone. Hirsutella parasitism was widespread and detected in 13 of the 20 field soils, indicating that these fungi may contribute to suppressiveness of soybean cyst nematode in nature and likely have high biological control potential for the nematode.     

Effects of arbuscular mycorrhizal inoculation and phosphorus supply on the growth and nutrient uptake of Kandelia obovata (Sheue,Liu & Yong) seedlings in autoclaved soil

This study evaluated the interactive effect of arbuscular mycorrhizal fungi (AMF) inoculation and exogenous phosphorus supply on soil phosphotases, plant growth, and nutrient uptake of Kandelia obovata (Sheue, Liu & Yong). We aimed to explore the ecophysiological function of AMF in mangrove wetland ecosystems, and to clarify the possible survival mechanism of mangrove species against nutrient deficiency. K. obovata seedlings with or without AMF inoculation (mixed mangrove AMF), were cultivated for six months in autoclaved sediment medium which was supplemented with KH₂PO₄ (0, 15, 30, 60, 120 mg kg−¹). Then the plant growth, nitrogen and phosphorus content, root vitality, AMF colonization and soil phosphatase activity were analyzed. The inoculated AMF successfully infected K. obovata roots, developed intercellular hyphae, arbuscular (Arum-type), and vesicle structures. Arbuscular mycorrhizal fungi colonization ranged from 9.04 to 24.48%, with the highest value observed under 30 and 60 mg kg⁻¹ P treatments. Soil P supply, in the form of KH₂PO₄, significantly promoted the height and biomass of K. obovata, enhanced root vitality and P uptake, while partially inhibiting soil acid (ACP) and alkaline phosphotase (ALP) activities. Without enhancing plant height, the biomass, root vitality and P uptake were further increased when inoculated with AMF, and the reduction on ACP and ALP activities were alleviated. Phosphorus supply resulted in the decrease of leaf N–P ratio in K. obovata, and AMF inoculation strengthened the reduction, thus alleviating P limitation in plant growth. Arbuscular mycorrhizal fungi inoculation and adequate P supply (30 mg kg⁻¹ KH₂PO₄) enhanced root vitality, maintained soil ACP and ALP activities, increased plant N and P uptake, and resulted in greater biomass of K. obovata. Mutualistic symbiosis with AMF could explain the survival strategies of mangrove plants under a stressed environment (waterlogging and nutrient limitation) from a new perspective.     

Soil amendments and watering influence the incidence of endophytic fungi in Amaranthus hybridus in South Africa

A study was conducted to determine the influence of soil amendments and irrigation on the incidence of endophytic fungi in Amaranthus hybridus. Five- and 6-month-old, asymptomatic tissues from A. hybridus were sampled from cultivated plots at Potchefstroom, South Africa in 1997 and 1998, respectively. Soil treatments consisted of the addition of commercial fertilizer or manure to irrigated soils, and wood ash to nonirrigated soils; control plots were neither amended nor irrigated. Ten leaves, 10 petioles, and 10 roots from each of five plants per soil treatment were surface disinfested and small sections from each were placed on corn-meal agar (8000 isolation attempts). After 5 days, the resulting fungal colonies were counted. Significant differences in recovery of fungi occurred among the soil treatments (P < 0.01) and among plant parts (P < 0.01). The highest recovery occurred from the commercial fertilizer and watered treatment (least stressed) for leaves and petioles in both years. Higher fungal recovery also occurred in the wettest year from leaves and petioles for all soil treatments. In contrast, roots yielded higher fungal recovery in the driest year for all soil treatments. These results show that soil attributes can influence frequency of endophytic fungi in both above- and below-ground tissues.