Microbial activity and nutrient dynamics in earthworm casts (Lumbricidae)

Summary Microbial respiration, microbial biomass and nutrient requirements of the microflora (C, N, P) were studied in the food substrate (soil taken from the upper 3 cm of the mineral soil of a beech wood on limestone), the burrow walls and the casts of the earthworm Aporrectodea caliginosa (Savigny). The passage of the soil through the gut caused an increase in soil microbial respiration of about 90% over a 4-week period. Microbial biomass was increased only in freshly deposited casts and decreased in aging faeces to a level about 10% lower than in soil. Microbial respiration of the burrow walls was only increased over a shorter period (about 2 weeks). The microflora of the soil and the burrow walls was limited by P, whereas in earthworm casts, microbial growth was limited by the amount of available C. In aging faeces the P requirement of the microflora increased and approached that of the soil. Immobilization of phosphate in earthworm casts is probably caused by mainly abiotic processes. C mineralization by soil microflora fertilized with glucose and P was limited by N, except in freshly deposited casts. Ammonium, not nitrate, was responsible for this process. N dynamics in earthworm casts are discussed.     

The role of the earthworm Lumbricus terrestris in the transport of bacterial inocula through soil

Two laboratory experiments were used to investigate the effect of the earthworm Lumbricus terrestris on transport of genetically marked Pseudomonas fluorescens inocula through soil microcosms. The microcosms comprised cylindrical cores of repacked soil with or without earthworms. Late log-phase cells of P. fluorescens, chromosomally marked with lux genes encoding bioluminescence, were applied to the surface of soil cores as inoculated filter paper discs. In one experiment, 5 and 10 days after inoculation, cores were destructively harvested to determine concentrations of marked pseudomonads with depth relative to the initial inoculum applied. Transport of the bacteria occurred only in the presence of earthworms. In a second experiment cores were subjected to simulated rainfall events 18 h after inoculation with lux-marked bacteria at 3-day intervals over a 24-day period. Resulting leachates were analysed for the appearance of the marked bacteria, and after 28 days cores were destructively harvested. Although some marked cells (less than 0.1% of the inoculum applied) were leached through soil in percolating water, particularly in the presence of earthworms, the most important effect of earthworms on cell transport was through burial of inoculated litter rather than an increase in bypass flow due to earthworm channels.     

Plant community impacts on the structure of earthworm communities depend on season and change with time

Declining plant diversity potentially threatens essential ecosystem functions driven by the decomposer community, such as litter decomposition and nutrient cycling. Currently, there is no consensus on the interrelationships between plant diversity and decomposer performance and previous studies highlighted the urgent need for long-term experiments.In the Jena Experiment we investigated the long-term impacts of plant community characteristics on the structure of earthworm communities representing key decomposers in temperate grassland. We repeatedly sampled plots varying in plant species richness (1-16 species), plant functional group richness (1-4 groups), and presence of certain plant functional groups (grasses and legumes) three, four, and six years after establishment of the experiment in spring and autumn.The results show that earthworm performance is essentially driven by the presence of certain plant functional groups via a variety of mechanisms. Plant productivity (root biomass) explained most of the detrimental grass impacts (decrease in earthworm performance), while beneficial legume effects likely were linked to high quality inputs of plant residues (increase in earthworm performance). These impacts depended on the functional group of earthworms with the strongest effects on surface feeding anecic earthworms and minor effects on soil feeding endogeic species. Remarkably, effects of plant community characteristics on the composition and age structure of earthworm communities varied between seasons. Moreover, plant diversity effects reported by a former study decreased and detrimental effects of grasses increased with time.The results indicate that plant community characteristics, such as declining diversity, indeed affect the structure of earthworm communities; however, loss of key plant functional groups is likely to be more important than plant species number per se. However, in frequently disturbed ecosystems plant species richness might be important for the recovery and resilience of belowground functions. Moreover, the results accentuate the importance of long-term repeated measurements to fully appreciate the impacts of plant community composition and diversity on ecosystem properties. Single point observations may be misleading and potentially mask the complexity of above-belowground interrelationships.     

Fate and dispersal of recombinant bacteria in a soil microcosm containing the earthworm Lumbricus terrestris

Summary A model ecosystem was designed containing Lumbricus terrestris in soil. The microcosm is suitable for assessing the dispersal and survival of genetically engineered microorganisms. After the inoculation of genetically engineered microorganisms their number in the worm excretions decreased continuously to reach the level of the detection limit within 50 days.     

Microbial biomass and activity in pine litter in the presence of Tomocerus minor (Insecta,Collembola)

Summary Laboratory microcosms were used to study microbial populations and biomasses developing in fragmented litter of Pinus nigra Arnold var. nigra (A. et G.). Direct observations (fungal standing crop and fluorescein-stainable mycelia), litter enzyme analyses (cellulase and dehydrogenase), and measurements by physiological methods (microbial CO₂ production and total microbial, fungal, and bacterial viable biomasses) were made at 3-week intervals for 15 weeks. Most variables showed great changes during this period, which were ascribed to a rise in litter moisture content during the initial phase of the experiment, and to substrate depletion towards its final phase. The addition of the collembolan Tomocerus minor (Lubbock) for 1 week enhanced cellulase activities by 4%. When the animals were introduced after 6 weeks, the fungal standing crop was enhanced, and the percentage of fluorescein-stainable mycelia was reduced. Dehydrogenase activity was increased by grazing when the microbial population had been established for 9 weeks or longer. Eucaryotic and procaryotic substrate-induced respiration were positively correlated, which was explained by partial segregation of resources for the two groups. Litter cellulase and dehydrogenase activity showed correlations by other techniques, indicating their suitability as parameters for microbial activity in general, and for the collembolan grazing impact on microbial activity in particular.     

Novel method for the study of the population dynamics of a genetically modified microorganism in the gut of the earthworm Lumbricus terrestris

Summary A laboratory microcosm study was used to investigate the survival and population dynamics of genetically modified microorganisms (GMM) in the gut of Lumbricus terrestris. Three methods of axenic earthworm production were investigated. An antibiotic mixture of streptomycin and cycloheximide was introduced either passively, mixed with sterile soil or cellulose, or actively, by teflon catheter. Worms treated by all methods lost weight but this was least for the catheter method which was also the only method to produce axenic earthworms. Axenic earthworms were used to determine the effect of competition with indigenous gut bacteria on ingested GMM. The GMM used was Pseudomonas fluorescens, strain 10586/FAC510, with chromosomally inserted Lux genes for bioluminescence, and chromosomal resistance to rifampicin. The bacteria were grown up to the mid-exponential phase before inoculation into earthworms. Bacteria in faecal material were enumerated by dilution plate counting using selective agar. The GMM were re-isolated from the casts of both antibiotic-treated and untreated earthworms. Lower concentrations of GMM and higher concentrations of indigenous bacteria in the casts of untreated compared to antibiotic-treated earthworms suggested that competition is a fundamental control on population dynamics of the introduced bacterial inocula ingested by earthworms.The catheter method, developed in this study, is proposed as a technique to contribute to the risk assessment of environmental release of GMM.     

The nature of eurytopy in polyploid earthworm species in relation to their use in biological soil amelioration

Summary Many of the most widely distributed species of Lumbricidae are polyploid. So far polyploid forms have been found to be distributed more widely than diploid ones. Taking Eisenia nordenskioldi as an example, it was shown that polyploids are able to occupy new ecological niches within the area of the diploid form. Ecological differences observed in representatives of different chromosomal races provide polyploids with the chance to coexist in one biocurrent, occupying different ecological niches.Dedicated to the late Prof. Dr. M.S. Ghilarov     

Gut load and food-retention time in the earthworms Lumbricus festivus and L. castaneus: A field study

Summary Lumbricus festivus and L. castaneus consume dung. In the field, below cow pats, their gut loads were about 0.15 and 0.14 g dry weight g^-1 ash-free dry weight of worm, respectively, but in free soil the loads were higher, about 0.21 and 0.19 g g^-1 ash-free dry weight of worm. The gut contents of dung were lighter than the total ingested material, at about 0.10 and 0.07 g dry weight g^-1 ash-free dry weight of worm, respectively. Field experiments showed that the retention time of dung ranged from >9 to 15 h for L. festivus, and from >3 to 6 h for L. castaneus. The experiments also indicated that L. festivus exploited 20- and 36-day-old dung in different ways, since the gut load was lower in those worms consuming 20-day-old dung than in those consuming 36- to 40-day-old dung. On the basis of these results the calculated consumption rate for L. festivus is 0.08 g dung day^-1 g live weight of worm^-1, and for L. castaneus 0.15 g dung day^-1, with retention times assumed to be at maximum, 15 h, for L. festivus and 6 h for L. castaneus. These calculations indicate that our field population of worms (75 g m^-2) consumes 10–15 t dung ha^-1 180 days^-1, corresponding to the amount of dung produced by 2–3 dairy cows.     

Leaf-litter consumption and assimilation by juveniles of Lumbricus terrestris L. (Oligochaeta,Lumbricidae) under different environmental conditions

Summary The rate of consumption of dandelion leaves per g earthworm dry weight per week was described by non-linear functions of soil temperature, soil water potential, and food availability. The optimum temperature and soil water potential for food consumption are about 22°C and-7 kPa, respectively. Zero consumption occurred at about-40 kPa. Food consumption increased with greater food availability, but only up to 1.05 (dandelion) and 1.23 (grass) g dry weight per g earthworm dry weight per week at 15°C. A general consumption function to account for all three environmental factors is given. The assimilation rate per g earthworm dry weight per week was defined as the sum of the growth rate and the maintenance rate. Maintenance was calculated according to respiratory measurements reported previously, whereas growth was measured. High temperatures and limiting environmental conditions, such as a low food availability and a low soil water potential, led to an increase in the assimilation efficiency of Lumbricus terrestris. At-7 kPa and 15°C, L. terrestris assimilated 55 and 43% of the ingested dandelion if 0.25 and 1.0 g dry weight of food was available per g earthworm dry weight per week, respectively. it is concluded that L. terrestris has a strong, direct effect on the decomposition of highly palatable plant materials.     

Evaluation of suppression of rhizomania disease by earthworm (Lumbricus terrestris L.) and its effects on soil microbial activity in different sugar beet cultivars

The ability of earthworm Lumbricus terrestris L. to suppress the multiplication of Beet necrotic yellow vein virus (BNYVV) transmitted by Polymyxa betae and its effects on soil microbial activity were investigated under controlled conditions. BNYVV-infested and a combination of earthworm with BNYVV-infested soil were compared to non-infested soils for their effects on plant and root weights and virus content of two different sugar beet cultivars by using partially resistant (cv. Leila) and susceptible (cv. Arosa) cultivars to the rhizomania disease. Soil testing with sugar beet baiting plants followed by enzyme-linked immunosorbent assay (ELISA) was used to diagnose virus. The results of the statistical analyses showed that total fresh plant and root weights were negatively correlated with BNYVV infection. Addition of L. terrestris significantly enhanced plant and root weights. The earthworm-added soils had higher microbial activity such as basal soil respiration and dehydrogenase activity. The presence of earthworms in the soil did not statistically suppress BNYVV infection (p < 0.05). Sugar beet production may be enhanced by using resistant cultivars with adding L. terrestris into soil where rhizomania is present.     

Abundance of the earthworm Lumbricus terrestris in relation to subsurface drainage pattern on a sandy clay field

Subsurface drainage induces systematic spatial variability in soil properties which may be reflected in the abundance and distribution of soil organisms. We compared the population density of the deep burrowing earthworm Lumbricus terrestris L. above and between tile subdrains in 41 sample pairs on an eight hectare grass field. Above the drains the median number of  individuals was twice as high and their total fresh mass five times as high as between the drains (4.5 vs. 2.1 individuals m^-2  and 9.6 vs. 1.9 g m^-2, respectively). The mean difference (above drain – between drains) was 2.5 individuals m^-2  (95% CI = 1.0 to 4.0) and 6.6 grams m^-2 (95% CI = 3.6 to 9.6). The relatively larger difference in fresh mass was due to a high proportion of adult individuals above drains. One likely explanation for the pattern of abundance is that the lowered water table level near the drains provides an environment beneficial for the population growth of L. terrestris. Due to the role of L. terrestris burrows as flow paths of percolating water the observations may have implications on subdrain function.     

不同种植年限宁夏枸杞根际微生物多样性变化

为了解长期人工种植枸杞根际土壤微生物的种群结构变化特征,利用Illumina Mi Seq测序平台分别对种植5 a、10 a和15 a宁夏枸杞根际土壤微生物基因组总DNA中16S和18S r DNA基因的部分区域进行测序,经UPARSE pipeline和RDP classifier软件进行聚类分析和物种注释。结果表明,长期种植宁夏枸杞不会改变其根际土壤p H,但会导致根际土壤全磷、有效磷、全盐含量和电导率升高。测序结果表明,不同种植年限枸杞根际土壤细菌群落α多样性无显著变化,但真菌群落多样性在种植10 a枸杞中较种植5 a时显著降低(p0.05),表明根际细菌群落多样性受种植年限影响较小。从门的分类水平看,酸杆菌门、放线菌门、拟杆菌门、厚壁菌门、绿弯菌门、泉古菌门、蓝菌门、芽单胞菌门、变形菌门以及真菌子囊菌门、担子菌门、接合菌门的比例在不同种植年限的枸杞根际土壤中显著改变(p0.05)。属水平的分析也表明,共有27个细菌属和16个真菌属的比例发生改变(p0.05),这些结果表明枸杞根际土壤微生物群落组成受种植年限的影响更大。相关性分析结果表明,种植年限、土壤全磷及有效磷含量是影响枸杞根际微生物群落结构的主要因子。     

Spring dynamics of soil carbon, nitrogen, and microbial activity in earthworm middens in a no-till cornfield

Earthworm activity may be an important cause of spatial and temporal heterogeneity of soil properties in agroecosystems. Structures known as “earthworm middens,” formed at the soil surface by the feeding and casting activities of some earthworms, may contribute significantly to this heterogeneity. We compared the temporal dynamics of carbon (C), nitrogen (N), and microbial acitivity in Lumbricus terrestris middens and in surrounding non-midden (bulk) soil during the spring, when seasonal earthworm activity was high. We sampled soil from middens and bulk soil in a no-till cornfield on four dates during May and June 1995. Soil water content and the weight of coarse organic litter (>2mm) were consistently higher in middens than in bulk soil. Total C and N concentrations, C:N ratios, and microbial activity also were greatest in midden soil. Concentrations of ammonium-nitrogen and dissolved organic N were greater in middens than in bulk soil on most dates, suggesting accelerated decomposition and mineralization in middens. However, concentrations of nitrate were usually lower in middens, indicating reduced nitrification or increased leaching and denitrification losses from middens, relative to bulk soil. Fungal activity, as well as total microbial activity, was consistently greater in middens. The contribution of fungae to overall microbial activity differed significantly between middens and bulk soil only on one date when both soils were very dry; the contribution of fungae to microbial activity was lower in the middens on this date. We conclude that the midden-forming activity of L. terrestris can be a major determinant of spatial heterogeneity in some agricultural soils, and that this can potentially affect overall rates of soil processes such as organic matter decomposition, N mineralization, denitrification, and leaching. Received: 4 April 1997     

Changes in microbial biomass and microbial activity of pig slurry after the transit through the gut of the earthworm Eudrilus eugeniae (Kinberg, 1867)

Here we studied the effects of gut transit through the earthworm Eudrilus eugeniae, on the physicochemical, biochemical, and microbial characteristics of pig slurry, by analyzing fresh casts. The reduction in the dissolved organic C contents in casts we recorded suggests that during digestion, earthworms assimilated labile organic C preferentially, which is a limiting growth factor for them. Furthermore, both microbial biomass and activity in pig slurry were significantly decreased by earthworm gut transit. It appears that E. eugeniae is able to digest microorganisms, although the addition of glucose to the food increased respiration, indicating that growth of microorganisms in casts could be limited by depletion of labile C. Despite reduced microbial biomass and activity, the metabolic diversity of microbial communities was greater in casts than in original pig slurry. Community level physiological profiles obtained from Biolog Ecoplate data revealed that, after earthworm gut transit, the microbial communities in casts and pig slurry were clearly differentiated by their physiological profiles. The results indicate that first stage in vermicomposting of pig slurry by E. eugeniae, i.e., casting, produced changes that will influence the dynamics of the organic matter degradation by reducing forms of N and C available to microorganisms, hence restricting their growth and multiplication. Nevertheless, the reduced microflora of casts was characterized by an increased catabolic potential that might lead to thorough degradation of pig slurry.     

Potential contribution of Lumbricus terrestris L. to carbon dioxide, methane and nitrous oxide fluxes from a forest soil

 Potential effects of earthworms (Lumbricus terrestris L.) inoculated into soil on fluxes of CO₂, CH₄ and N₂O were investigated for an untreated and a limed soil under beech in open topsoil columns under field conditions for 120 days. Gas fluxes from L. terrestris, beech litter and mineral soil from soil columns were measured separately in jars at 17  °C. The inoculation with L. terrestris and the application of lime had no effect on cumulative CO₂ emissions from soil. During the first 3–4 weeks earthworms significantly (P<0.05) increased CO₂ emissions by 16% to 28%. In contrast, significantly lower (P<0.05) CO₂ emission rates were measured after 11 weeks. The data suggest that earthworm activity was high during the first weeks due to the creation of burrows and incorporation of beech litter into the mineral soil. Low cumulative CH₄ oxidation rates were found in all soil columns as a result of CH₄ production and oxidation processes. L. terrestris with fresh feces and the beech litter produced CH₄ during the laboratory incubation, whereas the mineral soil oxidised atmospheric CH₄. Inoculation with L. terrestris led to a significant reduction (P<0.02) in the CH₄ oxidation rate of soil, i.e. 53% reduction. Liming had no effect on cumulative CH₄ oxidation rates of soil columns and on CH₄ fluxes during the laboratory incubation. L. terrestris significantly increased (P<0.001) cumulative N₂O emissions of unlimed soil columns by 57%. The separate incubation of L. terrestris with fresh feces resulted in rather high N₂O emissions, but the rate strongly decreased from 54 to 2 μg N kg^–1 (dry weight) h^–1 during the 100 h of incubation. Liming had a marked effect on N₂O formation and significantly (P<0.001) reduced cumulative N₂O emissions by 34%. Although the interaction of liming and L. terrestris was not significant, N₂O emissions of limed soil columns with L. terrestris were 8% lower than those of the control. Received: 2 September 1999     

Microbial biomass and activity in soil and litter underPinus sylvestris, Picea abies andBetula pendula at originally similar field afforestation sites

Microbial biomass C and N, and activities related to C and N cycles, were compared in needle and leaf litter, and in the uppermost 10 cm of soil under the litter layer in Scots pine (Pinus sylvestris L.), Norway spruce (Picea abies L.) and silver birch (Betula pendula L.) stands, planted on originally similar field afforestation sites 23–24 years ago. The ground vegetation was differentiated under different tree species, consisting of grasses and herbs under birch and pine, and mosses or no vegetation with a thick layer of needles under spruce. The C:N ratio of the soils was 13–21 and the soil pH_{CaCl 2} 3.8–5.2. Both showed little variation under different tree species. Microbial biomass C and N, C mineralization, net ammonification, reduction) did not differ significantly in soil under different tree species either. Birch leaf litter had a higher pH_{CaCl 2} (5.9) than spruce and pine needle litter (pH 5.0 and 4.8, respectively). The C:N ratio of spruce needles was 30, and was considerably higher in pine needles (69) and birch leaves (54). Birch leaves tended to have the highest microbial biomass C and C mineralization. Spruce needles appeared to have the highest microbial biomass N and net formation of mineral N, whereas formation of mineral N in pine needles and birch leaves was negligible. Microbial biomass C and N were of the same order of magnitude in the soil and litter samples but C mineralization was tenfold higher in the litter samples.     

Microfungal communities in soil,litter and casts of Lumbricus terrestris L. (Lumbricidae): a laboratory experiment

The anecic earthworm Lumbricus terrestris L. was kept in laboratory microcosms containing beech forest soil without litter, with beech leaf litter or with lime leaf litter. The structure of microfungal communities in soil, litter and fresh and aged (100 days) earthworm faeces was analysed using the washing and plating technique. The passage of mineral soil through the gut of L. terrestris affected the structure of the fungal community only little. In contrast, in the litter treatments the structure of the fungal community in fresh earthworm casts significantly differed from that in soil and litter. The majority of soil and litter inhabiting fungi survived passage through the gut of L. terrestris and the fungal community in casts consisted of a mixture of soil and litter inhabiting fungi. However, the frequency of Cladosporium spp., Alternaria spp., Absidia spp., and other taxa was strongly reduced in fresh casts. The degree of colonization of litter particles (number of isolates per number of plated particles) also decreased, but some fungi (mainly Trichoderma spp.) benefited from gut passage and flourished in fresh casts. During ageing of cast material the dominance structure of the fungal community changed. Both the degree of colonization of organic particles and the species diversity increased and approached that in soil. However, the structure of the fungal community in casts remained cast specific even after 100 days of incubation. It is concluded that the feeding and burrowing activity of L. terrestris accelerates the colonization of litter by the edaphic mycoflora but also extends the range of occurrence of litter-associated fungi into mineral soil layers.     

Microbial biomass and respiratory activity in soil aggregates of different sizes from three beechwood sites on a basalt hill

We studied the effects of aggregates of different sizes on the soil microbial biomass. The distribution of aggregate size classes (<2, 2–4, 4–10, >10 mm) in the upper mineral soil horizon (A_h layer) was very different in three sites (upper, intermediate, lower) in a beechwood (Fagus sylvatica) on a basalt hill (Germany). Aggregates of different sizes (<2, 2–4, 4–10 mm) contained different amounts of C and N but the C:N ratios were similar. C and N contents were generally higher in smaller aggregates. The maximum initial respiratory response by microorganisms in intact aggregates and in aggregates passed through a 1-mm sieve declined with the aggregate size, but the difference was more pronounced in intact aggregates. Disruption of aggregates generally increased this response, particularly in 4- to 10-mm aggregates in the lower site. Basal respiration differed strongly among sites, but was similar in each of the aggregate size classes. Aggregate size did not significantly affect the specific respiration (g O₂ g^–1 microbial C h^–1) nor the microbial: organic C ratio, but these parameters differed among sites. Microbial growth was increased strongly by passing the soil through a 1-mm sieve in each of the aggregate materials. The growth of microorganisms in disrupted aggregates was similar, and the effect of aggregate disruption depended on the growth of microorganisms in intact aggregates.     

Structure and activity of soil-inhabiting methanotrophic communities in northern forest of Japan

In a Japanese forest, CH₄ uptake rate and methanotrophic community structure in the soil were investigated at four sites of different vegetation. At two of these sites, undergrowth was dominated by Sasa senanensis, and that of another was dominated by Sasa kurilensis. At the rest site, undergrowth had been removed artificially. The tree-layer composition differed between the two sites with S. senanensis, but tree layer of the other two sites were dominated by the same species. At the site lacking undergrowth, observed CH₄ uptake rate was twice as high as that at the other sites. Under laboratory conditions, soil sample from the site lacking undergrowth exhibited CH₄ consumption rate higher than that of the adjacent site with the same dominating tree species. The community structures of methanotrophs were investigated with denaturing gradient gel electrophoresis (DGGE) of the gene encoding particulate methane monooxygenase (pmoA). The banding patterns observed were different depending on the type of undergrowth vegetation. The sequences of the DGGE bands were closely related to each other and belonged to the “upland soil cluster alpha” (USCα). These results imply possible close relationship between the undergrowth vegetation and methanotrophic communities in forest soils.     

Microbial biomass and activity indices after organic substrate addition to a selenium-contaminated soil

High concentrations of Se in soil might have negative effects on microorganisms. For this reason, the effect of organic substrate addition (glucose + maize straw) on Se volatilisation in relation to changes in microbial biomass and activity indices was investigated using an artificially Se-contaminated soil. Microbial biomass N was reduced on average by more than 50% after substrate addition, but adenylate energy charge (AEC) and metabolic quotient qCO₂ were both increased. The Se content decreased by nearly 30% only with the addition of the organic substrate at 25°C. No significant Se loss occurred without substrate at 25°C or with substrate at 5°C. In the two treatments with substrate addition, the substrate-derived CO₂ evolution was about 30% lower with Se addition than without. In contrast, Se had no effect on any of the other soil microbial indices analysed, i.e. microbial biomass C, microbial biomass N, adenosine triphosphate (ATP), AEC, ATP-to-microbial biomass C, and qCO₂.