Assessing daily egestion rates in earthworms: using fungal spores as a natural soil marker to estimate gut transit time

Earthworms have an important role in ‘bioturbation’—the mixing of soil due to biological processes. Quantification of earthworm bioturbation relies on estimating earthworm egestion rates which in turn depend on two parameters: the gut content of the worms and the gut transit time (GTT). Gut content can be determined relatively easily, but determining GTT is problematic. The present study aimed at estimating daily soil egestion rates of Aporrectodea caliginosa and Lumbricus terrestris, refining the most common approach for estimating GTT by using fungal spores as natural markers in ingested soil. This approach avoids the use of artificial markers that may adversely affect the earthworms. Gut transit time was estimated by tracking the passage of marked soil through the gut by the appearance of the spores in the egested faeces. Gut transit time was estimated to be 9.6?±?0.3 h for A. caliginosa and 11.6?±?0.5 h for L. terrestris. Gut content averaged 465?±?40(± standard error (SE))?mg dw g^?1 dw worm for A. caliginosa and 265?±?80 mg dw g^?1 dw worm for L. terrestris. From these values, daily egestion rates of 1.16 and 0.66 g dw faeces g^?1 dw worm d^?1 were calculated for A. caliginosa and L. terrestris, respectively. Both values compare well to literature values for each species. The presented method for GTT estimation is inexpensive, rapid and easy to evaluate, with spores being a good alternative to existing markers.     

影响丛枝菌根真菌孢子萌发的几种因素研究

对丛枝菌根真菌孢子萌发的几种影响因素进行了研究。结果表明,土壤是丛枝菌根真菌孢子萌发最适宜的培养基;寄主植物根的分泌物对孢子萌发有显著的促进作用。重金属Cd和Pb含量过高时(50mg kg)抑制真菌孢子的萌发。培养基中有效P含量较低时(KH2PO4添加量为0～80mg L),对孢子萌发影响较小,高浓度的有效P(KH2PO4添加量大于100mg L)对孢子萌发有一定的抑制作用。培养基的pH值过高(pH8.0以上)或过低(pH5.5以下)抑制孢子萌发。生长激素对孢子萌发率没有显著性影响。对于有休眠现象的丛枝菌根真菌,4℃低温处理4～6周,可打破休眠孢子的休眠,显著提高休眠孢子的萌发率。     

A framework for examining inter-regional aerial transport of fungal spores

Aerial transport of fungal spores has no doubt been responsible for occasionally spreading plant diseases over distances of 500 km or more. With currently available methods, however, there is little basis for estimating the likelihood of such occurrences. Models capable of estimating the relative probability of infection of a crop from known sizes of local or distant sources of pathogenic spores could help farmers with decisions about fungicide use, local sanitation and quarantine. A logical framework for estimating long-distance transport of viable spores is presented which encompasses: production of spores at the source region, escape of spores into the air above the diseased crops, transport and dilution of spores in the atmosphere, survival of airborne spores and deposition of spores onto a distant crop. Hypothetical examples of spore transport over 700 km are calculated to illustrate the relative magnitudes of the five parts of the spore transport process. Although the combined uncertainties in spore transport are estimated to be a factor of 10⁴–10⁶, this should be compared to the expected factor of 10¹³ dilution of spore concentration experienced by a cloud of spores after having been transported through the atmosphere for 700 km. The largest uncertainties lie in our ability to estimate daily spore production in a geographic region, survival of spores while airborne and deposition of spores during rainfall. Survival of spores and deposition of spores during rainfall are difficult to predict because of temporal and spatial variations in weather. To account for this, it will be necessary in the future to combine the physical model of spore transport described here with weather records and air parcel trajectory calculations to develop a “climatology” of danger of disease transmission between geographic regions.     

乙草胺、甲胺磷及其复合污染对黑土土壤真菌的影响

Using plate counting and ergosterol assay, single and joint effects of acetochlor and methamidophos on the dynamics of soil fungal population and total fungal biomass in the black soil zone of Northeast China were investigated. The results demonstrated that acetochlor at high concentration levels （150 and 250 mg kg^-1） had an acute and mostly chronic toxicity on both the soil fungal population and total fungal biomass, but at a low concentration （50 mg kg^-1） generally had a stimulating effect that was stronger with total fungal biomass than with the soil fungal population. Methamidophos at a high concentration level （250 mg kg^-1） alone and almost all of its combinations with various dosages of acetochlor increased the soil fungal population, whereas at most sampling dates with 250 mg methamidophos kg^-1 soil, total fungal biomass increased, but in combination with acetochlor it was decreased in the early period of incubation and then increased 28 days after incubation. Thus, through measuring the number of colony forming unit of the soil fungal population along with the total fungal biomass, a better understanding on effects of agrochemicals on soil fungi could be made.     

Aqueous biphasic system to extract arbuscular mycorrhizal spores from soils

The aqueous biphasic system (ABS) method using PEG and ammonium sulphate improved the recovery of spores from tropical soils as compared with the sucrose gradient method. The ABS was not affected by low temperatures. The optimal recovery of spores was achieved when 100% PEG and 50% ammonium sulphate concentration and PEG₈₀₀₀ instead of PEG₄₀₀₀ were used. Under these conditions an optimal interface formation which facilitates the localization and recovery of spores was obtained. Both phases of the ABS are aqueous and did not affect either the viability of spores or the level of root colonization reached by these spores. Soil composition did not affect the number of spores obtained by the ABS method either.     

Suppression of root-knot nematodes in natural and agricultural soils

The potential of the invertebrate communities in agricultural and natural soils to suppress Meloidogyne incognita on coleus (Coleus blumei) was evaluated in two greenhouse experiments. Soil from adjacent natural and agricultural habitats was collected from four locations that had very different soil types (loamy sand, muck, sand and rock). Soil of each type was placed into pots, planted with coleus seedlings, inoculated with 2000 eggs of M. incognita, and arranged in a 4 × 2 factorial design. Root-knot nematodes were suppressed in natural soil compared to agricultural soil for loamy sand, sand and rock soils, but not for muck soil. Plants grew larger and had less root galling in natural soils than in agricultural soils. Free-living nematodes (bacterivores, fungivores, omnivores and predators) were monitored but did not show population patterns consistent with the suppression of root-knot nematodes observed in natural soils. Reasons for the opposite result (higher root-knot levels in natural than in agricultural soil) on muck soil are unknown, but this soil type was very different from the other soils in its unusually high organic matter content and very low levels of omnivorous and predatory nematodes. The agricultural muck in particular was unusually low in fungivorous nematodes and extremely high in levels of sodium and macronutrients. Despite the exception on the muck soil, the relative suppression of root-knot nematodes in natural compared to agricultural soils of the other three soil types (loamy sand, sand and rock) support the idea that potential for nematode suppression may be greater in natural than in agricultural soils.     

Rubidium ‐ availability and plant uptake in natural soils

Abstract

High soil acidity (low metal ion saturation) favours the absorption of Rb from natural soils by vascular plants and fungi, whereas the absorption of K is little or not influenced. The difference in Rb uptake may be as great as one order of magnitude between soils of high and low acidity. Results of several studies are presented and discussed.     

Bacterial and fungal response to nitrogen fertilization in three coniferous forest soils

Forest soil carbon (C) pools may act as sinks for, or sources of, atmospheric carbon dioxide, while nitrogen (N) fertilization may affect the net exchange of C in forest ecosystems. Since all major C and N processes in soil are driven by soil microorganisms, we evaluated the effects of N fertilization on biomass and bacterial and fungal activity in soils from three Norway spruce forests with different climatic and N availability conditions. N deposition and net N mineralization were higher at the sites in southern Sweden than at the site in northern Sweden. We also studied the extent to which N fertilization altered the nutrient(s) limiting bacterial growth in soil. We found that on average microbial biomass was reduced by ～40% and microbial activity by ～30% in fertilized plots. Bacterial growth rates were more negatively affected by fertilization than fungal growth rates, while fungal biomass (estimated using the phospholipid fatty acid (PLFA) 18:2ω6,9) decreased more than bacterial biomass as a consequence of fertilization. The microbial community structure (indicated by the PLFA pattern) was changed by fertilization, but not in the same way at the three sites. Soil bacteria were limited by a lack of carbon in all forests, with the carbon limitation becoming more evident in fertilized plots, especially in the forests that had previously been the most N-limited ones. This study thus showed that the effects of N fertilization differed depending on the conditions at the site prior to fertilization.     

Particle size fractionation of fungal and bacterial biomass in subalpine grassland and forest soils

Characterization of soil aggregates according to particle size fractions is a useful tool in process-oriented research into soil organic matter and biological properties. Substrate-induced respiration (SIR) inhibition was used to quantify microbial, fungal and bacterial biomass in particle size fractions of soils ranging from forest to grassland in a subalpine region of central Taiwan. In addition, ergosterol content was determined in the same samples to verify fungal biomass measured by SIR inhibition technique. Surface soil (0–10 cm) was fractionated into four particle size fractions: coarse sand (250–2000 μm), fine sand (53–250 μm), silt (2–53 μm) and clay (0.2–2 μm). The larger sized fractions (>250 μm and 53–250 μm) contained higher levels of fungal ergosterol than the smaller sized ones (2–53 μm and 0.2–2 μm). The largest particle size fraction (250–2000 μm) from all studied habitats showed the highest level of microbial biomass, with no clear trend in microbial biomass level among the other size fractions. SIR-calculated fungal biomass level and ergosterol converted fungal biomass content were positively correlated (r=0.71, p<0.05), and such correlation decreased as biomass levels were high. Ratios of fungi to bacteria ranged between 0.6 and 1.3 in fractions obtained in this study. This study indicates a high variability of microbial (fungal and bacterial) biomass level among particle size fractions in soil, and that the large-sized fractions tend to contain a high level of microbial biomass in a given ecosystem.     

Metal-tolerant bacterial populations from natural and metal-polluted soils

The toxicity of several metals towards bacterial populations from natural and metal-polluted soils could be described either partially by a single exponential equation or completely by the sum of two exponential functions. Bacterial populations from both soils contained two subgroups, one of which could tolerate metals over a greater range of concentrations than the other. Most bacteria comprising the more mctal-tolerant subgroup were Gram-negative and were multiple drug resistant. Exceptions were organisms, tentatively identified as coryncforms, isolated on nickel-supplemented medium. It is suggested that, in general, Gram-negative bacteria arc more metal-tolerant than Gram-positivc organisms and, in soils containing comparatively low levels of metal pollution, may be able to function without the need for plasmid-mediated metal-tolerance.     

Molecular identification of arbuscular mycorrhizal fungal spores associated to the rhizosphere of Retama raetam in Tunisia

ABSTRACT

Arbuscular mycorrhizal fungi (AMF) are found in the soil of most ecosystems where they form mutualistic associations that affect plants growth. We have investigated the community structure of AMF associated to Retama raetam growing in five regions of Tunisia. The total number of spores was significantly different across sites, ranging from 633 to 1062 spores per 100 g dry soil. A dominance of small spores was revealed. The large subunit region of the rDNA of AMF spores associated to the rhizosphere of R. raetam was sequenced. Sequences clustered into 13 operational taxonomic units. Phylogenetic analysis revealed that the majority of sequences were grouped within Glomeraceae and Claroideoglomeraceae families. Only two sequences were affiliated to the Scutellospora genus. These results suggest the dominance of the genus Glomus in the soil rhizosphere of R. raetam. A correlation between phylogenetic analysis, soil chemicals properties, and AMF community richness was also detected.     

Multiple species-specific controls of root-feeding nematodes in natural soils

One of the major limitations to enhance sustainability of crop production systems is the inability to control root-feeding nematodes without using chemical biocides. In soils under wild vegetation, however, root-feeding nematodes affect plant performance and plant community composition varying from substantially to insignificantly. Previous studies in natural ecosystems have already shown that mutualistic symbionts, such as arbuscular mycorrhizal fungi and endophytes, may influence plant exposure to root-feeding nematodes. In order to learn more from natural systems, we examined nematode control in the root zone of a wild coastal foredune grass by microorganisms, other nematodes and microarthropods. We cultured all eight root-feeding nematode species that occur in the root zone of marram grass (Ammophila arenaria) in coastal foredunes of the Netherlands. Then, in an indoor growth experiment we exposed each nematode species to the potential natural antagonists collected from the same dune soil. Most of the eight dominant root-feeding nematode species could be controlled to some extent by more than one group of soil organisms added. The effectiveness of control varied among nematode species, which seemed to be controlled in a species-specific way. We conclude that in a natural soil the effectiveness of control by microorganisms, other nematodes or microarthropods varies among root-feeding nematode species. Most are controlled, at least to some extent, by soil microbes. However, some root-feeding nematode species are controlled only by microarthropods. Our results strongly suggest that sustainable agriculture will benefit from using a range of biological control mechanisms when controlling root-feeding nematodes, rather than relying on single control agents. Our suggestion also implies that conserving soil biodiversity is crucial in order to enhance the reliability of biological crop protection against soil-borne pests and diseases.     

Soil ameliorants alter physicochemical properties and fungal communities in saline-sodic soils of Northeast China

To understand the mechanisms of soil ameliorants affecting microbial communities is important for saline-sodic soils reclamation. High-throughput sequencing was used to characterize the fungal community in soils amended with four types of ameliorants over an 8-year period. Besides a control without any additional ameliorant (CK), other four treatments including 1) amendment with sandy soil (SS), 2) amendment with desulfurization gypsum (DG), 3) amendment with farm manure (FM), and 4) amendment with a mixture of SS, DG, and FM (M) were analyzed. Soil pH and electrical conductivity significantly decreased with the addition of soil ameliorants, whereas the soil organic carbon (SOC), total nitrogen (TN), and SOC/TN ratio (C/N) significantly increased in the FM and M treatments compared with the CK treatment. Fungal richness increased significantly with the mixed ameliorants addition (M). Distinct fungal community structures were observed in the treatments with soil ameliorants. The fungal community composition was significantly associated with the SOC, C/N, aggregate content with a diameter > 0.25 mm and geometric mean diameter. The changes in these soil characteristics were highly correlated with the ameliorants additions, suggesting that the impacts of ameliorants on the soil fungal community occurred indirectly as a result of alterations to soil physiochemical properties.     

Nitrous oxide production in soils and the ratio of the fungal to bacterial biomass

The proportion between the fungal and bacterial biomass, the potential activity of denitrification, and the intensity of N₂O production were determined in the soils (chernozem and soddy-podzolic) of secondary biocenoses formed upon the abandoning of agricultural lands. The substitution of meadow and forest vegetation for agrocenoses has led to an increase in the percentage of the fungal biomass in the upper soil horizons. The rate of the net N₂O production after the soil moistening positively correlated with the content of nitrates. In the soddy-podzolic soil (pH 3.7–5.6), the rate of nitrous oxide production was higher than that in the chernozem (pH 6.1–6.8). The rate of N₂O production was inversely proportional to the bacterial biomass in the soils.     

Distribution of arbuscular mycorrhizal fungi spores in soils of smallholder agroforestry and monocultural coffee systems in southwestern Ethiopia

Arbuscular mycorrhizal fungi (AMF) are associated with the root system of coffee (Coffea arabica L.) plants, but their distribution in smallholder agroforestry and monocultural coffee systems is not well known. This study investigates the spatial distribution of AMF spores in a field study in southwestern Ethiopia. Soil samples from different depths (0–50 cm) were collected under the tree canopies of Acacia abyssinica, Albizia gummifera, Ficus sur, Ficus vasta and randomly selected unshaded coffee plants at different sampling points (canopy base, radius, edge and outside canopy). Significantly higher AMF spore densities were recorded at canopy bases and at 0–30 cm soil depth. Spore populations were found to belong to five genera: Acaulospora, Entrophospora, Glomus, Gigaspora and Scutellospora, with Glomus and Acaulospora dominating. Sampling points, sites and depths, shade tree species and shade tree/coffee plant age affected AMF spore density. Agroforestry practices including the use of leguminous shade trees effectively maintained AMF numbers in soils even at depth compared with unshaded coffee plants (monocultures).     

Nitrogen losses from two grassland soils with different fungal biomass

Nitrogen losses from agricultural grasslands cause eutrophication of ground- and surface water and contribute to global warming and atmospheric pollution. It is widely assumed that soils with a higher fungal biomass have lower N losses, but this relationship has never been experimentally confirmed. With the increased interest in soil-based ecosystem services and sustainable management of soils, such a relationship would be relevant for agricultural management. Here we present a first attempt to test this relationship experimentally. We used intact soil columns from two plots from a field experiment that had consistent differences in fungal biomass (68 ± 8 vs. 111 ± 9 μg C g⁻¹) as a result of different fertilizer history (80 vs. 40 kg N ha⁻¹ y⁻¹ as farm yard manure), while other soil properties were very similar. We performed two greenhouse experiments: in the main experiment the columns received either mineral fertilizer N or no N (control). We measured N leaching, N₂O emission and denitrification from the columns during 4 weeks, after which we analyzed fungal and bacterial biomass and soil N pools. In the additional ¹⁵N experiment we traced added N in leachates, soil, plants and microbial biomass. We found that in the main experiment, N₂O emission and denitrification were lower in the high fungal biomass soil, irrespective of the addition of fertilizer N. Higher ¹⁵N recovery in the high fungal biomass soil also indicated lower N losses through dentrification. In the main experiment, N leaching after fertilizer addition showed a 3-fold increase compared to the control in low fungal biomass soil (11.9 ± 1.0 and 3.9 ± 1.0 kg N ha⁻¹, respectively), but did not increase in high fungal biomass soil (6.4 ± 0.9 after N addition vs. 4.5 ± 0.8 kg N ha⁻¹ in the control). Thus, in the high fungal biomass soil more N was immobilized. However, the ¹⁵N experiment did not confirm these results; N leaching was higher in high fungal biomass soil, even though this soil showed higher immobilization of ¹⁵N into microbial biomass. However, only 3% of total ¹⁵N was found in the microbial biomass 2 weeks after the mineral fertilization. Most of the recovered ¹⁵N was found in plants (approximately 25%) and soil organic matter (approximately 15%), and these amounts did not differ between the high and the low fungal biomass soil. Our main experiment confirmed the assumption of lower N losses in a soil with higher fungal biomass. The additional ¹⁵N experiment showed that higher fungal biomass is probably not the direct cause of higher N retention, but rather the result of low nitrogen availability. Both experiments confirmed that higher fungal biomass can be considered as an indicator of higher nitrogen retention in soils.