Protozoan bioassays of soil amended with sewage sludge and heavy metals,using the common soil ciliate Colpoda steinii

The common soil protozoan Colpoda steinii was used to study the toxicity of sulphate solutions of Ni, Cd, Cu, and Zn. The growth of C. steinii was reduced by 50% in the presence of 0.10, 0.22, 0.25, and 0.85 mg litre^-1 of Ni, Cd, Cu and Zn, respectively, during 24 h of incubation at 25°C, as calculated from a regression analysis of probit-transformed data. The same growth assay was used to assess the toxicity of soil solution extracted by centrifugation from soil samples of field plots of a grass/clover ley on a sandy loam treated with sewage sludge spiked with additional Cd, Cu, Cr, Ni, Pb, or Zn at concentrations either equivalent to or twice the limits for heavy metals recommended in recent EC guidelines (Commission of European Communities directive 86/278/EEC). The toxicity of these soil solutions varied with the season of the year. None of the soil solutions extracted in winter (February 1991) inhibited the growth of C. steinii. In summer (July 1991), the growth was reduced in solutions extracted from plots that were amended with sludge plus additional Zn or Ni at twice the maxima recommended by the EC. The changes in toxicity to C. steinii of the soil solutions between February and July were positively correlated with increases in heavy metal concentrations of Zn and Ni between winter and summer. These preliminary results suggest that regular protozoan bioassays may be used to monitor the biological availability of heavy metals in soils, especially when combined with other microbial assays and with chemical analyses of soil solutions.     

The effects of mancozeb and lindane on the soil microfauna of a spruce forest: A field study using a completely randomized block design

Summary The effects of mancozeb (fungicide) and lindane (insecticide) were investigated in active soil ciliates, testaceans, rotifers, and nematodes. The effects were evaluated 1, 7, 15, 40, 65, and 90 days after application of a standard and a high (10x) dose. Individual numbers were estimated with a direct counting method. Mancozeb, even at the high dose, had no pronounced acute or long-term effects on absolute numbers of the taxa investigated. The number of ciliate species, which decreased 1 day after treatment with the normal dose (0.05 <P < 0.1), soon recovered. However, the community structure of ciliate species was still slightly altered after 90 days. Testaceans were not reduced before day 15 at the higher dose or before day 40 at the normal one (0.05 <P < 0.1). A normal dose of lindane caused acute toxicity in ciliates and rotifers (P < 0.05) but the latter soon recovered. The number and community structure of ciliate species were still distinctly altered after 90 days (0.05 <P <0.1), indicating the critical influence of lindane. Testaceans were reduced only after day 15, and nematodes only on day 40 (0.05 <P < 0.1). At the high dose of lindane severe long-term effects occurred in soil moisture, total rotifers (P < 0.05), total nematodes (0.05 <P <0.1), and in the structure of the ciliate community. Generally, there were marked differences in the effect of the normal and the high dose of lindane but not with mancozeb. Ciliates showed very pronounced changes after the pesticide applications, indicating their usefulness for testing biocides under field conditions. Testaceans were more resistant than ciliates.     

Comparison between direct counts of active soil ciliates (Protozoa) and most probable number estimates obtained by Singh's dilution culture method

The abundance of active soil ciliates was determined by Singh's dilution culture method any by the direct counting method of Lüftenegger et al. (1988) in 13 samples of 3 types of soil (meadow topsoil, spruce and beech litter). This comparative investigation revealed that the dilution culture methods are likely to overestimate active ciliates by orders of magnitude (with an average factor of 2000). One of the disadvantages of dilution methods is their difficulty meeting the assumptions of the MPN (most probable number) estimation model, on which they are based. A further serious problem is the proper separation of encysted and active cells. Due to a wide range of systematic errors, MPN estimates become seriously biased. But even if the basic assumptions could be met, the maximal precision of the estimate is far too low for a useful application within the context of soil ecology. This was revealed by the calculation of approximate confidence limits. Subjecting the data to analysis of variance showed that the direct counts can clearly differentiate ciliate abundances in forest litter versus meadow soil, whereas the dilution culture method is not precise enough to achieve this distinction. The MPN estimates did not even correlate significantly with the direct counts. Our investigation thus proved that dilution culture methods are beset with methodological deficiencies, resulting in unreliable estimates of protozoan abundances in soil.     

Protozoa and their bacterial prey colonize sterile soil fast

We know little about the ability of protozoa to colonize soils, including their successional patterns. To elucidate this issue, we investigated in which order different protozoan morpho-types colonize sterile soil. We used sterilized soils with different carbon content, and exposed them to the atmosphere for a period of 14 months. Bacteria and flagellates occurred in high numbers after 25 days. Ciliates reached high abundance levels after 137 days, followed by amoebae after 245 days. Numbers of distinguishable protozoan morpho-types increased until 245 days after exposure and declined thereafter. DGGE bands showed a remarkable shift in bacterial diversity with time and a significant increase of detectable gene-types until day 137.     

Fermentation Capacity of Fecal Microbial Inocula of Przewalski Horse,Kulan, and Chapman Zebra and Polysaccharide Hydrolytic Activities of Fecal Microbial Constituents (Ciliates and Bacteria) of Kulan and Chapman Zebra

We examined fermentation capacity of fecal microbial inocula of Przewalski horse (Equus ferus przewalskii), Asian wild ass - kulan (Equus hemionus hemionus), and Chapman zebra (Equus quagga chapmani) in vitro. Interactions of the substrates (amorphous cellulose, wheat straw, meadow hay, xylan from oat spelt, and ground barley grain) and type of fecal inocula in the gas volume and in vitro dry matter digestibility were detected in all substrates after 72 hours of fermentation in five replicates for each substrate and type of inocula. No effects of fecal inocula sources were detected on total short-chain fatty acids concentrations. No live fecal ciliate population was present in kulan feces. Complex ciliate populations in zebra feces and the number and genera resembled ciliates from the colon of horses. Fresh feces of kulan and zebra were fractionated by galvanotaxis and centrifugation to separate fecal ciliates and bacteria. Specific activities (μmol of reducing equivalents/mL min mg protein) of carboxymethyl cellulase (CM-cellulase), xylanase, α-amylase, and inulinase were measured in crude cell-free extract of fecal ciliates (zebra), fecal bacteria (zebra and kulan), and total fecal preparation (zebra and kulan). All examined specific enzymatic activities were present in zebra fecal samples. We were unable to measure the inulinase activity and CM-cellulase activity in kulan fecal samples. Zebra ciliates are actively involved in the digestion of plant storage (α-amylase, 0.53 ± 0.02; inulinase, 1.77 ± 0.01, specific activities) and structural polysaccharides (CM-cellulase, 0.4 ± 0.15; xylanase, 0.26 ± 0.06). For the first time, we measured inulinase activity in intestinal ciliates.     

Population dynamics and ecology of ciliates (Protozoa,Ciliophora) in an anoxic rice field soil

Wetland rice fields cover 1.46 million km² globally; the flooded soil of these fields is largely anoxic. While biogeochemistry and microbiology have been studied in detail, the microbial loop and especially the dynamics and function of ciliates are largely unknown. We used anoxic microcosms prepared with soil from an Italian rice field and recorded species composition, abundance and volume of ciliates together with numbers, volume and size distribution of bacteria. Ciliates were the dominating protists observed in the microcosms, but could be outnumbered by flagellates if the soil was amended with rice straw. The number of ciliate taxa was 23. Metopus species were dominant, but 16 of the species recorded in the anoxic soil were facultative anaerobes. Another 29 species were found in accompanying experiments that included the oxic soil surface. Total abundance in the anoxic soil was on average 110 cells g^–1 dry weight and comparable to that of other soils. The population of ciliates declined around 30 days after flooding, but recovered later. The period before the population declined was characterized by a rapid species turnover, many facultative anaerobes and large species. After recovery, the average cell size was much smaller, but even then a facultative anaerobe, Plagiacampa pentadactyla , was common. About 90% of all species were bacteriovores while the others—mainly Gymnostomatidae—were predators. Grazing ciliates may have controlled bacteria during the first 5 days after flooding, as could be shown by a negative correlation between the respective volumes and by the size spectra of the bacteria.     

Protozoa from aboveground and ground soils of a tropical rain forest in Puerto Rico

Decomposition occurs in the aboveground and ground litter and soils of tropical rain forests, but little is known about the protozoa that stimulate bacterial activity and turnover. I examined litter and ground soils, epiphytic bryophyte soils on tree trunks and branches, and adventitious roots of lianas attached to tree trunks, within 2 m above ground in the Luquillo Experimental Forest, within the Caribbean National Forest, Puerto Rico. Amoebae numbered 69,000–170,000, ciliates 1000–25,000, and testate amoebae 58,000–190,000 g⁻¹ dry wt. of litter, but were reduced by 0.25–0.5 of these abundances in the underlying soils. In the aboveground soils, amoebae numbered 64,000–145,000, ciliates 1000–8000, and testate amoebae 84,000–367,000 g⁻¹ dry wt. of soil. Eighty species of ciliates and 104 species of testate amoebae were found. About 50% of the individuals in ciliate and 33% in testate amoebae populations were small r-selected species, illustrating that functional differences between species determine community composition. Although protozoan numbers are best described as “protozoan potential” because many individuals may be dormant, the high moisture content of tropical rain forest litter and soils suggest almost continually connected soil water films (necessary for protozoan transport), and together with the large numbers and biodiversity of protozoa, suggest that a major proportion of these protozoa contribute to the bacterial decomposition channel of organic matter.     

Distribution of and insights from soil protozoa of the Olympic coniferous rain forest

Decomposition occurs in the surface litter and soil to support temperate rainforests, but little is known about the protozoa that stimulate bacterial activity and turnover. I examined surface litter and top soils, fallen logs, and epiphytes within 2 m from the soil surface in Olympic National Park, USA, of the Pacific Northwest Temperate Coniferous Rain Forest. Ciliates in surface litter numbered 180-580 g⁻¹ dry weight, but were reduced by 20-60% in the underlying soils. Testate amoebae numbered 18,000-77,000 g⁻¹ dry weight in both litter and soil although they were often more abundant in underlying soils. Rotting logs, essential for tree regeneration, supported similar numbers of ciliates, but twice the numbers of testate amoebae. In three epiphytic soils, ciliates numbered 350-550, and testate amoebae 35,000-195,000 g⁻¹ dry weight of soil. In these soils, 26 species of gymnamoebae, 64 species of ciliates, and 113 species of testate amoebae were found. About 65% of the individuals in ciliate and 45% in testate amoebae populations were small, r-selected taxa. Rain forest soil protozoa have distinct testate amoebae populations, and are characterized by enormous biodiversity, the dominance of acrostome species, the proliferation of Euglypha and Nebela species, and the appearance of aquatic taxa. Ecological succession of ciliates and testate amoebae follows an additive (non-replacement) pattern according to a neutral model. The large numbers of persistent r-selected species respond to ecosystem disturbances by mobilizing quickly to resume the bacterivory necessary to help restore the recovering above-ground plant community.     

Filter-out-grazers (FOG): A filtration experiment for separating protozoan grazers in soil

Summary In the present experiment, natural protozoan fauna and other microbial components in water extracts from shortgrass prairie soil were separated on the basis of size by differential filtration (8-, 5-, and 3-m porosities). All extracts contained bacteria and fungi, along with a few very small flagellates (3-m pore size filtrate); flagellates and a few small amoebae (5-m pore size filtrate); and flagellates, small amoebae, and small ciliates (8-m pore size filtrate). All microorganisms, except a few species of flagellates, were present in the centrifuge treatment. Each filtrate was added to sterile soil, and the population of each microbial group was determined after inoculation at intervals up to 80 days (at room temperature). Populations of all added groups decreased on initial addition to soil but then increased during the incubation. By following nitrogen, phosphorus, and CO₂ dynamics, we observed impacts of protozoan grazing on bacteria, including mineralization of N from microbial biomass.     

Motility and migration rate of protozoa in soil columns

Velocity and distances travelled each day by protists in the soil was previously unknown. Soil columns were designed to monitor the distribution of cells along the length of each column as a way to measure migration potential. Motility was affected by pore space, water flow and gravity. The results show that the soil microhabitats that can be explored and exploited by flagellate, amoeboid or ciliated species is in the order of centimetres per day in sandy and sandy loam-textured soil matrices. This value is useful to understand the soil volume that can be explored for food and protist dispersal potential without lateral water flow.