Slug Arion Ater Research Articles

Genital morphology differentiates three subspecies of the terrestrial slug Arion ater (Linnæus, 1758) s.l. and reveals a continuum of intermediates with the invasive A. vulgaris Moquin-Tandon, 1855

1. Adomaitis M., Skujienė G. 2016. Comparative analysis of ligulas of slugs Arion lusitanicus from Lithuania and Denmark. Scientific Bulletin of the Uzhgorod University (Series Biology) 40: 5–10. Google Scholar

Introgression and Differentiation of the Invasive SlugArion vulgarisfrom NativeA. ater

ABSTRACT The large arionid slug Arion vulgaris is an invasive pest dispersing through large parts of Europe and causing considerable damage in gardens, horticulture and agriculture. It is also possible that this so-called “Iberian slug” has an impact on Norwegian ecosystems, displacing or hybridizing with the native black slug Arion ater. The taxonomy of the large arionids is complex and confusing, encompassing different anatomical forms and colour varieties. The present study integrates, for the first time, coloration, ligula morphology, genital morphometry, nuclear and mitochondrial DNA in order to differentiate the large arionids found in Norway, A. vulgaris, A. ater and A. rufus. These data revealed a clear separation between A. vulgaris and A. ater based on the morphology of the genitalia and mtDNA. However, introgression with the red slug A. rufus was apparent in approximately half of the A. ater specimens analysed, evidenced by ligula morphology, mitochondrial and nuclear DNA. In addition, the mito...

Characterization of cellulolytic activity in the gut of the terrestrial land slug Arion ater: Biochemical identification of targets for intensive study.

The level of cellulolytic activity in different areas of the gut of the terrestrial slug Arion ater was assayed at different temperatures and pH values. To do this, crude gut proteins were isolated and assayed using modified dinitrosalicylic acid reducing sugar assay. Crude protein samples were also separated and cellulolytic activity identified using in gel CMC zymography and esculin hydrate activity gel assays. pH and temperature profiling revealed optimum cellulolytic activity between pH5.0 and 6.0 for different gut regions and retention of up to 90% of activity at temperatures up to 50°C. Zymograms and activity gels revealed multiple endoglucanase and β-glucosidase enzymes. To further investigate the source of this cellulolytic activity bacterial isolates from the gut were tested for endoglucanase and β-glucosidase activity using growth plate assays. 12 cellulolytic microbes were identified using 16S rDNA gene sequencing. These include members of the genera Buttiauxella, Enterobacter, Citrobacter, Serratia and Klebsiella. Gut metagenomic DNA was then subjected to PCR, targeting a 400bp region of the 16SrDNA gene which was subsequently separated and individuals identified using DGGE. This identified members of the genera Citrobacter, Serratia, Pectobacterium, Acinetobacter, Mycoplasma, Pantoea and Erwinia. In summary, multiple glycoside hydrolase enzymes active over a broad range of temperature and pH values in a relatively under studied organism were detected, indicating that the gut of A. ater is a viable target for intensive study to identify novel carbohydrate active enzymes that may be used in the biofuel industry.

Simulation of moderated, severe metal pollution and remediation scenarios using ex situ transplant experiments with real soils: Effects on mortality, growth, and cell and tissue-level biomarkers in slugs Arion ater

Simulation of moderated, severe metal pollution and remediation scenarios using ex situ transplant experiments with real soils: Effects on mortality, growth, and cell and tissue-level biomarkers in slugs Arion ater

Applicability of the black slug Arion ater for monitoring exposure to polycyclic aromatic hydrocarbons and their subsequent bioactivation into DNA binding metabolites

The applicability of terrestrial black slugs Arion ater (Mollusca, Gastropoda) was studied for biomonitoring environmental exposure to polycyclic aromatic hydrocarbons (PAHs). In laboratory experiments, slugs were orally exposed to benzo[a]pyrene (BaP) for a short term (3 days) or a long term (119 days) period. Test animals were collected in the field, or were reared under laboratory conditions to ensure that they had no history of PAH-exposure. Benzo[a]pyrene hydroxylase (BPH) activity was measured in the digestive gland as a biomarker for BaP exposure. Bulky DNA adduct formation in kidney was measured as an effect biomarker for BaP bioactivation into DNA-binding metabolites. Although success of clutching was relatively low (5 out of 18 slugs produced egg packages), sufficient number of slugs were obtained to perform exposure experiments due to high hatching (89%) and survival rates (79%). After a short exposure to a relatively high BaP doses of 20 and 200 μg/g fresh feed, a dose-dependent and significant increase of BPH activity and bulky DNA adduct levels could be demonstrated in A. ater. Induction factors were low (two times control level), but optimization of the test conditions yielded a higher BPH induction factor of 4.8 times control level. BPH activity and bulky DNA adduct levels, however, did not increase after a long-term exposure to environmentally relevant BaP doses (upto 0.25 μg/g fresh feed). Based on this lack of response after realistic exposure it is concluded that A. ater is not sensitive to BaP exposure and, therefore, not suitable for monitoring environmental exposure to PAHs.

Preference for Acyanogenic White Clover (Trifolium repens) in the Vole Arvicola terrestris: I. Experiments with Two Varieties

We report experimental results showing that, under both laboratory conditions as well as in outdoor enclosures, the fossorial vole Arvicola terrestris preferentially feeds on acyanogenic white clover (Trifolium repens) when offered the choice between two varieties (Ladino and Aran) differing highly in their content in cyanogenic glycosides. We also observed that the voles adapted their diet and reduced their relative consumption of the cyanogenic variety during experiments conducted for two to three weeks in outdoor enclosures as compared to shorter tests conducted for 48 hr in laboratory cages. In addition, we report a similar preference for the acyanogenic Ladino variety for the slugs Arion ater and A. subfuscus.

The association of mannitol oxidase with a distinct organelle in the digestive gland of the terrestrial slugArion ater

Post nuclear supernatants prepared from homogenates of the digestive gland of the slugArion ater were analysed by rate-dependent and density-dependent density gradient zonal centrifugation. Mannitol oxidase carrying structures exhibited distinct centrifugal behaviour sedimenting more slowly than mitochondria, lysosomes and peroxisomes but faster than microsomes, and banding sharply at a density of 1.15 g/ml in sucrose gradients. By combining rate and isopycnic centrifugations mannitol oxidase carrying structures were largely separated from contaminating organelles. Electron microscopic examination of such mannitol oxidase-enriched fractions revealed the predominant presence of distinct structures of tubular form. SDS PAGE analysis indicated that the major polypeptide present had a mass of ∼ 68 kDa corresponding to the major subunit previously reported for partially purified mannitol oxidase ofHelix aspersa.

Effects of adenine nucleosides and nucleotides on the isolated heart of the snail Helix aspersa and the slug Arion ater

1. Adenine nucleosides and nucleotides were examined for pharmacological activity in hearts isolated from the snail Helix aspersa and the slug Arion ater. 2. Adenosine, AMP, ADP and ATP (above 100 μM) produced either an excitation or an inhibition in the isolated hearts of the snail and slug. 3. 2-Chloroadenosine, α, β -methylene ATP and 2-methylthio ATP were inactive at concentrations up to 1 mM. 4. Responses were not blocked by any commonly accepted vertebrate purinoceptor antagonists, indicating that these purinoceptors are dissimilar to vertebrate purinoceptors and cannot be classified according to accepted purinoceptor classifications. 5. Electrical field stimulation of the snail heart produced frequency-dependent responses: 1–4 Hz produced predominantly excitation, 8–32 Hz predominantly inhibition. These responses were unaffected by the purines up 3 mM.

Preferential Grazing by Molluscs of Plants Infected by Rust Fungi

In the field infection by pathogenic rust fungi was observed to increase invertebrate herbivory of a number of plant species. This phenomenon was investigated in detail in Allium ursinum L. infected by Puccinia sessilis Schroter and Tussilago farfara L. infected by Puccinia poarum Niels and Coleosporium tussilaginis (Pers.) Berk. In the field, 57% of P. sessilis pustules on A. ursinum were grazed and the area consumed by herbivores was significantly greater in rusted than control leaves in nine out of the ten host populations studied. Examination of herbivores associated with rusted A. ursinum suggested that much of the observed grazing could be attributed to several species of mollusc. In a controlled preference test the slug Arion ater L. consumed 6-23 times more rusted tissue than controls. Aeciospores of P. sessilis were present in the faeces of 44 ? 4% of the snail Trichia striolata Pfeiff. 48% of P. poarum-infected T. farfara leaves showed some evidence of selective grazing. Such leaves had 29.25 ? 3.52 rust pustules of which 4.57 ? 0.66 (ca. 17%) were grazed. The slugs Arion hortensis Fer. and A. fasciatus Nilss. consumed signif

Aromatic alcohol oxidase: A new membrane‐bound H2O2‐generating enzyme in alimentary tissues of the slug Arion ater

AbstractWe employed a sensitive spectrophotometric technique to detect H2O2‐generating enzymic activities capable of utilizing aldehydes or alcohols as substrates. We analysed tissue homogenates and subcellular fractions of the terrestrial slug Arion ater. Of the tissues tested, those of the alimentary system (especially crop and digestive gland) had the greatest activity and generated H2O2 when supplied with a variety of alcohol and aldehyde substrates. Aldehyde oxidase activity was soluble and had high apparent affinity for benzaldehyde. This activity is probably analogous to mammalian aldox (aldehyde oxidase; E.C.1.2.3.1), since it could use an artificial electron acceptor system and greatly preferred benzaldehyde to hypoxanthine. However, it differed from guinea pig liver aldehyde oxidase in that salicylaldehyde was a potent inhibitor rather than a substrate. Alcohol oxidase activity was particulate but not localised in mitochondria or peroxisomes. In digestive gland, it was localised differently from mannitol oxidase. In crop, the subcellular sites are either the same or inseparable by isopycnic density gradient centrifugation in sucrose gradients. Activity was at least ten times greater with aromatic alcohols than with ethanol, propanol, or butanol. No activity was detected with methanol, and the enzyme could not utilise an artificial electron acceptor system or pyridine nucleotides. Aromatic alcohol oxidase differed from mannitol oxidase with respect to temperature and pH sensitivity and in exhibiting greater apparent affinity for its substrates. The alcohol oxidase accepted a wide variety of aromatic alcohols as substrates. In several respects it resembled the corresponding enzyme of wood‐rot fungi, implying that the slug utilises dietary lignin or lignin precursors. Possible dietary, microbicidal, and detoxifying roles are considered for slug aldehyde and alcohol oxidases.

The involvement of opioid peptides in stress-induced analgesia in the slug Arion ater

Application of tail-pinch stress to the terrestrial slug, Arion ater, produced a significant increase in the response time when tested on the hot-plate for foot-lifting response. The analgesia was completely reversed by injections of the opiate antagonists, naltrexone and ICI 174864, in a dose-dependent manner. Analgesia could also be elicited by the injection into the foot of beta-endorphin and the enkephalin analogues, DAGO and DADLE. No effect was seen with dynorphin A (1-8) or dynorphin A (1-17). The stress-induced analgesia disappeared after 30 minutes but could be maintained for 100 min following the injection of a mixture of bestatin and the enkephalinase inhibitor, N-carboxymethyl-L-phenylalanyl-L-leucine. This work suggests that in the slug, a physical stressor produces an analgesia which may be due to the release of endogenous opiates.

Zinc treatment of the digestive gland of the slug Arion ater L. 2. Sublethal effects at the histological level.

Several authors have pointed out the necessity when measuring the level of pollution within a biological system on an assessment of the biological effects of pollutants. Within this field, histological parameters for the indication of environmentally induced stress response (i.e., destabilization of the lysosomal systems, reduction of the mean epithelia thickness of digestive gland cells-MET) could offer an initial approach to determine the impact of a pollutant on the biota. The present work is the completion of the histochemical study on cellular distribution of zinc in the digestive gland of the slug Arion ater. Thus, the histochemical and planimetric study of the effects of zinc-exposure on A. ater is presented as the other aspect in which histology is concerned in pollution research.

Zinc treatment of the digestive gland of the slug Arion ater L. 1. Cellular distribution of zinc and calcium.

Extensive literature has been published on the molluscan capability for metal bioaccumulation. As a consequence, a variety of species have been proposed for environmental pollution biomonitoring. The knowledge of biological mechanisms of accumulation and elimination of environmental pollutants is essential to get a proper use of bioindicator species. Histological procedures may indicate (a) the events involved in the metabolic regulation of pollutant bioavailability and release and (b) their specific toxic effects. The relevance of the digestive gland as a major site for Zn accumulation has been reported. The Zn distribution in this organ might indicate the level of environmental bioavailable Zn. The present study is a histochemical investigation of the distribution and release of zinc by the different cell types of the digestive gland of A. ater.

A comparative study of the uptake and distribution of silver in a slug Arion ater and a snail Achatina fulica

1. 1. Studies over 10 weeks showed Achatina accumulated more silver than Arion. 2. 2. This response was not due to the presence of a shell in Achatina. 3. 3. The digestive gland was the main site of silver deposition. 4. 4. Faecal silver was higher in Arion than Achatina. 5. 5. Silver uptake appeared to cause an increase in magnesium but a decrease in calcium and zinc in the digestive gland of Achatina. 6. 6. The location of the silver in the digestive gland was associated with calcium granules in the cell.

Assay and subcellular localization of H2O2 generating mannitol oxidase in the terrestrial slug Arion ater

AbstractAn assay is described that allows the measurement of H2O2‐generating mannitol oxidase activity in sucrose‐containing homogenates and subcellular fractions. Using this assay high activity was found in digestive gland and crop of the slug Arion ater. The subcellular localization of mannitol oxidase in these tissues was examined using sucrose density gradient centrifugation in a B‐XIV zonal rotor. Mannitol oxidase was found to reside in a lowdensity membrane tentatively identified as golgi or plasma membrane, rather than in mitochondria or peroxisomes as had previously been supposed.

Copper treatment of the digestive gland of the slug Arion ater L. 2. Morphometrics and histophysiology.

In order to have a clear image of processes involved in copper bioaccumulation-detoxication mechanisms in the terrestrial slug, Arion ater, the authors planned a histophysiological analysis based on morphometrics and on the variations in morphological characteristics of epithelia. This study is the completion of the histochemical analysis, in which the authors suggest a close relationship between copper and calcium in the digestive gland of Arion ater. They had observed that copper was exclusively stored within calcium cells and that calcium cells secretion was more intensive at higher copper dosages and at larger bioassay times. Therefore, histophysiological analysis was necessary for explaining the nature of copper bioaccumulation-detoxication mechanisms.

FEEDING AND GROWTH RESPONSES TO COPPER, ZINC, MERCURY AND LEAD IN THE TERRESTRIAL GASTROPOD ARIONATER (LINNE)

Food consumption and growth variations related to several dosages of heavy metals (Cu, Zn, Hg and Pb) in diet were studied in the terrestrial slug Arion ater for 27 days of experimental treatment. The work reported here is part of a larger project to investigate the histological effects of metal exposure in pulmonate gastropods. Graphics on food consumption and growth vs. time for each metal, and regression for each behaviour observed are presented.

Seasonal changes in zinc, manganese, magnesium, copper and calcium content in the digestive gland of the slug Arion ater L.

1. 1. Except for zinc there was an increase in the total metal content of the digestive gland from May to October. This was due essentially to an increase in the weight of the gland. 2. 2. The drop in total metal content during November was probably the result of a decrease in the size of the digestive gland. 3. 3. Except for copper in May and calcium in November there was an increase in the metal concentration from May to November. In some cases it was not significant. 4. 4. Variations in the metal content are discussed in relation to seasonal uptake and age.

Alternative risk‐taking styles: The case of time‐budgeting strategies of terrestrial gastropods

The terrestrial slugs Arion ater, Limax maximus and Ariolimax columbianus have similar morphological designs but differ remarkably in their life history tactics and behavioural time budgets. The adaptive value of particular risk-taking styles was investigated using a comprehensive computer simulation model. The model allowed each species' success (growth rates, food acquisition) and costs (distance travelled, hydration deficits, injury from aggressive encounters) to be evaluated in various types of weather (benign, harsh and surprise). This was interpreted in terms of the species' life history design. In addition, each species was simulated with the behavioural strategy of the other two species substituted for its natural programming. The simulation experiments demonstrated how variations in a few simple rules could lead to divergent cost-benefit consequences. The model also illustrated that coarsely-tuned sensitivity to the environment may be better than finely-tuned responses, depending on the animal's ability to respond quickly and the degree of risk. The simulations suggested that competition has shaped the time-budgeting tactics of A. ater and L. maximus. Each species actually performed better in harsh weather, using the behavioural program of the other. L. maximus is an aggressive species with a narrow, nocturnal activity period. The concentrated activity of L. maximus may allow it to displace competitors more effectively, whereas the broader time span of A. ater's activity may be necessary to avoid L. maximus.

Sites of water, zinc and calcium uptake and distribution of these metals after cadmium administration in Arion ater (Gastropoda: Pulmonata)

1. 1. Water and zinc were taken up parentally in the slug Arion ater. 2. 2. Using the everted sac technique, radioactive calcium was not actively transported across the intestine of A. ater. 3. 3. Metal accumulation was studied in the foot, intestine and digestive gland. 4. 4. Cadmium accumulated in the digestive gland and was proportional to the degree of environmental contamination. There was a significant decrease in the concentration of calcium in the intestinal tissue after cadmium treatment. 5. 5. Cadmium in the intestinal tissue was associated with a protein eluted from a Sephadex column in the same place as the cadmium-binding protein in the digestive gland.