Soil Protozoa Research Articles

Meeting on the Microbiology of Soils, Autumn 2001Estimation of protozoan diversity in soil

Different methods of estimating protozoan diversity in soil are discussed in this paper, with the major emphasis on heterotrophic flagellates. Although many species of ciliates and testate amoebae seem to be unique to the soil environment, the communities of heterotrophic flagellates and naked amoebae are probably best considered as restricted versions of their aquatic counterparts. Soil protozoa are difficult to observe directly, hence culture techniques are usually used. These techniques enable us to explore certain functional aspects, but their major drawback is that some soil protozoa cannot be cultured. Molecular methods, however, have the potential to detect such non-culturable forms.

お詫びと訂正

お詫びと訂正

Meeting on the Microbiology of Soils, Autumn 2001Aspects of soil protozoa on a grassland farm

This paper reviews the principal outcomes from analyses of the soil protozoan community of an upland grassland site in Scotland. An assessment of the protozoan community growth potential of 150 soil samples revealed that abundance increased in the order ciliates < testate amoebae < naked amoebae < flagellates and was inversely proportional to organism size within each functional group, whilst species richness appeared to be invariant with size. These results are consistent with the observed fractal structure of flooded Sourhope soil. Protozoan species are randomly distributed across the site and species which are globally abundant are also abundant at Sourhope, which is consistent with the hypothesis of ubiquitous dispersal of protozoan species. The potential contribution of protozoa to carbon cycling in soil was confirmed in soil microcosms in which the presence of protozoa increased carbon dioxide production significantly.

Meeting on the Microbiology of Soils, Autumn 2001Spatial distribution of soil protozoa in an upland grassland

A 12 × 12 m plot from a visually uniform upland pasture was sampled in a spatially referenced manner. There was no correlation between inter-sample distance and the protozoan community (colpodid and heterotrich ciliates, flagellates, naked amoebae, and total biomass). Evidence from the other parameters measured, and from other studies reported in the literature, indicates that spatial organisation in protozoan communities occurs at scales below 10 cm.

Meeting on the Microbiology of Soils, Autumn 2001

This paper reviews the principal outcomes from analyses of the soil protozoan community of an upland grassland site in Scotland. An assessment of the protozoan community growth potential of 150 soil samples revealed that abundance increased in the order ciliates < testate amoebae < naked amoebae < flagellates and was inversely proportional to organism size within each functional group, whilst species richness appeared to be invariant with size. These results are consistent with the observed fractal structure of flooded Sourhope soil. Protozoan species are randomly distributed across the site and species which are globally abundant are also abundant at Sourhope, which is consistent with the hypothesis of ubiquitous dispersal of protozoan species. The potential contribution of protozoa to carbon cycling in soil was confirmed in soil microcosms in which the presence of protozoa increased carbon dioxide production significantly.

Biodiversity of Terrestrial Protozoa Appears Homogeneous across Local and Global Spatial Scales

Free-living microbes are by far the most abundant group of organisms in the biosphere, yet estimates of global species richness remain nebulous, and there is no consensus regarding the likely geographical distribution of species. Both uncertainties are addressed by the suggestion that the vast abundance of microbes may drive their ubiquitous random dispersal; for this would also make it likely that global species richness is relatively low. Here we test the idea of ubiquitous dispersal of testate amoebae and ciliates living in soil. We analysed their abundance and species richness in 150 soil samples collected from the one-hectare grassland site at Sourhope in Scotland, and in comparable published data from 1500 soil samples collected worldwide. Following taxonomic revision and removal of synonyms, there remained a total of 186 taxa (91 testate and 95 ciliate) recorded from both Sourhope and other places in the world. A fundamental pattern of random spatial distribution of species was revealed in species that are relatively rare. This probably arises from random dispersal, for when localised population growth occurs, the distributions become aggregated, as in virtually all metazoan species. We find no evidence for geographically-restricted protozoan morphospecies at spatial scales of 4 m2, 10,000 m2, or worldwide. Species that are locally rare or abundant are similarly rare or abundant on a global scale. Approximately one third of the global diversity of soil protozoa was found at the one-hectare grassland site in Scotland, but this is a minimum figure, for recorded species richness is proportional to sampling effort, as shown here.

Impact of cultivation management in an agroecosystem on hot spot effects of earthworm middens

In comparison to the surrounding soil, hot spot effects of middens of the epianecic earthworm Lumbricus terrestris were determined under different field conditions on a loamy sand. The impact of 3 different management systems was studied: 1. unmulched; 2. straw mulching; 3. intermediate crop: Sinapis alba. Furthermore, we considered 2 different crops (bean, lupine) and their growth stages. We monitored different enzyme activities and abiotic parameters of soil at 6 sampling dates between April and August 1998 parallel to abundances of microfauna (soil protozoa) and mesofauna (Enchytraeidae, Collembola, mites). The analysis of earthworm middens revealed enhanced enzyme activities and increased mesofaunal abundances. Differences between midden and reference samples were significant during the whole vegetation period and became maximal during ripening of crops. We found a distinct but not significant increase in individual numbers of soil flagellates in midden areas during leafing and heading of crops. Values of the studied parameters tended to be higher in bean plots but generally the crop impact on midden properties was not significant. Our results showed a significant positive synergistic effect between earthworm middens and intermediate crop. Straw mulching supported the hot spot effects of middens.

Contrasting effects of microbial partners in the rhizosphere: interactions between Norway Spruce seedlings ( Picea abies Karst.), mycorrhiza ( Paxillus involutus (Batsch) Fr.) and naked amoebae (protozoa)

The importance of the soil microbial community for plant mineral nutrition and nutrient cycling has long been recognized. One of the most important interactions is the symbiosis of plants with mycorrhizas. In contrast, the effects of soil microfauna on plant performance have so far received little attention, although soil protozoa in particular, have been shown to beneficially affect plant growth. We investigated in a laboratory experiment the impact of mycorrhiza and protozoa and their interaction on plant performance. Spruce seedlings with or without the ectomycorrhizal fungus Paxillus involutus (Batsch) Fr. were grown in microcosm chambers with defaunated forest soil with naked amoebae ( Acanthamoeba sp.) or without protozoa for 10 months. The presence of protozoa resulted in the development of a more complex root system by increasing root length (51%), length of fine roots (64%) and number of root tips (43%). The effects of protozoa were more pronounced in the absence of mycorrhiza. In contrast to protozoa, the presence of mycorrhiza resulted in a less complex root system, i.e. root length, length of fine roots and number of root tips were reduced by 47, 47 and 40%, respectively. Shoot height, and stem, shoot and needle mass were at a maximum in the combined treatment with both mycorrhiza and protozoa. The presence of mycorrhiza and protozoa also affected plant nutrient concentrations. In treatments with protozoa shoots of spruce seedlings contained less nitrogen, leading, e.g. to an increased C/N ratio in needles. Conversely, in treatments with mycorrhiza concentrations of phosphorus in needles were increased by a factor of almost two. Mycorrhiza and protozoa also affected rhizosphere microorganisms. Microbial biomass was reduced in the presence of mycorrhiza, mainly due to a reduction in bacterial numbers. Conversely, in the presence of protozoa the length of hyphae in the rhizosphere was reduced. It is concluded that the plant–mycorrhiza mutualism and the bacteria-mediated mutualism between plants and protozoa (microbial loop) complement each other; plant resources presumably are allocated to optimize simultaneous exploitation of both mutualistic relationships.

Reduced particle size of plant material does not stimulate decomposition, but affects the microbivorous microfauna

The influence of the size of plant litter particles on substrate induced respiration (SIR), inorganic N, respiration activity, protozoa and nematodes in soil was analysed. Finely ground (<2 mm sieve) and larger pieces (4×5 mm) of maize leaves ( Zea mays L.) (C toN=20) and barley ( Hordeum vulgare L.) straw (C to N=190) were added to an arable sandy loam, resulting in four treatments plus a control (no amendment). The microcosms were incubated at 10°C and sampled on days 0, 10, 16, 24 and 71. After day 10, nitrate concentrations in soils with barley and maize were <0.8 and >8.0 μg N g −1 soil, respectively. More N was re-mineralised in the soils amended with finely-ground maize than in those with the coarse maize or barley straw. Respiratory activity peaked during the first few weeks when soils with maize had a higher respiration than soils with barley. During this period, respiration was higher in soils amended with the large pieces on nine out of 10 occasions. Microbial biomass measured as SIR was significantly higher in soils with maize than in those amended with barley, but no effect of particle size was observed (three-way ANOVA, P<0.05). Protozoan numbers were not affected by type of plant material, but significantly higher numbers were found in soil with finely-ground maize than in soil with large pieces (two-way ANOVA, P<0.05). In contrast, large pieces of maize or barley resulted in significant higher nematode numbers than ground material, and maize supported higher populations than barley (three-way ANOVA, P<0.05). The different response of protozoa and nematodes to particle size can be related to their life strategies: protozoa are numerous and have restricted mobility whereas nematodes are larger, with more mobility towards resources.

Bacillus thuringiensis (Bt) toxin released from root exudates and biomass of Bt corn has no apparent effect on earthworms, nematodes, protozoa, bacteria, and fungi in soil

There were no significant differences in the percent mortality and weight of earthworms ( Lumbricus terrestris) after 40 days in soil planted with Bt (NK4640Bt) or non-Bt corn or after 45 days in soil amended with biomass of Bt or non-Bt corn. The toxin was present in the guts and casts of earthworms in soil planted with Bt corn or amended with biomass of Bt corn, but it was cleared within 2–3 days from the guts after placing in fresh soil. There were no significant differences in the colony-forming units of culturable bacteria (including actinomycetes) and fungi and in the numbers of protozoa and nematodes between rhizosphere soil of Bt and non-Bt corn or between soil amended with biomass of Bt and non-Bt corn. The Cry1Ab protein in root exudates and biomass of Bt corn appears not to be toxic to earthworms, nematodes, protozoa, bacteria, and fungi. The presence of the toxin in the guts and casts of earthworms confirmed that the toxin released in root exudates and from transgenic biomass was bound on surface-active particles in soil, which protected the toxin from biodegradation, as has been observed in this laboratory with purified toxin.

Microfaunal primary succession on the of Surtsey, Iceland

The island of Surtsey, Iceland, was formed in 1963 by a volcanic eruption. Since then, it has served as a unique natural laboratory for scientists interested in primary succession. In this study we investigated the state of the soil microfauna succession in 1995. We examined locations on the island with different vegetation types (unvegetated soil, soil with one or two plant species, and bird colony soil with a diverse vegetation). We recorded at least 16 nematode taxa and 13 flagellate taxa. Most of these were not reported in previous surveys from Surtsey. On the location with unvegetated soil, ciliates and nematodes were absent and only amoebae and heterotrophic flagellates were found. Most of the protozoan populations we examined were unable to survive salinity levels corresponding to seawater. We therefore conclude that many of soil protozoa populations on Surtsey arrived to the island as airborne cysts brought there from nearby land. However, in the bird colony soil with a high input of salts from the bird droppings, several flagellate species survived and multiplied at seawater salinity. This indicates that the bird colony soil harbours microhabitats where marine flagellate populations have been established.

Protozoan response to addition of the bacteria Mycobacterium chlorophenolicum and Pseudomonas chlororaphis to soil microcosms

Protozoa are important predators of bacteria in soil and protozoan predation is one of the main factors responsible for the decline of bacterial populations introduced into soil. Bacteria, however, are not equally susceptible to protozoan predation. We have studied the response of indigenous protozoan populations to the introduction of the polychlorophenol-degrading bacterium Mycobacterium chlorophenolicum PCP-1 (DSM 43826), and Pseudomonas chlororaphis (ATCC 43928), into soil microcosms. Introduction of P. chlororaphis to the soil resulted in a huge increase in the numbers of heterotrophic flagellates and naked amoebae during the first 8 days of the experiment. Addition of M. chlorophenolicum to soil caused only a slight increase in protozoan numbers, which was similar to the increase caused by addition of water. There was no indication that addition of M. chlorophenolicum to soil resulted in any increase in the number of protozoa able to feed on this bacterium. The number of colony forming units (CFU) decreased rapidly in the treatment amended with P. chlororaphis cells, whereas there was no decrease in CFUs in the M. chlorophenolicum treatment. The only slight increase in protozoan numbers in the M. chlorophenolicum treatment, as well as the apparently low mortality rate of M. chlorophenolicum in the soil microcosms, coincided with significantly lower soil respiration in the soil microcosms amended with M. chlorophenolicum compared to those amended with P. chlororaphis. The results suggest that the indigenous soil protozoa did not graze on M. chlorophenolicum at all, presumably because it is not a suitable food source.

Microfaunal primary succession on the volcanic island of Surtsey, Iceland

The island of Surtsey, Iceland, was formed in 1963 by a volcanic eruption. Since then, it has served as a unique natural laboratory for scientists interested in primary succession. In this study we investigated the state of the soil microfauna succession in 1995. We examined locations on the island with different vegetation types (unvegetated soil, soil with one or two plant species, and bird colony soil with a diverse vegetation). We recorded at least 16 nematode taxa and 13 flagellate taxa. Most of these were not reported in previous surveys from Surtsey. On the location with unvegetated soil, ciliates and nematodes were absent and only amoebae and heterotrophic flagellates were found. Most of the protozoan populations we examined were unable to survive salinity levels corresponding to seawater. We therefore conclude that many of soil protozoa populations on Surtsey arrived to the island as airborne cysts brought there from nearby land. However, in the bird colony soil with a high input of salts from the bird droppings, several flagellate species survived and multiplied at seawater salinity. This indicates that the bird colony soil harbours microhabitats where marine flagellate populations have been established.

Terrestrial invertebrate abundance across a habitat transect in Keble Valley, Ross Island, Antarctica

Summary The abundance and distribution of soil invertebrates was examined along a transect in Keble Valley, Cape Bird, Ross Island, Antarctica during and after snowmelt. During snowmelt, streams were flowing across the transect, and the moisture environment was highly heterogeneous, whereas after the cessation of stream flow the moisture environment became more homogeneous. Of the seven groups of invertebrates examined, only the prostigmatid mite Stereotydeus mollis showed any redistribution between the two sampling points. The streams had higher water availability, higher soil chlorophyll-a content, higher organic content and lower surface salinity than the areas beside the streams. Nematode ( Panagrolaimus davidi ), soil rotifer and soil protozoan abundance was significantly associated with this area of high productivity, and the cryptostigmatid mites S. mollis and Nanorchestes antarcticus were found near the stream centres. By contrast, the collembolan Gomphiocephalus hodgsoni was found at riparian margins, and it is hypothesised that this could be due to habitat and food preferences.

Soil carbon dynamics of conventional tillage and no-till agroecosystems at Georgia Piedmont — HSB-C models

This model was designed to simulate the dynamics of soil organic matter (SOM) and biomass change of detritus soil food webs after the application of crop residue in agroecosystems. Background data and literature parameters were used to initialize the model and a short-term field experiment was conducted to calibrate the model. Our model is a synthesis and integration of many others’ previous studies because many others’ ideas on food web or SOM modeling were incorporated into our model. In our model, soil microorganisms were considered to be the major decomposers of organic matter with soil animals playing an important role in decomposing organic matter by controlling the population of soil microorganisms through trophic interactions. Though we did not model the soil food web into species or functional groups, we attempted to represent the hierarchical structure and trophic interactions of soil food webs as complete as possible. The whole soil food web was conceptualized as a ‘super organism’ to process organic matter in the model diagram. State variables of our model were categorized into soil organic matter pools, inorganic carbon pool, and soil organism pools. Flows were modeled based on first order kinetics. A tillage effect was introduced in our model by assuming that soil microorganisms, microarthropods and earthworms decrease temporarily immediately after the tillage operation. We also introduced a ‘minimum population’ protection mechanism for each group of soil organisms by assuming that predation ceases when energy obtained can not offset the energy expended in prey-searching and feeding. We found that the simulation outputs fitted the measured data very well, the simulation outputs were in the range of field measurements in most cases. However, there was a significant discrepancy in respiration between simulation output and field measurement in both conventional tillage (CT) and no-till (NT), particularly in NT agroecosystems. We hypothesized that plant roots should be responsible for this discrepancy. The sensitivity analysis of our model showed that the population of soil microorganisms was controlled by resources rather than predators. However, the populations of soil protozoa, nematodes and microarthropods were controlled by both resources and predators.

Estimating the Growth Potential of the Soil Protozoan Community

We have developed a method for determining the potential abundance of free-living protozoa in soil. The method permits enumeration of four major functional groups (flagellates, naked amoebae, testate amoebae, and ciliates) and it overcomes some limitations and problems of the usual 'direct' and 'most probable number' methods. Potential abundance is determined using light microscopy, at specific time intervals, after quantitative re-wetting of air-dried soil with rain water. No exogenous carbon substrates or mineral nutrients are employed, so the protozoan community that develops is a function of the resources and inhibitors present in the original field sample. The method was applied to 100 soil samples (25 plots x 4 seasons) from an upland grassland (Sourhope, Southern Scotland) in the UK. Median abundances for all four functional groups lie close to those derived from the literature on protozoa living in diverse soil types. Flagellates are the most abundant group in soil, followed by the naked amoebae, then the testate amoebae and ciliates. This order is inversely related to typical organism size in each group. Moreover, preliminary evidence indicates that each functional group contains roughly the same number of species. All of these observations would be consistent with soil having fractal structure across the size-scale perceived by protozoa. The method described will be useful for comparing the effects on the soil protozoan community of different soil treatments (e.g. liming and biocides).

Two new terricolous spathidiids (Protozoa, Ciliophora) from tropical Africa: Arcuospathidium vlassaki and Arcuospathidium bulli

Soil protozoa of the tropics are poorly known. However, the few data available indicate that tropical soils are inhabited by a highly diverse protozoan community. In the present paper, some of the many new ciliate species (about 300) that I discovered in African soils are described. Descriptions are based on standard methods, i.e. living and silver-impregnated specimens. Arcuospathidium vlassaki was discovered in a highly saline soil from the margin of the Etosha Pan in Namibia, Southwest Africa. It differs from congeneric species mainly by its rod-shaped macronucleus, fusiform micronuclei, and oblanceolate extrusomes. Arcuospathidium bulli was discovered in a savanna soil of Rwanda, East Africa. It is similar to Arcuospathidium cultriforme Penard in most features, but has a second contractile vacuole in the anterior body half.

Substrate heterogeneity and microfauna in soil organic ‘hotspots’ as determinants of nitrogen capture and growth of ryegrass

In this study we simultaneously manipulated the patchiness of complex organic resources and the composition of microfaunal populations (protozoa and nematodes) in soil, to influence microbial mineralization processes and to elucidate the underlying mechanisms of nutrient acquisition from decomposing plant residues by ryegrass plants.Hotspot treatments of decreasing patchiness were established by filling laboratory microcosms with defaunated soil and adding labelled (13C, 15N) grass residues as 1-layer, 4-layer or completely mixed within the soil. Microfaunal treatments were set up by inoculation of the soil with either protozoa or bacterivorous nematodes, a combination of both or neither (control). The microcosms were planted with surface sterile ryegrass seedlings.Growth of ryegrass plants was enhanced by both, increasing patchiness of the organic matter in soil (1-layer>4-layer>mixed) and microfloral–microfaunal interactions (protozoa+nematodes=protozoa>nematodes>control). The presence of microfauna enhanced the decomposition of hotspot material. Protozoan grazing in particular increased the availability of N in soil and leaching water and led to a concomitant increase in plant growth. While root foraging in organic hotspots enhanced the spatial coupling of mineralization and plant uptake, microfaunal grazing increased the temporal coupling of nutrient release and plant uptake. Consequently the greatest plant biomass was found in treatments combining aggregation of organic material in patches and the presence of microfauna.

Testing genetically engineered potato, producing the lectins GNA and Con A, on non‐target soil organisms and processes

Summary 1. Two lectins, concanavalin A (Con A) and Galanthus nivalis agglutinin (GNA), have anti‐feedant properties that suggest a potential for the control of invertebrate pests of plants. We tested potato plants genetically engineered to produce each of these lectins constitutively, as well as the purified lectins, for possible non‐target effects. 2. Laboratory studies with soil bacterial communities and a ciliate protozoan could detect no direct effect of either lectin over a range of concentrations. There was a significant inhibition in the host‐finding response of a bacterial‐feeding nematode when Con A or GNA was present in the medium at 0·5–50 µg ml−1. 3. A number of GNA‐ and Con A‐producing potato lines had no detectable effects on the rhizosphere microbial and microfaunal populations when examined in pot trials. The incorporation of leaves from transgenic plants into soil reduced protozoan populations significantly, but there was no subsequent effect on the decomposition of added cotton strips. 4. Controlled field‐release experiments demonstrated that, although GNA‐producing potato lines consistently altered the physiological profile of the rhizosphere microbial community at harvest, the effect did not persist from one season to the next over a trial period of two field seasons. There was no significant effect of the best performing GNA line on the development of a subsequent barley crop. 5. A single Con A‐producing line was tested in a controlled field‐release. The only significant effects were transient reductions of c. 40% in soil protozoan populations and of c. 10% in potential microbial activity.

Soil protozoa as bioindicators: pros and cons, methods, diversity, representative examples

This paper emphasizes some general aspects of soil protozoa as indicators of environmental quality—pros and cons, methods, and diversity. Protozoa are at the base of the heterotrophic eukaryotic food web and an essential component in marine, freshwater, and soil ecosystems because they consume a significant portion (usually >50%) of the bacterial productivity, enhancing nutrient cycles and energy flows to the benefit of microorganisms, plants and animals. Accordingly, studies of their dynamics and community structures provide a powerful means for assessing and monitoring changes in the biotic and abiotic environment. This is exemplified by some representative studies focusing on soil oxygen regime, differentiation of humus types, pesticides, global warming, forest decline, movement of protozoan pathogens in soil, and soil protozoan bioassays. Usually, protozoa are not replaceable by higher animals (meso- and macrofauna) as indicator organisms because they have unique physiological properties: they consume more food and have a higher respiration rate per mass unit, have shorter generation and life times, and reproduce much faster. Direct counting methods should be given preference over dilution culture techniques, which are beset with uncontrollable deficiencies. Thus, ciliates and, especially, testate amoebae, whose abundance and diversity can be reliably estimated in simple soil suspensions, should be preferred in environmental studies. About 1600 protozoan species are known to live in terrestrial habitats. However, data from studies of ciliates suggest that this is only a minor portion (20–30%) of the species actually present, most of which are still undescribed. Overcoming the methodological and taxonomic problems are urgent needs which, at present, limit the use of protozoa as bioindicators in terrestrial environments. Furthermore, species monographs are required to compile the taxonomic, faunistic, and ecological information available.