Fungal abundance and diversity in earthworm casts and in uningested soil

Earthworm casts and adjacent uningested soil from 30 different locations were compared to determine the abundance and diversity of fungal species. The casts contained larger fungal populations (g^-1 dry soil weight) and numbers of fungal species than the soil. Variations in these parameters between casts and soil were statistically significant (P=0.05). Fungal populations and the number of fungal species in casts and soil also varied significantly (P=0.05) between samples from different locations. A total of 27 fungal species were recorded from the casts and soil. Indices of dominance (0.084 casts; 0.14 soil) and general diversity (2.53 casts; 2.02 soil) demonstrated that the casts displayed more diverse fungal flora than the soil. The diversity of fungal species increased in earthworm casts after passing through the earthworm gut.     

Microbial populations,enzyme activities and nitrogen-phosphorus-potassium enrichment in earthworm casts and in the surrounding soil of a pineapple plantation

Summary Total populations of bacteria and fungi, dehydrogenase activity (as a measure of total potential microbial activity), and urease and phosphatase activities were determined in earthworm casts and surrounding laterite soils planted to pineapple. The casts contained higher microbial populations and enzyme activities than the soil. Except for fungal populations, statistically significant (P = 0.05) increases were found in all other parameters. Microbial populations and enzyme activities showed similar temporal trends with higher values in spring and summer and lower values in winter. The earthworm casts contained higher amounts of N, P, K and organic C than the soil (P = 0.05). Selective feeding by earthworms on organically rich substrates, which break down during passage through the gut, is likely to be responsible for the higher microbial populations and greater enzyme activity in the casts.     

Fate of prions in soil: Longevity and migration of recPrP in soil columns

Research into transmissible spongiform encephalopathy (TSE) diseases has become a high priority worldwide in recent years yet remarkably little is known about the behaviour of TSE infectivity in the environment. The resilience and stability of prion proteins could lead to soils becoming a potential reservoir of TSE infectivity as a result of contamination from activities such as infected carcass burial or the dispersion of effluents from slaughter houses, or by contamination of pastures by infected animals, (e.g. scrapie in sheep). Knowledge of the fate of prion proteins in soils, and associated physico-chemical conditions which favour migration, can be used to help prevent re-infection of animals through grazing, to protect watercourses and develop good management practices. In two consecutive experiments of 9 and 6 months, the migration of recombinant ovine PrP (recPrP) in soil columns was followed under contrasting levels of microbial activity (normal versus reduced), under varying regimes of soil water content and redox potential, and in two different soil types (loamy sand and clay loam). At each analysis time (1, 3, 6 or 9 months), in both soil types, full-length recPrP was detected in the original contaminated layer, indicating the resilience and stability of recPrP under varied soil conditions, even in the presence of active soil microbial populations. Evidence of protein migration was found in every soil column at the earliest analysis time (1 or 3 months), but was restricted to a maximum distance of 1 cm, indicative of limited initial mobility in soils followed by strong adsorption over the following days to weeks. The survival of recPrP in the soil over a period of at least 9 months was demonstrated. In this study, recPrP was used as an indicator for potential TSE infectivity, although infectivity tests should be carried out before conclusions can be drawn regarding the infection risk posed by prions in soil. However, it has been demonstrated that soil is likely to act as a significant barrier to the dispersion of contaminated material at storage or burial sites.     

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.     

Fate of prions in soil: Interaction of a recombinant ovine prion protein with synthetic humic-like mineral complexes

Prion proteins are regarded as the main agents of transmissible spongiform encephalopathies. Understanding their fate in soil may be crucial to elucidate the dissemination of the prion in the environment, associated with a possible transmission of infectivity. Studies were performed with simplified model systems, derived by the birnessite-mediated oxidative polymerization of catechol, which simulate processes naturally occurring in soil. A benign full-length recombinant purified ovine protein (PrP) (residues 23-234), as well as a truncated-form ovPrP (tPrP) (103-234) were utilized. Catechol and prion protein interacted under experimental conditions that reproduced the interaction of PrP with soluble organic matter or with insoluble organic matter during or after formation of catechol polymers. PrP stability in all buffers and chemicals was preliminarily monitored by circular dicroism (CD) measurements. The disappearance of protein molecules from the solution, the decrease of UV-Visible absorbance of supernatants, and the FT-IR spectra and the elemental analyses of solid-phase residues indicated that both PrP and tPrP were involved in catechol polymerization by birnessite. Furthermore, a clear flocculation of soluble catechol-protein polymeric products in solid aggregates was observed when PrP was added to the supernatants. Different kinds of extracting agents were not able to desorb/extract PrP as well as tPrP from the formed solid aggregates, thereby highlighting the high stability of protein-organic and -organo-mineral complexes.     

Destabilization of soil during the production of earthworm (Lumbricidae) and artificial casts

The mechanisms by which soil is destabilized in the digestive tract of endogéic earthworms were investigated with artificial casts, which were moulded with a syringe from slurries of a silty loam with or without gypsum and organic matter treatments, and compared to casts produced by Aporrectodea rosea (Lumbricidae). Both types of casts generally had the same levels of mechanical dispersion, observations of slaking, and particle size distribution when the casts were fresh, aged-moist for 30 days, or air-dried. Fresh casts were significantly more dispersive than the uningested soil despite the addition of gypsum or organic matter to the soil. However, the dispersion from aged-moist or air-dried casts was not substantially greater than that of uningested soil. Air-drying was more effective than moist-ageing in increasing the stability of casts and uningested soil. The concentration of soluble carbohydrate was greater in artificial casts produced from soil treated with sheep dung or xanthan gum, and in earthworm casts produced from soil treated with xanthan, than in the uningested soil of the same treatments. An increase in the concentration of soluble carbohydrate was related to an increase in dispersion. An attempt was made to simulate the addition of mucus to soil in the digestive tract of earthworms, by the addition of sucrose or xanthan gum to the slurry during the production of artificial casts. The addition of xanthan, but not sucrose, to the slurry increased mechanical dispersion relative to that of the uningested soil in the fresh treatment. Although the production of artificial casts destabilized soil to the same degree as earthworm casts, the artificial casts did not simulate all chemical, biochemical, and microbiological aspects of digestion. Received: 24 November 1995     

Effects of earthworm casts and compost on soil microbial activity and plant nutrient availability

Vermicomposting differs from conventional composting because the organic material is processed by the digestive systems of worms. The egested casts can be used to improve the fertility and physical characteristics of soil and potting media. In this study, the effects of earthworm casts (EW), conventional compost (CP) and NPK inorganic fertilizer (FT) amendments on N mineralization rates, microbial respiration, and microbial biomass were investigated in a laboratory incubation study. A bioassay with wheat (Triticum aestivium L.) was also conducted to assess the amendment effects on plant growth and nutrient uptake and to validate the nutrient release results from the incubation study. Both microbial respiration and biomass were significantly greater in the CP treatment compared to EW treatment for the initial 35 days of incubation followed by similar respiration rates and biomass to the end of the study at 70 days of incubation. Soil NO₃⁻ increased rapidly in the EW and CP treatments in the initial 30 days of incubation, attaining 290 and 400 mg N kg⁻¹ soil, respectively. Nitrate in the EW treatment then declined to 120 mg N kg⁻¹ soil by day 70, while nitrate in the CP treatment remained high. While ammonium levels decreased in the CP treatment as nitrate level increased with increasing incubation time, a low level of ammonium was maintained in the EW treatment throughout the incubation. The wheat bioassay study included two additional cast treatments (EW-N and EW2) to have treatments with higher levels of N input. Plants grown with CP or FT treatment had a lower shoot biomass and higher shoot N content than in EW-N and EW-2 treatments, and also showed symptoms of salinity stress. Ionic strength and other salinity indicators in the earthworm cast treatments were much lower than in the CP treatment, indicating a lower risk of salinity stress in casts than in compost. All cast and compost amendments significantly increased wheat P and K uptake compared to either the non-amended control or the mineral fertilizer treatment. The results show that casts are an efficient source of plant nutrients and that they are less likely to produce salinity stress in container as compared to compost and synthetic fertilizers.     

Degradation dynamics of surface earthworm casts in grasslands of the eastern plains of Colombia

 Earthworms are generally considered to fit the definition of ecosystem engineers. The casts they produce are recognised to have a great importance in the regulation of soil processes. Lifetimes and degradation rates of these structures remain poorly known. In this study, the dynamics of disappearance and the changes in the physical properties of the surface casts of the anecic earthworm Martiodrilus carimaguensis were assessed in a native savanna and an intensive pasture. In both systems, casts were composed of superposed layers deposited by earthworms over a period of at least a few days. The half-life of casts ranged between 2 months and 11 months in the pastures (trampled and protected, respectively), and 5 months in the savanna. Some dry casts remained at the soil surface for more than 1 year after having been excreted. The disappearance of casts was mostly attributed to rain-drop impact and the effect of animal trampling. The bulk density of fresh casts was higher (+17%, P<0.05) or equivalent (–1%, P>0.05) to that of the surrounding soil, in the savanna and the pasture, respectively. Fresh cast aggregates were of larger size than bulk soil aggregates (about +70% in both systems, P<0.05). Bulk density and the size of cast aggregates decreased with cast ageing (from –29% to –24% for bulk density, and from –68% to –80% for size, in the pasture and the savanna, respectively). Macroinvertebrates were observed digging into casts, and were assumed to be partly responsible for the physical degradation of casts with time. Received: 12 November 1999     

Digestion and residue stabilization of bacterial and fungal cells,protein, peptidoglycan,and chitin by the geophagous earthworm Metaphire guillelmi

Microbial biomass is an important source of soil organic matter, which plays crucial roles in the maintenance of soil fertility and food security. However, the mineralization and transformation of microbial biomass by the dominant soil macrofauna earthworms are still unclear. We performed feeding trials with the geophagous earthworm Metaphire guillelmi using ¹⁴C-labelled bacteria (Escherichia coli and Bacillus megaterium) cells, fungal (Penicillium chrysogenum) cells, protein, peptidoglycan, and chitin. The mineralization rate of the microbial cells and cell components was significantly 1.2–4.0-fold higher in soil with the presence of M. guillelmi for seven days than in earthworm-free soil and 1–11-fold higher than in fresh earthworm cast material. When the earthworms were removed from the soil, the mineralization of the residual carbon of the microbial biomass was significantly lower than that in the earthworm-free soil, indicating that M. guillelmi affects the mineralization of the biomass in soil in two aspects: first stimulation and then reduction, which were attributed to the passage of the microbial biomass through the earthworm gut, and that the microorganisms in the cast could play only minor roles in the stimulated mineralization and residual stabilization of microbial biomass. Large amounts (8–29%) of radiolabel of the tested microbial biomass were assimilated in the earthworm tissue. Accumulation of fungal cells (11%) and cell wall component chitin (29%) in the tissue was significantly higher than that of bacterial cells (8%) and cell wall component peptidoglycan (15%). Feeding trails with ¹⁴C-lablled microbial cells and cell components provided direct evidence that microbial biomass is a food source for geophagous earthworm and fungal biomass is likely a more important food source for earthworms than bacterial biomass. Findings of this study have important implications for the roles of geophagous earthworms in the fate of microbial biomass in soil.     

The interactions between soil type and earthworm species determine the properties of earthworm casts

Earthworms are recognized to increase soil porosity, reorganize soil structure, and stimulate soil microflora and nutrient mineralization. The properties of earthworm casts should depend both on earthworm species or ecological group and on soil properties. Interactions between earthworm species and soil types have been suggested, but only poorly demonstrated. In order to better understand those interactions, two hypotheses led our study: (1) Soil type has a greater influence on cast properties than earthworm; (2) Earthworms from different species influence cast properties differently; (3) The intensity and direction of the impact of each earthworm species on cast properties vary with soil properties. Fifteen physical and chemical variables (N–NH₄⁺, N–NO₃⁻, total organic C and N, C/N ratio, CaCO₃, pH, P, K⁺, Mg²⁺, Mn²⁺, Na⁺, CEC, moisture, wettability) were measured in casts of three earthworm species (Lumbricus terrestris, Allolobophora chlorotica and Aporrectodea rosea) produced in three temperate soils. Univariate and multivariate analyses showed that earthworm species and soil types significantly impacted cast properties. pH, N_t, K and Mg contents were interactively altered by both factors. Multivariate analysis showed that a difference of soil type had a major impact on casts properties (62%) compared to the impact of a difference of earthworm species (10%). Cast properties were most impacted by L. terrestris, then by A. chlorotica and last by A. rosea. The response ratio (ratio of the properties of the casts to the properties of the bulk soil) was used to quantify the effect of earthworm species compared to the control soil. It showed a higher response of variables in casts in nutrient-rich soils, especially in casts of L. terrestris. The interactions between earthworm species and soil types on cast properties were discussed with regards to earthworm ecology, properties of the soil, and earthworm modifications of cast microflora.     

Inability of Near Infrared Reflectance Spectroscopy (NIRS) to identify belowground earthworm casts in no-tillage soil

Several studies have emphasised the ability of Near Infrared Reflectance Spectroscopy (NIRS) to identify surface earthworm casts in the field. However, less is known about casts deposited within the soil, which usually represent the majority found in the field. This study tested the ability of NIRS to identify belowground casts in agricultural systems. Casts and surrounding soils were sampled at depths of 20–30 cm in a loamy soil under no tillage for 12 years. To distinguish different types of cast, sizes and orientations relative to the horizontal plane were measured. NIRS analyses and analyses of carbon and nitrogen content were also performed to compare casts to surrounding soils. Casts were classified into 4 size classes, with no preferential orientation. Cast carbon and nitrogen content were not influenced by their size and did not differ from surrounding soils. PCAs performed on the NIRS data did not allow casts to be differentiated from surrounding soils, regardless of size class. However, soil aggregates were clearly differentiated probably due to their spatial distribution in the soil. Although this study did not identify specific NIRS signatures for casts, it shows the utility of this method to investigate the origin of the soil consumed by earthworms. In our case, NIRS analyses suggest that the high bulk density of the soil (1.42 g cm⁻³) forced ingestion by endogeic earthworms, simply to move around, without preferential selection for organic matter. Consequently, their casts were deposited a few mm from where they had ingested soil with similar organic matter quality.     

Variables controlling denitrification from earthworm casts and soil in permanent pastures

Summary Denitrification (using the acetylene block method) was determined in earthworm casts and soils from permanent, drained or undrained pasture plots fertilized with 0 or 200 kg N ha^-1 year^-1 as ammonium nitrate. Rates of N₂O production from soil cores were about three times higher from the fertilized than from the unfertilized plots while drainage had a relatively small effect. Denitrification rates from casts were 3–5 times higher than those from soil irrespective of the drainage treatment. Casts generally had higher NO _inf3 ^sup- , NH _inf4 ^sup+ , and moisture contents, and higher microbial respiration rates than soil. Rates of N₂O production were determined primarily by NO _inf3 ^sup- supply, secondarily by moisture; available C did not appear to limit denitrification in these pastures. Estimates of the potential contribution of casts to denitrification ranges from 10.1% of 29.3 kg ha^-1 year^-1 from the unfertilized, drained plot to 22% of 82.5 kg ha^-1 year^-1 from the fertilized undrained plot.     

Potential for higher rates of denitrification in earthworm casts than in the surrounding soil

Summary Earthworms (Lumbricus terrestris L.) were cultured in the laboratory and fed on lucerne (Medicago sativa L.). Denitrification rates in the surface casts and the surrounding soil were quantified using C₂H₂-inhibition of nitrous oxide reductase. The investigation also included determination of the N₂O-formation by nitrification as well as CO₂-formation as a measure of respiration. The denitrification rates of wet earthworm casts were found to be significantly higher than those occurring in wet samples from the soil. The low N₂O-formation observed seemed to be due to denitrification. Respiration was higher in casts, indicating higher oxygen demand which resulted in more anaerobic conditions. The energy supply was probably better in casts compared with the surrounding soil.     

Influence of cropping intensity on the production and properties of earthworm casts in a leucaena alley cropping system

The effect of cropping intensity on surface cast production by earthworms was studied more than 4 years in an alley cropping system. The plots were cultivated to maize–cassava intercrop every year (100% cropping intensity), after 2 years of fallow (33% cropping intensity) and after 3 years of fallow (25% cropping intensity). Cast production was assessed by the continuous sampling technique proposed by Hauser and Asawalam (Z Pflanzenernahr Bodenkd 161:23–30, 1998). Results showed that cropping intensity enhanced cast production in the first year, did not affect it in the second and third years and decreased cast production in the fourth year. Amount of casts deposited within the alleys was highest under the hedgerow and decreased towards the middle of the alley. The concentrations of organic matter and plant nutrients were significantly higher in worm casts than in soil, irrespective of the cropping intensity and position of sample collection within the interrow space. Continuous cropping up to 6 years significantly reduced the concentrations of organic carbon, total nitrogen, available phosphorus, and exchangeable calcium, potassium and magnesium in the top 0–5 cm of the soil, irrespective of the distance from the hedgerow. Regardless of the intensity of cropping, there is a strong linear association between the quantity of casts and the amount of organic carbon, and other nutrients in casts.     

Heavy metal accumulation by two earthworm species and its relationship to total and DTPA-extractable metals in soils

The concentrations of Zn, Cd, Pb and Cu in earthworm tissues were compared with the total and DTPA-extractable contents of these heavy metals in contaminated soils. Samples were taken from a pasture polluted by waste from a metallurgic industry over 70 y ago. Three individuals of Aporrectodea caliginosa and Lumbricus rubellus and soil samples were collected at six points along a gradient of increasing pollution. Total metal contents of earthworms, soil, and metals extracted by DTPA from the soil were measured. Total heavy metal contents of the soils ranged from 165.7 to 1231.7 mg Zn kg⁻¹, 2.7 to 5.2 mg Cd kg⁻¹, 45.8 to 465.5 mg Pb kg⁻¹ and 30.0 to 107.5 mg Cu kg⁻¹. Their correlations with metals extracted by DTPA were highly significant. Contents of the metals in earthworm tissues were higher in A. caliginosa than in L. rubellus, with values ranging from 556 to 3381 mg Zn kg⁻¹, 11.6 to 102.9 mg Cd kg⁻¹, 1.9 to 182.8 mg Pb kg⁻¹ and 17.9 to 35.9 mg Cu kg⁻¹ in A. caliginosa, and from 667.9 to 2645 mg Zn kg⁻¹, 7.7 to 26.3 mg Cd kg⁻¹, 0.5 to 37.9 mg Pb kg⁻¹ and 16.0 to 37.6 mg Cu kg⁻¹ in L. rubellus, respectively. Correlations between body loads in earthworms with either total or DTPA-extractable contents of soil metals were significant, except for Cd in L. rubellus and Cu in A. caliginosa. Considering its simple analytical procedure, DTPA-extractable fraction may be preferable to total metal content as a predictor of bio-concentrations of heavy metals in earthworms. Biota-to-Soil Accumulation Factor (BSAF) of these four metals are Cd>Zn>Cu>Pb, with range of mean values between: Cd (6.18-17.02), Zn (1.95-7.91), Cu (0.27-0.89) and Pb (0.08-0.38) in A. caliginosa, and Cd (3.64-6.34), Zn (1.5-6.35), Cu (0.29-0.87) and Pb (0.04-0.13) in L. rubellus. The BSAF of Ca, Fe and Mn are Ca>Mn>Fe, with mean values of: Ca (0.46-1.31), Mn (0.041-0.111), Fe (0.017-0.07) in A. caliginosa and Ca (0.98-2.13), Mn (0.14-0.23), Fe (0.019-0.048) in L. rubellus, respectively. Results of principal component analysis showed that the two earthworm species differ in the pattern of metal bioaccumulation which is related to their ecological roles in contaminated soils.     

Nitrogen mineralization and reorganization in casts of the geophagous tropical earthworm Pontoscolex corethrurus (Glossoscolecidae)

Summary Mineral N concentrations ranged from 133.1 to 167.8 g g^-1 dry soil in fresh casts of the endogeic earthworm Pontoscolex corethrurus fed on an Amazonian Ultisol; this was approximately five times the concentration in non-ingested soil. Most of this N was in the form of NH _inf4 ^sup+ . N also accumulated in microbial biomass, which increased from a control value of 10.5–11.3 to 67.5–74.1 g g^-1 in fresh casts. During a 16-day incubation, part of the NH _inf4 ^sup+ -N was nitrified and/or transferred to the microbial biomass. Total labile N (i.e., mineral+biomas N) decreased sharply at first (ca. 50% in the first 12 h), and then more slowly. The exact fate of this N (microbial metabolites, denitrification, or volatilization) is not known. After 16 days, the overall N content of the casts was still 28% higher than that of the control soil. Incubation of the soil before ingestion by the earthworms significantly increased the production of NH _inf4 ^sup+ in casts. We calculate that in a humid tropical pasture, 50–100 kg mineral N may be produced annually in earthworm casts. Part of this N may be conserved in the compact structure of the cast where the cast is not in close contact with plant roots.     

Habitat quality, conspecific density, and habitat pre-use affect the dispersal behaviour of two earthworm species, Aporrectodea icterica and Dendrobaena veneta, in a mesocosm experiment

Dispersal capacity is a life-history trait that may have profound consequences for earthworm populations: it influences population dynamics, species persistence and distribution and community structure. It also determines the level of gene flow between populations and affects processes such as local adaptation, speciation and the evolution of life-history traits. It may play a great role in soil functioning by determining the spatial distribution of ecosystem engineers such as earthworms. Dispersal is an evolutionary outcome of the behaviour in response to the ecological constraints of the species. Hence different dispersal behaviour is expected from the different ecological types of earthworms. Nevertheless the dispersal behaviour of earthworms has been little documented.In this work we test a series of basic mechanisms that are fundamental and complementary to understand earthworms dispersal behaviour. We focus on the dispersal triggered by environmental conditions, a fundamental process usually termed “conditional dependent dispersal”. We show experimentally in mesocosms that in one week: 1) earthworm dispersal can be triggered by low habitat quality, either through soil quality or the presence/absence of litter. 2) Earthworms can be subject to positive density dependent dispersal, that is the rate of dispersal increases when density increases; and 3) earthworm dispersal can be reduced by the pre-use of the soil by conspecific individuals that are no longer present.Our results suggest that earthworms may be more mobile than expected from previous estimations, and that they present high capacities of habitat selection. In the light of our findings we elaborate a behavioural scenario of earthworm foraging, and propose several priority working directions.     

Short-term responses of two contrasting species of earthworms in an agricultural soil amended with coal fly-ash

With the renewed interest in the use of coal fly-ash for amendment of agricultural soils in Australia, we assessed how earthworms, as indicators of soil health, responded to this ameliorant. We assessed survival, weight, burrowing and elemental concentrations for earthworms of a native unnamed Megascolecid species and of exotic Aporrectodea trapezoides in intact soil cores treated with an alkaline fly-ash at rates equivalent to 0, 5 and 25 t/ha over 6 weeks. Fly-ash did not affect survival, growth, number of burrows created or phosphorus solubilisation. Transfer of the earthworms to the new environment having vastly different pH from where they were collected, and possibly overcrowding, caused mortality in the soil cores for all treatments. A. trapezoides that had smaller individuals suffered mortality of 12% compared with 23% for the larger earthworms of Megascolecids. Earthworms of Megascolecids each increased their weight by 0.24g (25% of their original weight) while those of A. trapezoides lost 0.18g each (21% of their original weight). The difference in growth between the two earthworms was associated with grazing habit and probably with the large difference in the pH between source soil and that of the core soil. Megascolecids appeared to minimize grazing on ash-tainted soil and so ingested less Zn, which was more abundant in the fly-ash than in the soil, compared with A. trapezoides that had elevated concentration of this metal. Extractable P in the soil was increased with both species of earthworms, more so with the exotic species that solubilized 11% more P than the native Megascolecids. The benign influence of fly-ash on survival and growth of worms was associated with the pH of soil remaining unchanged during the six weeks of incubation.     

Volume density of earthworm burrows in compacted cores of soil as estimated by direct and indirect methods

After earthworms of the species Aporrectodea caliginosa and A. rosea had burrowed in compacted cores of soil for 68 days the cores were sectioned horizontally. The upper surface of each sectioned layer of soil was photographed before it was dissected and the dimensions of all burrows within the layer measured. Volume densities calculated from the direct measurement of burrows were compared with the values calculated by stereology from data obtained from two image analysis methods, computerised image analysis and point counting with a systematic lattice. The assumption implicit in all stereology calculations was satisfied for this experiment in that the burrows of both species showed no preferred orientation in the compacted soil. Computerised image analysis could not measure the density of all burrows in the photographs because of the lack of contrast between cast-filled burrows and the soil and the complex shapes of the burrows. Although the volume densities of A. caliginosa burrows calculated from point counts were correlated with the values calculated from direct measurement, point counting over-estimated volume densities by two to three times. In the experiment, A. rosea produced too few airfilled burrows for the lattice to detect. The relative ratios of air-filled to cast-filled burrows calculated from the point counts suggested that approximately two-thirds and eight-ninths of the burrows of A. caliginosa and A. rosea, respectively, were filled with casts.