Free‐living nematodes as prey for higher trophic levels of forest soil food webs

Nematodes are the most abundant invertebrates in soils and are key prey in soil food webs. Uncovering their contribution to predator nutrition is essential for understanding the structure of soil food webs and the way energy channels through soil systems. Molecular gut content analysis of consumers of nematodes, such as soil microarthropods, using specific DNA markers is a novel approach for studying predator–prey interactions in soil. We designed new specific primer pairs (partial 18S rDNA) for individual soil‐living bacterial‐feeding nematode taxa (Acrobeloides buetschlii, Panagrellus redivivus, Plectus velox and Plectus minimus). Primer specificity was tested against more than 100 non‐target soil organisms. Further, we determined how long nematode DNA can be traced in the gut of predators. Potential predators were identified in laboratory experiments including nine soil mite (Oribatida, Gamasina and Uropodina) and ten springtail species (Collembola). Finally, the approach was tested under field conditions by analyzing five mite and three collembola species for feeding on the three target nematode species. The results proved the three primer sets to specifically amplify DNA of the respective nematode taxa. Detection time of nematode DNA in predators varied with time of prey exposure. Further, consumption of nematodes in the laboratory varied with microarthropod species. Our field study is the first definitive proof that free‐living nematodes are important prey for a wide range of soil microarthropods including those commonly regarded as detritivores. Overall, the results highlight the eminent role of nematodes as prey in soil food webs and for channelling bacterial carbon to higher trophic levels.     

Separation characteristics of liquid nematode cultures and the design of recovery operations

Production of nematode-based pesticides involves the recovery of a viable nematode life stage known as the infective juvenile (IJ) from fermentation broth. Waste components to be separated from the IJs include non-IJ life stages, dead nematodes, nematode debris, spent media, and the nematode's associated bacteria. This paper reports separation characteristics of liquid cultures and suspensions of the nematodes Phasmarhabditis hermaphrodita, Steinernema feltiae, and Heterorhabditis megidis measured at small scale. Separation characteristics were determined for dead-end filtration, gravity settling and flotation. Results were used to identify large-scale recovery procedures. Separation of culture liquid by dead-end filtration of the crude fermentation broth was not possible due to rapid blinding of filters. However, nematode-water suspensions prepared by gravity settling could be concentrated using this separation method. Settling tests indicated that IJs could be efficiently separated from culture liquid by centrifugation but not by gravity settling. Examination of the effects of nematode concentration indicated an optimum concentration for gravity settling that may entail modest dilution of the fermentation broth. Flocculation of insoluble spent media in suspensions of P. hermaphrodita prevented its separation from nematodes by gravity settling. However, attachment of air bubbles to spent media allowed removal by flotation. Finally, adjustment of continuous phase density using sucrose allowed separation of non-IJ life stages, dead nematodes, and discarded cuticles from the IJs by flotation. The efficiency of this separation decreased with increasing nematode-solute contact time.     

Collembola as alternative prey sustaining spiders in arable ecosystems: prey detection within predators using molecular markers

Collembola comprise a major source of alternative prey to linyphiid spiders in arable fields, helping to sustain and retain these predators as aphid control agents within the crop. Polymerase chain reaction primers were developed for the amplification, from spider gut samples, of DNA from three of the most abundant species of Collembola in wheat crops in Europe, namely Isotoma anglicana, Lepidocyrtus cyaneus and Entomobrya multifasciata. The primers amplified fragments of the mitochondrial cytochrome oxidase subunit I (COI) gene and were designed following alignment of comparable sequences for a range of predator and prey species. Each of the primer pairs proved to be species-specific to a Collembola species, amplifying DNA fragments from 211 to 276 base pairs in length. Following consumption of a single collembolan, prey DNA was detectable in 100% of spiders after 24 h of digestion. We report the first use of DNA-based techniques to detect predation by arthropods on natural populations of prey in the field. All three species of Collembola were consumed by the spiders. By comparing the ratios of the Collembola species in the field with the numbers of spiders that gave positive results for each of those species, it was possible to demonstrate that the spiders were exercising prey choice. Overall, a single target species of Collembola was eaten by 48% of spiders while a further 16% of spiders contained DNA from two different species of Collembola. Preference was particularly evident for I. anglicana, the species most frequently found in spider guts yet the least numerous of the three target species in the field.     

Seasonal changes in soil arthropod abundance in the dry evergreen forest of north-east Thailand,with special reference to collembolan communities

A soil arthropod community was studied in a dry evergreen forest over a 3-year period from May 1998 to April 2001. Population abundance, species composition, and community structure were investigated over the 3-year study period. The soil arthropods consisted of Acari (75.38%), Collembola (16.11%), and others (8.51%), and their abundances showed a clear difference between the rainy and dry seasons. Population abundance of Collembola and Acari were low during drought conditions. The humidity was the most important factor determining distribution, abundance, and survival of soil Collembola in this tropical forest. High predation and low accumulation of organic matter caused low population abundance of Collembola in the tropical habitat. The collembolan community was dominated by a few dominant species over the study period. The pattern of seasonal changes in numbers of Collembola was similar over the 3-year study period. The species composition of the collembolan community was constant and persistent throughout a 3-year study period. Thus, the collembolan community showed constancy in its species composition with seasonal variability over the 3-year study period.     

Susceptibility and immune response of Deroceras reticulatum, Milax gagates and Limax pseudoflavus exposed to the slug parasitic nematode Phasmarhabditis hermaphrodita

We exposed three slug species (Deroceras reticulatum (Müller), Milax gagates (Draparnaud) and Limax pseudoflavus L.) to the parasitic nematode Phasmarhabditis hermaphrodita Schneider. P. hermaphrodita was able to cause mortality and feeding inhibition to both D. reticulatum and M. gagates but did not negatively affect L. pseudoflavus. On dissection of surviving L. pseudoflavus large numbers of P. hermaphrodita were found encapsulated in the shell of the slug. We found that by increasing shell size, the slug was able to trap invading nematodes, which could be an immune response to P. hermaphrodita invasion. This is the first report of a slug defense mechanism to inhibit P. hermaphrodita.     

Prey preference,intraguild predation and population dynamics of an arthropod food web on plants

The theory of intraguild predation (IGP) largely studies effects on equilibrium densities of predators and prey, while experiments mostly concern transient dynamics. We studied the effects of an intraguild (IG) predator, the bug Orius laevigatus, on the population dynamics of IG-prey, the predatory mite Phytoseiulus persimilis, and a shared prey, the phytophagous two-spotted spider mite Tetranychus urticae, as well as on the performance of cucumber plants in a greenhouse. The interaction of the predatory mite and the spider mite is highly unstable, and ends either by herbivores overexploiting the plant or predators exterminating the herbivores. We studied the effect of IGP on the transient dynamics of this system, and compared the dynamics with that predicted by a simple population-dynamical model with IGP added. Behavioural studies showed that the predatory bug and the predatory mite were both attracted to plants infested by spider mites and that the two predators did not avoid plants occupied by the other predator. Observations on foraging behaviour of the predatory bug showed that it attacks and kills large numbers of predatory mites and spider mites. The model predicts strong effects of predation and prey preference by the predatory bugs on the dynamics of predatory mites and spider mites. However, experiments in which the predatory bug was added to populations of predatory mites and spider mites had little or no effect on numbers of both mite species, and cucumber plant and fruit weight.     

Evaluation of Blattisocius dolichus (Acari: Blattisociidae) for biocontrol of root-knot nematode,Meloidogyne incognita (Tylenchida: Heteroderidae)

The root-knot nematode, Meloidogyne incognita Kofoid and White (Tylenchida: Heteroderidae), is one of the most important plant parasitic nematodes attacking many plant roots. In this paper, the predation and biocontrol efficiency of Blattisocius dolichus Ma (Acari: Blattisociidae) on this nematode were studied. The predation rate and the environmental factors affecting predation rate of B. dolichus on second-stage juveniles of M. incognita (Mi-J2) were studied in experimental arenas in plastic dishes. Both female and male mites had greater capability in consuming Mi-J2, and daily predation rate of female and male mites was not less than 35 and 29.6 nematodes within seven days, respectively. Temperature, nematode density and starvation affected B. dolichus predating on Mi-J2, and when offered 300 nematodes per dish, the predation rate of mites starved for 96 h was the highest at 25 °C, with female and male mites consumed 50.5 and 54 nematodes in 24 h, respectively. The biocontrol of B. dolichus against M. incognita was carried out on potted water spinach Ipomoea aquatic in a greenhouse. The water spinach roots were inoculated with 1,000 Mi-J2 ten days after releasing 100, 200, 300, 400 and 500 mites per pot. Compared to the nematodes–alone control, the number of root-knots of all mite treatments and the number of egg masses of the treatments with 300, 400 and 500 mites significantly decreased. Effect of releasing 500 mites on control of the root nematode M. incognita was best among all mite treatments, reduced the root-knots and egg masses 37.1 and 55.1 %, respectively, but no significant difference was observed compared to 400 mites treatment.     

Brain organization in Collembola (springtails)

Arthropoda is comprised of four major taxa: Hexapoda, Crustacea, Myriapoda and Chelicerata. Although this classification is widely accepted, there is still some debate about the internal relationships of these groups. In particular, the phylogenetic position of Collembola remains enigmatic. Some molecular studies place Collembola into a close relationship to Protura and Diplura within the monophyletic Hexapoda, but this placement is not universally accepted, as Collembola is also regarded as either the sister group to Branchiopoda (a crustacean taxon) or to Pancrustacea (crustaceans + hexapods). To contribute to the current debate on the phylogenetic position of Collembola, we examined the brains in three collembolan species: Folsomia candida, Protaphorura armata and Tetrodontophora bielanensis, using antennal backfills, series of semi-thin sections, and immunostaining technique with several antisera, in conjunction with confocal laser scanning microscopy and three-dimensional reconstructions. We identified several neuroanatomical structures in the collembolan brain, including a fan-shaped central body showing a columnar organization, a protocerebral bridge, one pair of antennal lobes with 20-30 spheroidal glomeruli each, and a structure, which we interpret as a simply organized mushroom body. The results of our neuroanatomical study are consistent with the phylogenetic position of Collembola within the Hexapoda and do not contradict the hypothesis of a close relationship of Collembola, Protura and Diplura.     

Consumption rate of phytonematodes by <Emphasis Type="Italic">Pergalumna</Emphasis> sp. (Acari: Oribatida: Galumnidae) under laboratory conditions determined by a new method

Although several generalist species of Pergalumna are known to be nematode predators, the potential of oribatid mites as natural enemies of phytonematodes has been underestimated. The objective of this work was to estimate the consumption rate of a Pergalumna sp. when feeding on two major pest nematodes, Meloidogyne javanica and Pratylenchus coffeae, under laboratory conditions. A new method was used, in which live nematodes are offered to mites and subsequently consumption is quantified based on the sclerotized, well preserved structures in the mite's fecal pellets. The assay was evaluated during 5 days, at 25 degrees C and 96% relative humidity, with three replicates for each nematode species. Every replicate consisted of a group of four mites isolated in an arena, to which 400 nematodes were transferred daily. The daily produced fecal pellets were mounted in Hoyer's medium for examination under a microscope. The nematode buccal stylets and cephalic frameworks were counted to estimate the number of nematodes consumed. It was estimated that a single mite daily ingested 18.3 +/- 0.8 (mean +/- SE) M. javanica (J(2) juveniles) or 41.6 +/- 7.2 P. coffeae (juveniles + adults), the maximal daily consumption being 34 M. javanica and 73 P. coffeae. The method showed to be practical, precise and suitable for laboratory studies in which nematophagous mites classified as engulfers are included.     

Refuge from predation, the benefit of living in an extreme acidic environment?

Organisms living in extreme habitats require costly adaptations to cope with these conditions. Among the suggested potential benefits that trade off these costs is refuge from predation. To study these interactions in extreme environments, samples were taken in the cave Cueva de Villa Luz, Tabasco, Mexico, where more than 32 subterranean springs, some H(2)S rich, rise from the floor. Hydrogen sulfide gas plus oxygen is absorbed by freshwater, and oxidation forms concentrated sulfuric acid. Snottites, whitish hollow mucous tubes, hang from the ceiling of the cave. Fluid drops from these snottites were recorded as having pH values of 0-3. We report the discovery of a new species of nematode that thrives in the highly acidic environment of the snottite. Micro CT scan of snottites reveals a complex interaction between the acidic snottite, nematodes, and abundant nematode-eating mites. The nematode adaptation to low pH probably protects them against mite predation, for which nematodes are most likely the most important source of carbon in this sulfur-driven ecosystem.     

Collembolan associations in natural versus improved pastures of the New England Tableland,NSW: Distribution of native and introduced species

Native Australian species of Collembola dominated natural pasture in numbers of species present and in abundance, while introduced Collembola dominated fertilized pasture sown to exotic species. Those collembolan species which were restricted to one pasture type only were mainly epigeal and hemiedaphic species of the herbage and litter. Increased grazing by sheep decreased species richness and increased uniformity of Collembola in both pasture types. The abundance of introduced Collembola was positively associated with phosphorus content of litter.     

Predation of entomopathogenic nematodes by <Emphasis Type="Italic">Sancassania</Emphasis> sp. (Acari: Acaridae)

Predation of the entomopathogenic nematode, Steinernema feltiae (Rhabditida: Steinernematidae), by Sancassania sp. (Acari: Acaridae) isolated from field-collected scarab larvae was examined under laboratory conditions. Adult female mites consumed more than 80% of the infective juvenile (IJ) stage of S. feltiae within 24 h. When S. feltiae IJs were exposed to the mites for 24 h and then exposed to Galleria mellonella (Lepidoptera: Pyralidae) larvae, the number of nematodes penetrating into the larvae was significantly lower compared to S. feltiae IJs that were not exposed to mites (control). Soil type significantly affected the predation rate of IJs by the mites. Mites preyed more on nematodes in sandy soil than in loamy soil. We also observed that the mites consumed more S. feltiae IJs than Heterorhabditis bacteriophora (Rhabditida: Heterorhabditidae). No phoretic relationship was observed between mites and nematodes and the nematodes did not infect the mites.     

Effects of plant diversity on Collembola in an experimental grassland ecosystem

The response of species numbers and density of Collembola to manipulation of plant species richness (1, 2, 4, 8, 32 species) and number of plant functional groups (grasses, legumes and non-legume herbs) was studied in an experimental grassland at the Swiss BIODEPTH site (Lupsingen, Switzerland) in October 1997. Plant species richness or number of plant functional groups did not affect total diversity of Collembola, however, the number of Collembola species increased in the presence of legumes and the grass Trisetum flavescens . The abundance of Protaphorura armata increased but that of Hypogastruridae/Neanuridae significantly decreased with increasing number of plant functional groups. Other groups including the herbivorous Symphypleona did not respond to plant species richness and plant functional groups. Possibly, Hypogastruridae/Neanuridae species are weak competitors declining in density if the density of other Collembola groups increase. In general, the effect of the number of plant functional groups on the densities of collembolan taxa was stronger than that of plant species richness. Changes in Collembola density and diversity in part was likely caused by increased soil microbial and fine root biomass in treatments with higher plant functional group diversity. The presence of legumes resulted in an increase in the densities of total Collembola, Symphypleona/Neelipleona and Isotomidae indicating that they benefited from the high litter quality and the increased microbial biomass in the rhizosphere of legumes. The results suggest that microbivorous soil invertebrates are controlled by food quality rather than quantity. Furthermore, they indicate that presence of certain plant species and functional groups may be more important for collembolan community structure than the diversity of plant species and functional groups per se.     

Host-plant species modifies the diet of an omnivore feeding on three trophic levels

The diet choice of omnivores feeding on two adjacent trophic levels (either plants and herbivores or herbivores and predators) has been studied extensively. However, omnivores usually feed on more than two trophic levels, and this diet choice and its consequences for population dynamics have hardly been studied. We report how host-plant quality affects the diet choice of western flower thrips feeding on three trophic levels: plants (cucumber or sweet pepper), eggs of spider mites and eggs of a predatory mite that attacks spider mites. Spider mites feed on the same host plants as thrips and produce a web that hampers predator mobility. To assess the indirect effects of spider mites on predation by thrips, the thrips were offered spider-mite eggs and predatory-mite eggs on cucumber or sweet pepper leaf discs that were either clean, damaged by spider mites but without spider-mite web, or damaged and webbed. We show that, overall, thrips consumed more eggs on sweet pepper, a plant of low quality, than on cucumber, a high quality host plant. On damaged and webbed leaf discs (mimicking the natural situation), thrips killed more predator eggs than spider-mite eggs on sweet pepper, but they killed equal numbers of eggs of each species on cucumber. This is because web hampered predation on spider-mite eggs by thrips on sweet pepper, but not on cucumber, whereas it did not affect predation on predatory-mite eggs. We used the data obtained to parameterize a model of the local dynamics of this system. The model predicts that total predation by the omnivore has little effects on population dynamics, whereas differential attack of predator eggs and spider-mite eggs by the omnivore has large effects on the dynamics of both mite species on the two host plants.     

Convergent evolution of defense mechanisms in oribatid mites (Acari, Oribatida) shows no "ghosts of predation past"

Oribatid mites are diverse and abundant terrestrial soil arthropods that are involved in decomposition of organic matter and nutrient cycling. As indicated by fossils starting from the Devonian, they evolved varied mechanisms and structures for defense from predators. We investigated four of these defensive structures (ptychoid body, hologastry, mineralization and opisthonotal glands) and used ancestral character state reconstruction to determine whether they evolved convergently and how many times this may have happened. Phylogenetic trees based on 18S rDNA were constructed for 42 oribatid mite species and two outgroup taxa using likelihood and Bayesian algorithms. The results suggest that at least three of the four defensive structures evolved convergently several times; for opisthonotal glands convergent evolution remains equivocal. This high level of convergence indicates that predation has been an important factor throughout the evolution of oribatid mites, contributing to morphological diversity and potentially also to species richness, as there are indications that some taxa radiated after the evolution of defense structures. Despite the ancientness of oribatid mites, defense structures seems to have been rarely lost, suggesting that they still are functional and necessary to reduce predation, rather than being 'ghosts of predation past'.     

Zoophagy in Collembola

The feeding of collembolans on animal food, including nematodes and small enchytraeids, was studied in laboratory. Active preying on worms was found in more than 30 Collembola species. Fecundity and natural growth of the collembolan populations were similar in the case of separate diet (fungal and animal), but mixed feeding increased both parameters by several times. No special hunting behavior was found, and feeding on nematodes was very similar to gathering. Several collembolans attack small enchytraeids together, dismembering living worms.     

Effect of osmotic desiccation on longevity and temperature tolerance of Phasmarhabditis hermaphrodita (Nematoda: Rhabditidae)

Limited storage stability severely restricts the biological control potential of slug-parasitic nematodes. In a series of experiments, we evaluated the effects of temperature and osmotic desiccation on the short- and long-term survival of the slug-parasitic nematode Phasmarhabditis hermaphrodita. Nematode survival in petri dishes at 1,500 infective juveniles/ml did not differ significantly at 5, 10, and 15 C but declined rapidly at 25 and 30 C. At 25 C about 50% of the nematodes survived for 4 wk, but at 30 C no nematode survived past 1 day. About 50% of the nematodes survived for 32 wk at 20 C. About 35-40% of the nematodes survived up to a year at 5, 10, and 15 C. Phasmarhabditis hermaphrodita showed poor survival under osmotic desiccation in glycerol with 15 and 20% glycerol significantly reducing survival at 5 and 15 C. Although the nematodes tolerated 10% glycerol, this level of desiccation also did not enhance long-term survival at either 5 or 15 C. There was a significant decrease in nematode survival in 10% glycerol at 25 C during the first 2-3 wk, but about 16% of the nematodes survived for 6 wk in 10% glycerol as compared with only 1% survival in water. The greatest benefit of osmotic desiccation in glycerol was observed in the enhanced survival of P. hermaphrodita at temperature extremes. Over 96% of the nematodes survived a 6-hr exposure to 35 C in 10% glycerol, whereas only 9% survived in water. Similarly, over 90% of the nematodes survived an exposure to -20 C for 4 hr in 10% glycerol, but less then 2% survived in water. We conclude that 5-15 C is an optimum temperature range for the storage of P. hermaphrodita. We also conclude that osmotic desiccation in 10% glycerol can substantially increase survival of P. hermaphrodita at temperature extremes (35 and -20 C) for short periods but has no effect on nematode longevity at the optimum temperature range of 5-15 C.     

Diversity and distribution of Collembola (Arthropoda: Hexapoda) of Brazil

Objectives of this study are to summarize the current state of knowledge of the diversity and distribution of Collembola in Brazil; to identify areas of most importance, interest, and need of additional research; and to enable development of experimental hypotheses for future research on Collembola in this region. The total number of collembolan species currently known from Brazil is 199, distributed among 19 families and 80 genera. The greatest numbers of species have been recorded from the states of Rio de Janeiro and Amazonas (with 69 and 56 species, respectively). Few or no species are known from most Brazilian states. Most of the species for which specific Brazilian environmental habitat information is available (93, 66%) are known only from forest environments. Most of the species (127, 64%) are known only from Brazil (most likely being endemic); 33 species (17%) are known only from Brazil and other neotropical areas; and 39 species (20%) have a distribution beyond the neotropical region. Results of this study indicate that much remains to be learned about the Brazilian collembolan fauna. This is especially true for areas of the northeastern, central-western, and southern regions. Studies to determine the species composition of collembolan communities in specific environments in Brazil are needed. The great diversity of Collembola species in Brazil is largely unknown and there are many opportunities for additional research on these environmentally important organisms in this area. Such additional research on the Collembola in Brazil is also essential for a better understanding of the neotropical (and world) collembolan fauna.     

Factors limiting short-term persistence of entomopathogenic nematodes

Abstract:  Entomopathogenic nematodes are commonly applied to soil and crops for biocontrol of insects. Typically, when nematodes are applied, recoverable numbers decline quickly. Because ultra-violet light is known to kill entomopathogenic nematodes in the laboratory, many researchers blame poor field persistence on the action of ultra-violet light. We conducted a field experiment to test this hypothesis, using Heterorhabditis bacteriophora applied to turf as a model system. We compared persistence of surface-applied nematodes with subsurface-applied nematodes and found no difference in persistence. Numbers of potential nematode antagonists (mites and collembola) were also monitored. Poor persistence was positively correlated with numbers of mites and collembola in plots where nematodes were surface-applied, but not in plots where they were subsurface-applied.     

Population- and ecosystem-level effects of predation on microbial-feeding nematodes

We studied the role of nematode predation in the functioning of detrital food webs assembled in microcosms. The microcosms contained defaunated humus and litter materials, a diverse microbial community with bacteria, fungi and protozoa, and a birch (Betula pendula) seedling infected with mycorrhizal fungi. Different levels of top-down control upon microbivorous nematodes were set up by assembling food webs either without predators, or in combinations with a specialist and a non-specialist predatory mite (Mesostigmata). The nematode community was composed of either (1) three species of bacterivorous, or (2) three species of fungivorous nematodes or (3) both groups together. After two growing periods for the birch (38 weeks), the microcosms were destructively sampled for animal and microbial biomasses, concentration of mineral N in the soil, plant biomass and plant N concentration. The specialist predator reduced biomasses of both bacterial- and fungal-feeding nematodes by more than 50%, whereas the non-specialist predator weakly increased the biomass of fungivorous nematodes. Thus, under high predation pressure, the biomass of microbivores changed as predicted by trophic dynamic models assuming strong top-down control and uniformly behaving trophic levels. Despite this, microbial biomass was unaffected by the predators. However, microbial respiration increased slightly in the presence of predators. Assuming that microbial respiration correlates with microbial productivity, the increase in microbial respiration indicates a cascading productivity regulation. The composition of the microbivore community had only a minor effect on the outcome of the top-down control on microbes. The >50% reduction in nematode biomass and respiration coincided with <16% increase in microbial respiration and did not affect microbial biomass. Presence of the specialist predator slightly reduced soil NH⁺ ₄ concentration in communities with fungivore nematodes but plant growth and N uptake remained unchanged. Thus, the structure of the community only weakly controlled nutrient mineralisation. Received: 18 May 1998 / Accepted: 3 May 1999