Springtail Orchesella Cincta Research Articles

Problems and pitfalls in cable analysis in short tubular epithelia. Analysis of regional functional differences, a model

Data from cable analysis give an estimate of the actual physiological values only, if there is no substantial electrical leak in the set-up and if the Rte is evenly distributed along the preparation. A model is proposed to assess the transepithelial resistance (Rte) of short tubular epithelia with unequally distributed Rte. The midgut of the springtail Orchesella cincta (L.) is short and shows no obvious morphological differentiation. Nevertheless differences in transport function (e.g. Rte) along the length are to be expected. Midgut pieces of different lengths (0.2 to 1.4mm) were luminally perfused from the oral to the aboral end. The Rte increased significantly from 70 Ω·cm2 to 1268 Ω·cm2 (p<0.00001; average: 380 ± 36SEM Ω·cm2; n = 57). Estimates of Rte from measurements in preparations with high Rte, may be obscured by electrical leakage between the lumen and the surrounding bath. Extrapolating the input resistances of midgut pieces of different lengths to zero length gave an estimate of these seal resistances (372 kΩ). Midguts (0.25 to 1.5mm) were also perfused in the other direction, i.e. from aboral to oral end, to elucidate whether leakage or true differences in Rte along the midgut explained the previous results. Overall the data confirmed a higher aboral Rte. The Rte distribution along the midgut could be mimicked by an electrical equivalent circuit. The midgut was divided into 10 subsequent loops in the model circuit. Inserting values for leakage, resistances and cell potentials, based on the experimental data, revealed, that the actual midgut Rte μαψ ηαϖε ινχρεασεδ 45–ϕολδ ϕρομ 150 Ω·cm2 at the oral side to 6700 Ω·cm2 at the aboral side.

Metallothionein mRNA Expression and Cadmium Tolerance in Metal-stressed and Reference Populations of the Springtail Orchesella cincta

Metal contamination in soil ecosystems is a permanent and often strong selection pressure. The present study investigates metal tolerance in 17 Orchesella cincta (Collembola) populations from metal-contaminated and reference sites, and combines analyses at the phenotypic and molecular level. Metal tolerance was phenotypically assayed by measuring survival times of laboratory cultures during exposure to cadmium. Comparisons of survival curves showed that five out of eight metal-stressed populations tested evolved increased cadmium tolerance (Stolberg, Plombieres, Hoboken, Hygum and Gusum). In addition, the role of the metallothionein (MT) gene in cadmium tolerance of O. cincta was studied by means of quantitative RT-PCR. The constitutive and Cd-induced MT mRNA expression of the laboratory cultures was measured. Results show that the mean constitutive MT mRNA expression of populations from polluted sites was significantly higher than of populations from reference sites. However, no correlation between MT mRNA expression levels after laboratory exposure to cadmium and field cadmium concentrations was observed. Furthermore, no relation between survival rate during exposure to cadmium and MT mRNA expression was detected. Our results suggest that constitutive MT mRNA expression plays a role in early protection against cadmium toxicity, and indicate that mechanisms other then MT up-regulation are involved in tolerance to prolonged exposure to cadmium.

Population genetic structure of Orchesella cincta (Collembola; Hexapoda) in NW Europe, as revealed by microsatellite markers

Summary We studied genetic variation and population differentiation in the springtail Orchesella cincta L. An earlier approach, using allozymes, revealed extremely low variation among and within populations from NW Europe. Microsatellite marker analysis showed higher genetic variation than allozymes, and agreed with the previously reported low population differentiation in the species. Analysis of molecular variance showed that genetic variation within populations amounted to 96.5% of the total variation, whereas the remaining 3.5% is accounted for by population differentiation. Population differentiation, as described by microsatellite markers and with the exception of one population, can be explained by an isolation by distance. Although the microsatellites are short, viz. only up to 12 repeat units, they appear to be useful tools in revealing population genetic structures in O. cincta .

Evidence for multiple origins of Wolbachia infection in springtails

Summary Bacteria of the genus Wolbachia (Rickettsiae) are endosymbionts found in many species of invertebrates. The traditional division of Wolbachia lines into four supergroups has recently been challenged with the finding of a fifth supergroup. Until now, this E supergroup exists of a single host species of the order Collembola (springtails) and is hypothesized to form a ‘missing link’ between Wolbachia lines in higher arthropods and nematodes. We have sequenced the 16S rDNA, the ftsZ and the wsp genes of a Wolbachia lineage present in the sexually reproducing collembolan Orchesella cincta. We tested the hypothesis on the origin of Wolbachia in Collembola by adding this new Wolbachia lineage and a lineage found in the collembolan Mesophorura macrochaeta to the Wolbachia phylogenetic tree. The addition of these two species clearly shows that Wolbachia is not monophyletic in Collembola. We conclude that the origin of Wolbachia in Collembola is a combination of vertical and horizontal transfer between coexisting species.

Indications for the tracking of elevated nitrogen levels through the fungal route in a soil food web

The objective of the present study was to determine the effects of elevated N in dead organic matter on the growth of fungi and to establish the consequences for the development of microbivores. Therefore, three fungal species were cultured on Scots pine litter differing in N content. The growth of the soil fungal species Trichoderma koningii, Penicillium glabrum and Cladosporium cladosporioides was directly influenced by the N content (ranging from 1.25 to 2.19% N) of the substrate. For all three fungal species maximum growth was highest at intermediate N content (1.55%) of the substrate. The fungivorous collembolan Orchesella cincta reached highest asymptotic body mass when fed with C. cladosporioides, grown on litter medium with intermediate N content (1.55%). The growth of O. cincta was lower when fed with C. cladosporioides from litter medium with the highest N content (2.19%). Similar results were obtained in mesocosm experiments in which pine litter with three levels of N (1.11, 1.78, 2.03% N) was used as substrate for the fungi. On litter with the highest N content (2.03%) hyphal length and asymptotic body mass of O. cincta were reduced. The results show that the N content of the substrate determines the growth of both fungi and fungivores, and suggest that elevated levels of N in soil track through the fungal part of the soil food web.

Field-selected cadmium tolerance in the springtail Orchesella cincta is correlated with increased metallothionein mRNA expression

Populations of the springtail Orchesella cincta that live in metal contaminated soils have developed tolerance to cadmium by increased metal retention in the midgut epithelium and excretion at every moult. Regulation of the MT gene was studied in a tolerant population (Plombières, Belgium) and a laboratory culture. Animals were exposed to a range of concentrations of cadmium in the food (0–1.5 μmol Cd/g food). RNA was extracted after 5–14 days of cadmium exposure and used for Northern blot analysis to quantify MT mRNA. MT expression levels were significantly higher ( p<0.01) in individuals from laboratory-raised strains originating from the soils of the metal contaminated forest Plombières (2.4- to 7.8-fold expression) compared to the reference population (1.5- to 2.4-fold expression). No variable sites were found in the complete MT coding sequence. Southern blot analysis suggests that in both populations the gene is not tandemly repeated. This is the first evidence of evolution of metal tolerance via gene regulation of MT in a natural population. These data indicate a higher fitness of the tolerant population in the polluted environment due to selection of high MT expression phenotypes.

Dietary zinc reduces uptake but not metallothionein binding and elimination of cadmium in the springtail, Orchesella cincta

In metal-polluted environments, high concentrations of metals such as zinc, cadmium, copper, and lead are present. Many studies have been conducted to determine uptake and binding of the nonessential, toxic metal cadmium. Effects of essential metals on this uptake and binding remain unclear. The aim of the present study was to determine the effect of zinc on cadmium kinetics and metallothionein induction in the springtail Orchesella cincta. Exposure experiments were conducted with cadmium only, zinc only, and cadmium + zinc. Metallothionein was isolated to determine zinc binding to or induction of this cadmium-binding protein. In animals exposed to cadmium only and to cadmium + zinc, metallothionein was induced. In animals exposed to zinc only, no metallothionein was induced, and zinc did not bind to the metallothionein induced by cadmium. Cadmium uptake rate was significantly lower in animals exposed to cadmium + zinc than in animals exposed to cadmium only. No significant pattern of uptake and elimination of zinc was observed. Zinc concentrations in the animals were not significantly different for the different exposures, demonstrating that this element is regulated by O. cincta. These results suggest a protective role of zinc in the uptake of cadmium but no interference of zinc with the internal cadmium distribution and elimination.

Distant lovers: spermatophore deposition and destruction behavior by male springtails.

Males of the springtail Orchesella cincta transfer their sperm indirectly using spermatophores. The present study aimed to investigate how individual males maximize their reproductive success. In an experiment examining the role of chemical communication, it appeared that the males deposit their spermatophores preferentially on patches which have been conditioned with conspecifics. Varying conditioning intensity resulted in a response which can be described by a saturation curve. In a comparison involving a related species the conditioning effect was strictly species specific. An experiment where the sexes were compared showed that conditioning with males exerts a slightly stronger effect than conditioning with females. In another experiment spermatophore depositing males showed a strong preference for patches with spermatophores of previous males, which they replaced with their own. An explanation for this may be that the males parasitize each other's scent marks. When a number of males are kept together they destroy most of each other's spermatophores. The destruction behavior is specifically displayed by males in their reproductive phase. Video recordings showed that deliberate destruction of spermatophores takes place by eating them and that the males are able to recognize and spare their own spermatophores.

Microsatellite loci in the soil‐dwelling collembolan, Orchesella cincta

AbstractThree total genomic libraries of the springtail Orchesella cincta (Insecta; Collembola) were screened for the presence of microsatellites using phosphor and colour detection. PCR primers were successfully constructed for six dinucleotide, one trinucleotide, and one interrupted dinucleotide microsatellite repeat. None of the microsatellite arrays were longer than 11 repeat units. Individuals from three forests were investigated: five out of eight microsatellite markers were polymophic in all forests (2–5 alleles per locus) and the observed heterozygosity was 0.1–0.9 per locus.

Metallothionein-bound cadmium in the gut of the insect Orchesella cincta (Collembola) in relation to dietary cadmium exposure

Metallothionein is considered to be a potential biomarker for heavy metal exposure in the terrestrial environment. However, limited information is available on metallothioneins from insects, a major class of terrestrial invertebrates. In this study we have quantified metallothioneins in the springtail Orchesella cincta by determining metallothionein-bound cadmium after separation of these proteins using gel filtration and reversed phase chromatography from total body homogenates of animals dietary exposed to different concentrations of cadmium. Furthermore, we have studied in more detail where cadmium and metallothionein-bound cadmium is located within this animal. The concentration of metallothionein-bound cadmium increases with the exposure concentration in the same way as the total internal concentration. Both reach a plateau at an exposure concentration of approximately 1.0 μmol Cd/dry food. Cadmium is primarily located within the gut of O. cincta and isolation of metallothionein from this organ gives results identical to isolations from total bodies. Based on this results an estimation of the metallothionein level at the highest exposure concentration results in a concentration of about 115 μg metallothionein/g fresh gut. The O. cincta metallothionein gives the possibility of using this protein as a biomarker for heavy metal exposure in soil insects.

Primary structure of a cadmium-induced metallothionein from the insect Orchesella cincta (Collembola).

The induction of metallothionein was studied in the springtail Orchesella cincta (Collembola), a species of insect living in forest soils. Upon dietary exposure to Cd, two Cd-binding, cysteine-rich peptides were isolated from whole-body homogenates, using gel filtration and reversed-phase FPLC. Mass spectrometric analysis revealed that the molecular masses of these peptides were 2989 Da and 4139 Da, respectively. Amino acid sequencing of the 2989-Da peptide resulted in a sequence typical for a metallothionein. Sequencing of the 4139-Da protein was unsuccessful, probably due to N-terminal blockage. Using different PCR techniques (3' and 5' RACE) with (degenerate) primers based on the identified amino acid sequence of the 2989 Da peptide, a metallothionein cDNA was isolated. The sequence of this cDNA potentially codes for a protein of 77 amino acids. The 2989 Da peptide corresponds to the C-terminal part of this protein. The 4139-Da protein is probably encoded by the N-terminal part of this protein. These results suggest that the identified peptides are products of one gene, and that the primary gene product is subject to post-translational processing. The deduced amino acid sequence of the O. cincta metallothionein shows low sequence similarity with metallothioneins from Drosophila. The similarity between O. cincta MT and MTs of invertebrates is not higher than that between O. cincta and vertebrates.

Modelling the time–temperature relationship in cold injury and effect of high‐temperature interruptions on survival in a chill‐sensitive collembolan

1. Temperature‐ and time‐dependent mortalities were studied and modelled in insects exposed in regimes with constant and alternating temperatures. In these experiments, freezing was not a cause of death. 2. Survival rates at a range of constant low temperatures (– 5 to + 1 °C) and for different exposure periods (1–14 days) were measured in the summer acclimated springtail Orchesella cincta. 3. Daily interruptions of the cold exposure with short intervals at high temperature reduced mortality or slowed the increase of mortality. This effect was stronger at higher temperature (19 vs 5 and 12 °C) and increased with the duration of the interruption (0·25–2 h). 4. The injury was reversible when the cold exposure was limited to 2 days. 5. Survival in desiccated animals (14% water loss) was reduced. 6. It is suggested that the mortality of summer acclimated springtails is caused by a complex metabolic disorder and membrane changes at low temperatures.

Cold hardiness in the collembolan Orchesella cincta under different food conditions

The effect of dietary nitrogen on the ability of the collembolan Orchesella cincta to develop cold tolerance was studied. Animals fed for 6 weeks under summer conditions on the fungus Cladosporium cladosporioides containing 2.3% nitrogen.had a higher lipid and lower protein concentration than those fed on the same fungus containing 4.3% nitrogen. After 6 weeks, the animals were placed under winter conditions, and their cold tolerance and hemolymph osmolality was tested weekly. No effect of the food treatment was found for these parameters. Because starvation did not increase the cold tolerance of the animals, elimination of ice nucleators in the gut is not thought to be a mechanism for increasing cold tolerance in this species. Winter acclimation increased survival of exposure to cold; the animals were able to survive an exposure to −3°C for 1 week after 2 weeks acclimation at winter conditions. Hemolymph osmolality did not change with the increased cold tolerance of the animals. It is concluded that O. cincta does not use osmotically active cryoprotectants to develop cold tolerance. The animals may use antifreeze proteins to increase cold tolerance.

Effects of starvation on body composition and cold tolerance in the collembolan Orchesella cincta and the isopod Porcellio scaber

The effects of long-term starvation on the body composition of the isopod Porcellio scaber (Latreille) and the collembolan Orchesella cincta (L.) were studied, by determining the body composition in starved and fed animals. A period under summer conditions (19°C, 75% RH and L/D 16/8 photoperiod), was followed by a period under winter conditions (5°C, 75% RH and LD 6/18 photoperiod). O. cincta was held under summer conditions for 3 weeks, during which its protein and lipid content decreased, while its water content increased. In P. scaber, the same occurred during the 6 weeks they were kept under summer conditions. During subsequent weeks under winter conditions, changes in cold tolerance of the animals were investigated. Cold tolerance and haemolymph osmolality were measured once a week. Starved animals had lower cold tolerance than fed ones. For P. scaber a decreased haemolymph osmolality was found in starved animals compared to fed ones. This is assumed to be caused by a combination of the consumption of carbohydrates out of the haemolymph and of protein reserves and the accumulation of body water. O. cincta appeared to be capable of osmoregulation, as haemolymph osmolality did not differ between starved and fed animals, despite differences in body water content. Decreased cold tolerance in starved animals of both species may be caused by increased water content or, more probably, by the decrease in reserves needed to produce cryoprotective substances.

Effects of food quality on growth and body composition of the collembolan Orchesella cincta

Abstract. The effects of food quality on growth and body composition of the collembolan Orchesella cincta (L.) (Entomobryidae) were investigated to determine optimal values for dietary nitrogen.Hyphae of Cladosporium cladosporioides containing 2.2%, 2.7%, 4.0% and 6.2% nitrogen were used as food sources.These different nitrogen contents were obtained by adjusting the medium on which the fungi were grown.Collembola were fed on one of these four food sources for 8 weeks. O.cincta grew faster and reached a higher asymptotic body mass when fed on fungi with 4% or 6.2% nitrogen content: this effect decreased with higher nitrogen content.The lipid concentration in the animals was negatively correlated with dietary nitrogen content, whereas protein concentration was positively correlated.Glycogen concentration was not found to correlate with the composition of the food.A mechanism is proposed by which O.cincta compensates for low dietary nitrogen by consuming more to maintain a relatively constant amount of protein in its body and, as a result, accumulates more fat than animals fed on fungi with a higher nitrogen content.

Heavy-metal adaptation in terrestrial invertebrates: A review of occurrence, genetics, physiology and ecological consequences

Abstract 1. The occurrence of genetic adaptation to heavy metals in natural populations of terrestrial invertebrates is evaluated from literature data. Five criteria for adaptation evidence are applied, with concepts from ecotoxicology, ecology, life-history theory and quantitative genetics. 2. There is strong evidence for the occurrence of adaptation in natural populations of the isopod Porcellio scaber (Isopoda), the springtails Isotoma notabilis , Onychiurus armatus and Orchesella cincta (Collembola), the blowfly Lucilia cuprina and the fruit fly Drosophila melanogaster (Diptera). Adaptation to metal-containing pesticides has been demonstrated in ticks (Acarina). Population divergence indicates acclimation or adaptation in many other species. 3. Metal adaptation has been achieved within a few generations under laboratory conditions in some species; adapted populations occur at field sites that have been polluted for decades, or longer. 4. Genetic variation for tolerance and life-history characteristics, allowing for adaptation, was quantified in a reference population of Orchesella cincta . Tolerance and life-history patterns in exposed field populations matched predictions from genetic variation. 5. Adaptation involves modification and intensification of existing physiological mechanisms for metal assimilation, excretion, immobilization or compartmentalized storage. There are indications ofinter-population divergence in metal-binding proteins in a snail. In the fruit fly Drosophila melanogaster metal adaptation is achieved by duplication of the metallothionein gene. 6. An altered life-history is often part of the complex adaptation syndrome. Metal-adapted invertebrates have a shorter life-cycle and a higher reproductive effort. 7. Possible consequences of adaptation, consisting of costs of tolerance determined by genetic correlations, and probably of reduced genetic variation for tolerance and other features, are discussed. Reduced genetic variation is suggested by results for the springtail Orchesella cincta . 8. The distinction between “costs of tolerance” on the one hand and linkage disequilibrium or direct selection for altered life-history patterns on the other hand is discussed. 9. Species with high sensitivity (i.e. a low NOEC), that do not have populations maintaining sufficient genetic variation to evolve tolerance or modified life-history characteristics, or that have costly tolerance mechanisms, or both, are most at risk for extinction at sites with increasing metal pollution. 10. Metal adaptation in terrestrial invertebrates appears to be of degree rather than of kind: indications for a specific metal-fauna, equivalent to metal-vegetation, are lacking.

Genetic Variation and Covariation for Characteristics Associated with Cadmium Tolerance in Natural Populations of the Springtail Orchesella cincta (L.)

Genetic Variation and Covariation for Characteristics Associated with Cadmium Tolerance in Natural Populations of the Springtail Orchesella cincta (L.)

GENETIC VARIATION AND COVARIATION FOR CHARACTERISTICS ASSOCIATED WITH CADMIUM TOLERANCE IN NATURAL POPULATIONS OF THE SPRINGTAIL ORCHESELLA CINCTA (L.).

Heavy metals can be strong and stable directional selective agents for metal-exposed populations. Genetic variation for the metal-tolerance characteristic "cadmium excretion efficiency" was studied in populations of the collembolan Orchesella cincta from a reference- and a metal-contaminated forest soil. Previously it has been shown that "excretion efficiency" influences tolerance through midgut-mediated immobilization and excretion of toxic metal ions, and that an increased mean excretion efficiency is present in animals inhabiting metal-contaminated litter. In the present research, offspring-parent regressions showed that additive genetic variation for cadmium excretion efficiency was present in the population from the reference site. The heritability estimate was 0.33. In the natural population exposed to heavy metals from an industrial source, additive genetic variation was not significantly different from zero. Differences in the heritability between the reference and the exposed population were not significant. Genetic variation for cadmium excretion efficiency allows for a response to selection in the reference population. Such a response has probably occurred in the metal-exposed population. Half-sib analysis with animals from the reference population was used to estimate genetic variation and maternal effects for excretion efficiency, relative growth rate and molting frequency, and to determine genetic correlations between these characteristics. Additive genetic variation was demonstrated for all three characteristics, heritability estimates were 0.48, 0.75 and 0.46, respectively. Maternal effects were low for excretion efficiency and molting frequency, but may be present for relative growth rate. Phenotypic and genetic correlations among these characteristics were positive. The environmental correlation between relative growth rate and molting frequency was positive, others were negative. Direct selection for any of the characteristics, or genetic correlations between tolerance characteristics and growth characteristics, or both may have caused the responses previously observed in field populations.

Size at Maturity and Patterns of Growth and Reproduction in an Insect with Indeterminate Growth

If, in animals with indeterminate growth, fecundity is size-dependent and if a trade-off exists between reproduction and growth, current reproduction affects future reproduction via growth, and natural selection may affect age at first reproduction via mortality. We examined both conditions in the bivoltine springtail Orchesella cincta. Previous studies on this species showed mortality in the field to he high, and age at first reproduction to alternate between generations

Life cycle and food availability indices in Notiophilus biguttatus (Coleoptera, Carabidae)

Abstract. Food demands of the predatory carabid Notiophilus biguttatus F. and their fulfilment were studied in the laboratory and field. In the laboratory, larval consumption, growth, adult body size and egg production were established for different regimes of temperature and food, the springtail Orchesella cincta. Temperature strongly influenced feeding rate of larvae and adults and consequently growth and oviposition rates. Accordingly, growth rate and ovi‐position rate increased with prey supply. Prey supply also had an effect on fat content, but the number of ripe eggs in the ovaries was related neither to temperature nor to prey supply. Adult body size was affected by temperature during the larval period but more so by food supply. Dissection of field fresh females showed them to bear eggs for the whole year, except in late winter/early spring and in July. Fat content was low in spring and, from July on, high in summer and autumn. Survivorship during starvation differed widely between samples and seemed to be related to reproductive status rather than to fat content. Samples of beetles and springtails from eleven pine plantations revealed for the beetles significant differences in fat content and body size, the latter ranging between the maximum and minimum value obtained in the laboratory. Conclusions about food limitation based on body size and fat content were not corroborated by a relationship of these indices with springtail density.