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Several studies have reported the high bioindication capacity of Isopoda (Crustacea, Oniscidea), which is related to their important ability to accumulate contaminants, usefulness in soil ecotoxicology and bioindication activities. Any change in the isopod population, diversity and life cycle can indicate relevant pollution levels. The analysis of target tissues, such as the hepatopancreas, is another emerging approach (from a cytologic/histological level) to detect contaminant accumulation from different sources. In this study, tissue disaggregation procedures were optimised in the hepatopancreas, and flow cytometry (FC) was applied to detect cell viability and several cell functions. After disaggregation, two hepatopancreatic cell types, small (S) and big (B), were still recognisable: they differed in morphology and behaviour. The analyses were conducted for the first time on isopods from sites under different conditions of ecological disturbance through cytometric re-interpretation of ecological-environmental parameters. Significant differences in cell functional parameters were found, highlighting that isopod hepatopancreatic cells can be efficiently analysed by FC and represent standardisable, early biological indicators, tracing environmental-induced stress through cytologic/histologic analyses., (© 2024. The Author(s).)

Wolbachia-induced feminisation is an extended phenotype that has been observed in only three models: isopods, butterflies and leafhoppers. Even among these, the process of feminisation differs markedly, especially since sexual differentiation is coordinated body-wide by a hormone in isopods, whereas in insects it is a chromosomally determined process and enforced cell by cell. For isopods, Armadillidium vulgare has contributed substantially to an understanding of Wolbachia-mediated feminisation, because a wide array of know-how and methods has been tailored to this model. Here, we describe establishment and maintenance of Wolbachia-infected laboratory lineages from animals collected in the field, the grafting of an androgenic gland to demonstrate the mode of action of Wolbachia (i.e. the disruption of the Androgenic Hormone pathway), and transinfection of naïve adults with Wolbachia. Finally, we describe selection of a ZZ genetic background in lineages created from transinfected WZ females, which is necessary to benefit from a property intrinsic to the naturally infected lineages: In a ZZ background, the vertical transmission rate of Wolbachia directly equates to the female ratio in the progeny. These protocols provide the essential framework for investigating molecular aspects of Wolbachia-induced feminisation., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

The order Isopoda contains both aquatic and terrestrial species, among which Hemilepistus reaumurii, which lives in arid environments and is the most adapted to terrestrial life. Olfaction has been deeply investigated in insects while it has received very limited attention in other arthropods, particularly in terrestrial crustaceans. In insects, soluble proteins belonging to two main families, Odorant Binding Proteins (OBPs) and Chemosensory Proteins (CSPs), are contained in the olfactory sensillar lymph and are suggested to act as carriers of hydrophobic semiochemicals to or from membrane-bound olfactory receptors. Other protein families, namely Nieman-Pick type 2 (NPC2) and Lipocalins (LCNs) have been also reported as putative odorant carriers in insects and other arthropod clades. In this study, we have sequenced and analysed the transcriptomes of antennae and of the first pair of legs of H. reaumurii focusing on soluble olfactory proteins. Interestingly, we have found 13 genes encoding CSPs, whose sequences differ from those of the other arthropod clades, including non-isopod crustaceans, for the presence of two additional cysteine residues, besides the four conserved ones. Binding assays on two of these proteins showed strong affinities for fatty acids and long-chain unsaturated esters and aldehydes, putative semiochemicals for this species., Competing Interests: Conflicts of interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Circadian clocks in terrestrial animals are encoded by molecular feedback loops involving the negative regulators PERIOD, TIMELESS or CRYPTOCHROME2 and positive transcription factors CLOCK and BMAL1/CYCLE. The molecular basis of circatidal (~12.4 hour) or other lunar-mediated cycles (~15 day, ~29 day), widely expressed in coastal organisms, is unknown. Disrupting circadian clockworks does not appear to affect lunar-based rhythms in several organisms that inhabit the shoreline suggesting a molecular independence of the two cycles. Nevertheless, pharmacological inhibition of casein kinase 1 (CK1) that targets PERIOD stability in mammals and flies, affects both circadian and circatidal phenotypes in Eurydice pulchra (Ep), the speckled sea-louse. Here we show that these drug inhibitors of CK1 also affect the phosphorylation of EpCLK and EpBMAL1 and disrupt EpCLK-BMAL1-mediated transcription in Drosophila S2 cells, revealing a potential link between these two positive circadian regulators and circatidal behaviour. We therefore performed dsRNAi knockdown of Epbmal1 as well as the major negative regulator in Eurydice, Epcry2 in animals taken from the wild. Epcry2 and Epbmal1 knockdown disrupted Eurydice's circadian phenotypes of chromatophore dispersion, tim mRNA cycling and the circadian modulation of circatidal swimming, as expected. However, circatidal behaviour was particularly sensitive to Epbmal1 knockdown with consistent effects on the power, amplitude and rhythmicity of the circatidal swimming cycle. Thus, three Eurydice negative circadian regulators, EpCRY2, in addition to EpPER and EpTIM (from a previous study), do not appear to be required for the expression of robust circatidal behaviour, in contrast to the positive regulator EpBMAL1. We suggest a neurogenetic model whereby the positive circadian regulators EpBMAL1-CLK are shared between circadian and circatidal mechanisms in Eurydice but circatidal rhythms require a novel, as yet unknown negative regulator., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Lin et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The terrestrial crustacean Porcellio scaber is an established test organism in environmental research. We analysed the haemolymph proteome of P. scaber using a classical proteomic approach based on one-dimensional gel electrophoresis and tandem mass spectrometry. Using a publicly available protein database and our P. scaber transcriptome data, we have identified 76 proteins involved in cytoskeleton formation, protein degradation, vesicular transport, genetic information processing, detoxification, carbohydrate and lipid metabolism reflecting haemocyte metabolic activity, active intracellular transport, and intercellular communication. Compared with the data reported for other crustaceans, 28 of these P. scaber proteins have been linked to its immunity, among them hemocyanin, α-2-macroglobulin, phenoloxidase 3, superoxide dismutase, glutathione S-transferase, haemolymph clottable protein, and histones H4 and H2B. Our results thus provide a firm base for studying the innate immune response of P. scaber at the level of the haemolymph proteome. This knowledge is of particular importance in ecotoxicity studies with various environmental stressors where understanding physiological changes is important to reveal possible modes of action., Competing Interests: Declaration of competing interest Authors have no competing interests to declare., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

The use of degradable chelating agents to facilitate phytoextraction is a promising low-cost method for the remediation of heavy metal-contaminated soils. However, there are few studies on how plants and soils respond to the chelating agents. In this study, the responses of French marigold (Tagetes patula L.) and soil cadmium (Cd) to the chelator tetrasodium glutamate (GLDA) was investigated in a 180 d field trial. Five GLDA treatments (0, 292.5, 585, 1170, and 2340 kg hm -2 ) were carried out in a Cd-contaminated soil (0.47 mg kg -1 ) under French marigold plantation. The results showed that the application of GLDA promoted the transformation of other forms of Cd in soil to exchangeable state, and the exchangeable Cd and Fe-Mn oxide bound state increased by 42.13% and 32.97% (p < 0.05), respectively. The cell wall Cd accumulations significantly increased 9.39% (p < 0.05) and the percentages of soluble fractions increased by 460.33% (p < 0.05). Furthermore, increases occurred in soil pH, as well as DOC and DTPA-Cd contents with increasing the total amount of GLDA. The composite application of GLDA (2340 kg hm -2 ) with French marigold reduced the total soil Cd content by 7.59% compared with the soil background. Altogether, results of this study suggested that the application of GLDA can effectively activate soil Cd and enhance the capability of French marigold for the remediation of Cd-contaminated soils., (© 2022. The Author(s).)

The semiterrestrial isopod, Ligia exotica represents one of the oldest documented species introductions of marine organisms and is known as an intermediate form between marine and strictly terrestrial isopods. In order to explore the potential value of Ligia as an animal food source, this study focused on the growth rate under laboratory rearing conditions and conducted a detailed analysis of the overall nutrient content of the species in comparison to two other marine food media (krill and fish meal). Evaluation of the growth rate of juveniles suggests it is a relatively fast-growing species of the Ligiidae family. The essential amino acids content Ligia meal is the lowest amongst the three studied media but the proportion of flavor amino acids, and in particular taurine, was higher. The most restricted amino acids of isopod meal are methionine and cysteine. The significantly unbalanced amino acid composition of Ligia meal may affect the absorption and utilization by consumers. In terms of fatty acids, the total polyunsaturated fatty acids in the isopod is very low. A total of 12 vitamins were examined. The VK 1, VE, VB 2 , VB 3 , VB 5 content of isopod meal were significantly higher than those of krill meal and fish meal. Similarly, most of the 11 mineral elements are highest in the isopod meal. Ligia therefore offers potential as an alternative natural food source in animal given the growth rate under culture and the overall nutrient content. But Ligia collected in most of the field would be deemed unfit for human consumption because of the relatively low nutritional value and heavy metal content exceeding the provided standard. Further study is warranted to elucidate the biological characteristics of isopods and how its diet is reflected in its nutritional value to consumers.

Like in most Crustacea, the cuticle of terrestrial isopods is hardened by a calcareous mineral phase. This rigid cuticle is frequently shed during a process called moulting. To reduce calcium loss, Porcellio scaber eats the shed cuticle, the exuviae, and absorb the calcium from it through large tubular diverticula of the intestine, called the mid gut glands or hepatopancreas. After moulting the absorbed calcium should be transported immediately into the hemolymph from which it is used to rapidly mineralize the new cuticle. This suggests that the hepatopancreas epithelium transports calcium from the lumen to the hemolymph. We used TEM, energy-filtered TEM and electron-probe X-ray microanalysis to analyse the distribution of elevated calcium within the hepatopancreas cells of P. scaber. We used animals in the postmoult stage that have eaten their exuviae and, as a control, those that have not ingested the exuviae. To minimize calcium loss within the samples, we used high pressure frozen and freeze substituted samples and propane-1-3-diol as floatation medium for thin-sectioning. The results reveal intracellular dense deposits containing calcium, phosphorus and oxygen at the apical microvillus membrane, within the cytoplasm, attached to vesicles and to the basolateral membrane, as well as extracellular between cells and the basal lamina. Control animals were devoid of these deposits. The results indicate that calcium from the exuviae is absorbed and transported across the epithelium into the hemolymph. We propose that during transport, intracellular calcium is bound to phosphate avoiding toxic effects of high concentrations of ionized calcium., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Using the isopod Armadillidium vulgare as a case study, we review the significance of the "bacterial dosage model", which connects the expression of the extended phenotype to the rise of the Wolbachia load. In isopods, the Insulin-like Androgenic Gland hormone (IAG) induces male differentiation: Wolbachia feminizes males through insulin resistance, presumably through defunct insulin receptors. This should prevent an autocrine development of the androgenic glands so that females differentiate instead: feminization should translate as IAG silencing and increased Wolbachia load in the same developmental window. In line with the autocrine model, uninfected males expressed IAG from the first larval stage on, long before the androgenic gland primordia begin to differentiate, and exponentially throughout development. In contrast in infected males, expression fully stopped at stage 4 (juvenile), when male differentiation begins. This co-occurred with the only significant rise in the Wolbachia load throughout the life-stages. Concurrently, the raw expression of the bacterial Secretion Systems co-increased, but they were not over-expressed relative to the number of bacteria. The isopod model leads to formulate the "bacterial dosage model" throughout extended phenotypes as the conjunction between bacterial load as the mode of action, timing of multiplication (pre/post-zygotic), and site of action (soma vs. germen).

Woodlice and the earthworm alone or in combination were used to improve physical properties, nutrient release, and heavy metals stabilization during composting of sewage sludge. Chemical properties of raw sludge (IS) were compared to those of composted sludge (CS), sludge + earthworms (VS), sludge + woodlice (WS), and sludge + earthworms + woodlice (VWS) after 50 and 100 days of composting. Physical properties and heavy metals accumulation by the studied fauna was determined after 100 days of composting. Highest proportions of fine particles, porosity and water holding capacity were in the VWS treatment, electrical conductivity, ash content, inorganic N and the total concentrations of P, K, Ca and Mg increased, whereas the pH value, the organic C, C:N ratio and humic acids content decreased in following order: VWS > VC > WS > CS > IS. Total concentrations of Pb, Cd and Ni in composted sludge were lower whereas concentrations of Zn, Cu and Mn were higher than in raw sludge, and woodlice showed higher bioconcentration factor (BCFs) than earthworm for all heavy metals in all treatments. We concluded that woodlice and earthworms synergistically improved the physicochemical properties of the compost and enhanced its potential use as amendment in agricultural soils., (Copyright © 2019 Elsevier B.V. All rights reserved.)

In crustaceans, antimicrobial peptides (AMPs) are clustered into four major groups according to their amino acid composition and structure: (1) single-domain peptides containing cysteine residues such as a nti-lipopolysaccharide-factor (ALF), (2) multi-domain or chimeric AMPs such as crustins, (3) non-conventional AMPs, and (4) linear single-domain AMPs. The majority of AMPs has been described in commercially exploited crustaceans, particularly decapods living in aquatic environments (crab, shrimp, lobster, and crayfish). Here, we aimed at establishing the AMPs repertoire of terrestrial isopods (Oniscidea), an original suborder of crustaceans adapted to life outside of the aquatic environment. Using transcriptomic data from 21 species, we identified 110 ALF and 73 crustin sequences. We also characterized the full-length sequence of armadillidins from 17 species, similar to the AMP previously described in the terrestrial isopod Armadillidium vulgare . Furthermore, we tested the antimicrobial activity of three armadillidin peptides characterized from three distantly related species. This analysis revealed similar activity spectra against pathogens, despite extensive structural variation among the tested peptides. In addition to conventional crustacean AMPs, our work highlights armadillidins as a new and independent family of AMPs specific to the Oniscidea, thus opening new perspectives concerning the study of the immune system of terrestrial isopods., Competing Interests: The authors declare no conflict of interest.

The hepatopancreas of isopods has major functions in food digestion and storage of carbohydrates and lipids. Also, it stores essential and accumulates xenobiotic metals in lysosomal granules within the two major cell types, the S- and B-cells of the tissue. A µCT study on moulting Porcellio scaber has shown mineral within the hepatopancreas lumen, when the animal has ingested their shed cuticle after moulting, suggesting recycling of mineral from the exuviae. This study aims to reveal if the lysosomal metal containing granules store calcium originating from the ingested exuviae. Therefore, we investigated the effect of cuticle ingestion on the elemental composition of the hepatopancreas granules of P. scaber, using electron probe X-ray microanalysis. For the preservation of diffusible elements, samples were high pressure frozen and freeze substituted in acetone and we used Propane-1,3-diol as a floatation medium for sections. We analyzed S- and B-cells of animals in the postmoult and intermoult stage that have ingested their exuviae and, as a negative control, cells from postmoult animals that have not ingested their exuviae. STEM and TEM were used for the investigation of the ultrastructure. Unexpectedly, the cryo-fixed samples contain numerous extracellular vesicles (exosomes) and many multivesicular bodies containing pro-exosomes. We show a significant increase of calcium, copper, zinc and sulphur within the metal granules upon exuviae ingestion, and, after 9 days, a reduction of calcium and zinc. The results indicate transitory storage of calcium from the exuviae within the metal granules and its subsequent utilization in cuticle mineralization., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Parasitic isopods perforate and attach to the host integument via the mandibles and then feed on hemolymph and exudate from the wounds. Such isopods attack a variety of commercially important fish and crustacean hosts. Similar to other hematophagous parasites, isopods may also employ biomolecules that affect host blood conglutination and defense systems. In the present study, a tandem mass tag-based quantitative proteomic approach was used to identify differentially expressed proteins in Tachaea chinensis parasites of shrimp, by comparing parasitic (fed) and pre-parasitic (unfed) individuals. We identified 888 proteins from a total of 1,510 peptides, with a significant difference in 129 between the fed and unfed groups. Among these, 37 were upregulated and 92 were downregulated in unfed T. chinensis . This indicates that T. chinensis may require more energy before parasitism during its search for a host. In addition, as is the case for other blood-sucking parasites, it might secrete antihemostatic, anti-inflammatory, and immunomodulatory molecules to facilitate blood meal acquisition. To our knowledge, this study is the first to use a TMT-based proteomic approach to analyze the proteome of isopod parasites, and the results will facilitate our understanding of the molecular mechanisms of isopod parasitism on crustaceans., (Copyright © 2019 Li, Li, Han, Xu, Li and Chen.)

Historically, the European Atlantic is probably the most important oil-spill hotspot worldwide. One of the most recent accidents occurred in 2002 when the oil-tanker Prestige sank over the Galician Bank causing two major oil-spills followed by several small leaks until March 2003. This resulted in contamination of virtually all types of marine habitat. Considering that parasites have proved to be good effect and accumulation bioindicators, the present study addresses the medium-term changes in trace element content after the Prestige oil-spill in a model host-parasite system, the bogue, Boops boops (Sparidae) and the isopod Ceratothoa oestroides. To our knowledge, this study is the first to address trace element concentrations in natural fish and parasite populations associated with the effects of an oil-spill. We observed that both test organisms examined, the host and the parasite, indicate a detectable change in the relative composition of trace element concentrations before and after the Prestige oil-spill. Multivariate analyses also indicated a differential response of the different tissues to the temporal sampling sequence. However, analyses of both host and parasite tissues supported the pattern of a gradual temporal transition to a state of relative trace element content distinctly departing from the pre-spill situation. Moreover, the parasite-host element accumulation ratios better depicted this temporal pattern. Additionally, changes in V concentrations in fish liver tissues and Ni concentrations in the parasite tissues suggest that this host-parasite system may be a useful tool to assess these two element contaminations linked to heavy fuel oil-spill., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Environmental temperatures and oxygen availability are important for the balance between oxygen supply and demand. Terrestrial organisms are generally perceived to be less limited by access to oxygen than their aquatic counterparts. Nevertheless, even terrestrial environments can be deficient in oxygen, especially for organisms occurring in soil, litter, wood, rotten fruit or at high elevations. While isopods are the best adapted to a terrestrial lifestyle among crustaceans, many species, including woodlice, occupy environmental gradients of temperature and oxygen. To investigate whether mismatches between oxygen supply and demand can result in a loss of performance in a terrestrial organism, we studied the effects of atmospheric oxygen concentration on the thermal performance of the common rough woodlouse (Porcellio scaber). We compared the thermal preference, thermal sensitivity of running speed, and tolerance to extreme temperatures of woodlice exposed to one of two oxygen concentrations (21% - normoxia, 7% - hypoxia). Under hypoxia, P. scaber preferred microhabitats with temperatures that were on average 3°C lower than those preferred under normoxia. The running speed tended to reach its maximum at a lower temperature under hypoxia than under normoxia (25.13°C vs 28.87°C, respectively, although p was equal to 0.09), and normoxic woodlice ran approximately 1.5-fold faster than hypoxic woodlice at the point of maximum speed. Heat tolerance was significantly lower under hypoxia (38.9°C) than under normoxia (40.7°C), but there was no difference in cold tolerance (5.81°C under normoxia and 5.44°C under hypoxia). Overall, our results indicate that environmental gradients of temperature and oxygen may shape the physiological performance of terrestrial ectotherms, likely via their effects on the balance between oxygen supply and demand, which may have fitness consequences for these organisms in nature., Competing Interests: The authors have declared that no competing interests exist.

Isopods of the genus Ligia are deemed as excellent pollution monitoring organisms for coastal zones. A species of Ligia cinerascens was chosen to be tested, which was collected from three coastal zones in Dalian, China. The concentrations of metals followed the order of Cu>As ≈ Cd>Pb. Ligia collected from a municipal sewage outlet in Heishijiao showed the highest level of metals. Ligia from a natural bathing beach in Changshan Island showed the lowest level of metals as expected. Ligia grown in the same site could accumulate different levels of metals due to their different body weight. Both males and females conformably accumulated heavy metals in bodies. Taken together, this work suggests that L. cinerascens can serve as a biomonitor species. Within the range of 0.3-0.9 g body weights, body weight rather than gender should be taken into account as a factor in the process of environmental biomonitoring.

Woodlice are top consumers of a three-trophic system (soil, leaf litter, woodlice), and they are closely related to pollutant absorbing surfaces such as soil, leaf litter, and organic matter. We studied the effects of urbanization on trace element concentrations and fluctuating asymmetry of Armadillidium vulgare (Crustacea: Isopoda) individuals in and around Debrecen city, Hungary. Along an urbanization gradient (urban, suburban, and rural areas), trace element concentrations (Ba, Cu, Fe, Mn, Pb, and Zn) of woodlice and bioconcentration factor (BCF) were analyzed. Asymmetry was also measured in metric traits of woodlice: the length of three segments of antennae, the body length, and the width of the 3rd segment of pereon. We found significant differences in Ba and Cu concentration of A. vulgare individuals along the urbanization gradient. The highest Cu concentration was found in woodlice from the urban area and the highest Ba concentration was found in the individuals from the rural area. The Ba concentration was higher in females than in males. The BCF values of Cu indicated that A. vulgare accumulated this element from soil and leaf litter. There was no significant difference in symmetry of the bilateral traits of woodlice along the urbanization gradient based on FA levels. Our results showed that the urbanization had remarkable effect on the Ba and Cu concentration of woodlice which were originated from traffic pollution. At the same time, the anthropogenic activities did not affect the symmetry of the tested traits of A. vulgare individuals.

Perfluoroalkylated acids (PFAAs) are persistent chemicals that have been detected globally in the environment and in wildlife. Although it is known that PFAAs sorb to solid matrices, little is known on PFAA concentrations in soils. PFAA pollution has often been studied in aquatic invertebrates. However, this has rarely been done on terrestrial species. In the present study, we examined whether the concentrations of 15 PFAAs in isopods (Oniscidae), collected at a fluorochemical plant and in four other areas, representing a gradient in distance from the pollution source (1 km to 11 km), were related to those in the soil and in eggs of a songbird, the great tit (Parus major), collected in the same areas. Additionally, we examined the effect of physicochemical properties such as total organic carbon (TOC) and clay content on the relationship between the concentrations in soil and isopods. Finally, we examined the composition profile in the soil and isopods. Mean PFOS and PFOA concentrations of 1700 ng/g dw and 24 ng/g dw were detected in the soil at the plant. PFOS and PFPeA were the dominant PFAAs in isopods and were detected at mean concentrations of 253 and 108 ng/g ww, respectively. The great tit eggs showed elevated mean PFOS concentrations of 55,970 ng/g ww. In most cases, PFAA concentrations decreased with increasing distance from the plant. As PFAA concentrations in isopods were correlated with concentrations in the soils, isopods could serve as a bioindicator for PFAA concentrations in soils. Additionally, there were indications that isopods could also serve as a bioindicator for PFAA concentrations in eggs of great tits. However, these indications were only the case at two locations, showing the need to further monitor the possibilities of using isopods as a bioindicator for PFAA concentrations in song bird eggs. CAPSULE: Elevated PFAA concentrations in isopods reflected concentrations in songbird eggs and in soil, indicating that trophic transfer of PFAAs from soil, via isopods, to songbirds might play a role in the PFAA exposure of terrestrial songbirds., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The presence of human pharmaceuticals in the environment has garnered significant research attention because these compounds may exert therapeutic effects on exposed wildlife. Yet, for many compounds, there is still little research documenting their stability in the water column and uptake in organism tissues. Here, we measured the uptake and stability of methylphenidate (Ritalin®, a frequently prescribed central nervous system stimulant) and its primary metabolite, ritalinic acid, in (1) water only or (2) with nine-spine stickleback and water louse. Methylphenidate degraded to ritalinic acid in both studies faster at a higher temperature (20 °C versus 10 °C), with concentrations of ritalinic acid surpassing methylphenidate after 48-100 h, depending on temperature. The concentration of methylphenidate in stickleback was highest at the first sampling point (60 min), while the concentration in water louse tissues reached comparatively higher levels and peaked after ~ 6 days. Neither stickleback nor water louse took up ritalinic acid in tissues despite being present in the water column. Our findings provide valuable data for use in future risk assessment of methylphenidate and will aid in the design of studies aimed at measuring any ecotoxicological effects on, for example, the behaviour or physiology of aquatic organisms.

The first protein which has been described to interact with the malacostracan Androgenic Gland Hormone (AGH) is a binding protein called IGFBP-rP1. It has been identified and studied in several species of decapods, in which its interaction with the masculinizing hormone and its expression patterns have been established in several ways. However, this protein remains uncharacterised to date in the other malacostracan orders, like Amphipoda and Isopoda, although they were historically the first ones in which the androgenic gland and the corresponding hormone were respectively described. In this article, we identified the IGFBP-rP1 of isopods and established its implication in the pathway of the AGH with a silencing approach in the model species Armadillidium vulgare. We also showed that this gene is expressed in all the tissues of males and females, with a similar pattern in animals infected with Wolbachia, a feminizing endosymbiont of several isopod species. The expression pattern did not differ during the development of uninfected and infected animals either. We finally studied the evolution of the IGFBP-rP1 in 68 isopod species, looking for conserved motifs and evidence of natural selection. Altogether, our results showed that this gene is constitutively expressed and strongly conserved in isopods, in which it likely constitutes a key element of the insulin/IGF signalling pathway. However, we also illustrated that IGFBP-rP1 is not sufficient on its own to explain the different developmental paths taken by the males and the females or feminized genetic males., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The present work aims to study the effects that acute exposure to low concentrations of silver nanoparticles (AgNPs) cause in digestive glands of terrestrial isopods (Porcellio scaber). The experiments were designed to integrate different analytical techniques, such as transmission electron microscopy, atomic absorption spectroscopy, proton induced X-ray emission, and Fourier transform IR imaging (FTIRI), in order to gain a comprehensive insight into the process from the AgNPs' synthesis to their interaction with biological tissues in vivo. To this aim, terrestrial isopods were fed with AgNPs having different shapes, sizes, and concentrations. For all the tested conditions, no toxicity at the whole organism level was observed after 14 days of exposure. However, FTIRI showed that AgNPs caused detectable local changes in proteins, lipids, nucleic acids and carbohydrates at the tissue level, to an extent dependent on the interplay of the AgNPs' properties: shape, size, concentration and dissolution of ions from them.

Parasites are widespread in natural environments, and their impacts on the fitness of their host and, at a broader scale, on ecosystem functioning are well recognized. Over the last two decades, there has been an increasing interest in the effects of parasites in conjunction with other stressors, especially pollutants, on the health of organisms. For instance, parasites can interfere with the bioaccumulation process of contaminants in their host leading to parasitized organisms exhibiting lower pollutants burdens than unparasitized individuals for example. However, the mechanisms underlying these patterns are not well understood. This study examined how the bopyrid parasite Gyge branchialis could lower the cadmium (Cd) uptake of its mud shrimp host Upogebia cf. pusilla. When exposed to water-borne Cd, parasites were able to bioaccumulate this trace metal. However, the uptake of Cd by the parasite was low and cannot entirely explain the deficit of Cd contamination of the host. The weight of gills of parasitized organisms was significantly reduced compared with unparasitized organisms. We suggest that by reducing the surface for metal uptake, parasites could lower the contaminant burden of their host.

Woody (lignocellulosic) plant biomass is an abundant renewable feedstock, rich in polysaccharides that are bound into an insoluble fiber composite with lignin. Marine crustacean woodborers of the genus Limnoria are among the few animals that can survive on a diet of this recalcitrant material without relying on gut resident microbiota. Analysis of fecal pellets revealed that Limnoria targets hexose-containing polysaccharides (mainly cellulose, and also glucomannans), corresponding with the abundance of cellulases in their digestive system, but xylans and lignin are largely unconsumed. We show that the limnoriid respiratory protein, hemocyanin, is abundant in the hindgut where wood is digested, that incubation of wood with hemocyanin markedly enhances its digestibility by cellulases, and that it modifies lignin. We propose that this activity of hemocyanins is instrumental to the ability of Limnoria to feed on wood in the absence of gut symbionts. These findings may hold potential for innovations in lignocellulose biorefining.

The rate of molecular evolution varies widely among species. Life history traits (LHTs) have been proposed as a major driver of these variations. However, the relative contribution of each trait is poorly understood. Here, we test the influence of metabolic rate (MR), longevity, and generation time (GT) on the nuclear and mitochondrial synonymous substitution rates using a group of isopod species that have made multiple independent transitions to subterranean environments. Subterranean species have repeatedly evolved a lower MR, a longer lifespan and a longer GT. We assembled the nuclear transcriptomes and the mitochondrial genomes of 13 pairs of closely related isopods, each pair composed of one surface and one subterranean species. We found that subterranean species have a lower rate of nuclear synonymous substitution than surface species whereas the mitochondrial rate remained unchanged. We propose that this decoupling between nuclear and mitochondrial rates comes from different DNA replication processes in these two compartments. In isopods, the nuclear rate is probably tightly controlled by GT alone. In contrast, mitochondrial genomes appear to replicate and mutate at a rate independent of LHTs. These results are incongruent with previous studies, which were mostly devoted to vertebrates. We suggest that this incongruence can be explained by developmental differences between animal clades, with a quiescent period during female gametogenesis in mammals and birds which imposes a nuclear and mitochondrial rate coupling, as opposed to the continuous gametogenesis observed in most arthropods.

Isopods of the genus Idotea have an unusual ability to feed on algae containing high amounts of chemical defense molecules, such as species of the genera Fucus and Ulva. In this study, we compared gene expression patterns of Idotea balthica individuals fed with Fucus vesiculosus to individuals fed with Ulva lactuca. We generated the first-ever transcriptome assembly for this species, and found 3,233 differentially expressed genes across feeding regimes. However, only a handful of biological functions were enriched with regard to differentially expressed genes, the most notable being "alkaloid metabolic process". Within this category, we found eight differentially expressed cytochrome P450 (CYP) unigenes, all of which had a higher expression in the U. lactuca diet treatment. A phylogenetic analysis showed that the differentially expressed CYP genes are closely related to a CYP gene described from the hepatopancreas of the spiny lobster Panulirus argus, and we hypothesize that these transcripts are involved in metabolite detoxification. This is a first step in the understanding of this algae-grazer interaction, and will form a basis for future work to characterize cytochrome P450 functioning in marine crustaceans.

Background: Woodlice are recognized as keystone species in terrestrial ecosystems due to their role in the decomposition of organic matter. Thus, they contribute to lignocellulose degradation and nutrient cycling in the environment together with other macroarthropods. Lignocellulose is the main component of plants and is composed of cellulose, lignin and hemicellulose. Its digestion requires the action of multiple Carbohydrate-Active enZymes (called CAZymes), typically acting together as a cocktail with complementary, synergistic activities and modes of action. Some invertebrates express a few endogenous lignocellulose-degrading enzymes but in most species, an efficient degradation and digestion of lignocellulose can only be achieved through mutualistic associations with endosymbionts. Similar to termites, it has been suspected that several bacterial symbionts may be involved in lignocellulose degradation in terrestrial isopods, by completing the CAZyme repertoire of their hosts., Results: To test this hypothesis, host transcriptomic and microbiome shotgun metagenomic datasets were obtained and investigated from the pill bug Armadillidium vulgare. Many genes of bacterial and archaeal origin coding for CAZymes were identified in the metagenomes of several host tissues and the gut content of specimens from both laboratory lineages and a natural population of A. vulgare. Some of them may be involved in the degradation of cellulose, hemicellulose, and lignin. Reconstructing a lignocellulose-degrading microbial community based on the prokaryotic taxa contributing relevant CAZymes revealed two taxonomically distinct but functionally redundant microbial communities depending on host origin. In parallel, endogenous CAZymes were identified from the transcriptome of the host and their expression in digestive tissues was demonstrated by RT-qPCR, demonstrating a complementary enzyme repertoire for lignocellulose degradation from both the host and the microbiome in A. vulgare., Conclusions: Our results provide new insights into the role of the microbiome in the evolution of terrestrial isopods and their adaptive radiation in terrestrial habitats.

Sphaeroma terebrans, an economically and ecologically important marine wood-boring isopod, is mainly distributed in tropical and subtropical mangroves. Nevertheless, available genomic and transcriptomic information for this isopod is extremely deficient. Here, we first performed the assembly of S. terebrans transcriptome by Illumina sequencing. A total of 51,092 high-quality unigenes with an average length of 641 bp were obtained and 19,915 unigenes were successfully annotated in four public databases. Only 9932 out of 19915 unigenes were commonly annotated by all four databases. In addition, 9609 unigenes were categorized into 54 function categories of Gene Ontology (GO), and 14,512 unigenes were successfully grouped into 25 functional categories of the EuKaryotic Orthologous Groups (KOG) database. Moreover, 11,507 unigenes were assigned to 228 pathways of the Kyoto Encyclopedia of Genes and Genomes (KEGG). Out of 51,092 unigenes, a total of 4257 different microsatellites with motifs range from di- to hexa-nucleotide were identified from 3324 unigene sequences. Among 64 primer pairs selected for validation, 35 were successful in PCR amplification and 13 exhibited obvious repeat polymorphisms in the wild population of S. terebrans in Dongzhaigang (Hainan Island). The transcriptome dataset and the identified microsatellite markers (SSRs) will provide abundant information for researches on the discovery of new genes, metabolic mechanism and genetic diversity of S. terebrans.

Background/aims: There is no treatment, without side effects, efficiently preventing or curing skin burns, caused by radiotherapy. A new experimental topical treatment protocol was assessed in mice receiving orthovoltage X-rays at an equivalent dose to that applied to human breast cancer patients in conventional radiotherapy., Methods: SKH-HR2 female hairless mice were irradiated on their dorsum with a total dose of 4,300 cGy during a 1-month period (20 fractions). The treatment group received a combination of 3 topical products, an oil-in-water cream, a gel containing Pinus halepensis bark aqueous extract, and an ointment containing olive oil extract of the marine isopod Ceratothoa oestroides. The positive control group was treated with a conventionally used commercial gel, whereas the negative control group did not receive any topical treatment. Skin alterations were evaluated by macroscopic examinations, measurements of transepidermal water loss (TEWL), melanin content, erythema intensity, hydration, and histopathology assessment., Results: Sixty days after radiation, TEWL and hydration values were abnormal and elements of acute, chronic, and granulomatous inflammation were present in all cases. The severest damage was detected in the deeper dermis. Treatment showed a comparatively beneficial effect on chronic and granulomatous inflammation while positive control was beneficial on acute inflammation., Conclusion: Skin anti-inflammatory treatment was the most effective but must be applied for several months. Further preclinical studies should be conducted, assimilating a human cancer radiation therapeutic schema with the aim of optimizing skin inflammation treatment., (© 2018 S. Karger AG, Basel.)

The evolution of current terrestrial life was founded by major waves of land invasion coinciding with high atmospheric oxygen content. These waves were followed by periods with substantially reduced oxygen concentration and accompanied by the evolution of novel traits. Reproduction and development are limiting factors for evolutionary water-land transitions, and brood care has probably facilitated land invasion. Peracarid crustaceans provide parental care for their offspring by brooding the early stages within the motherly brood pouch, the marsupium. Terrestrial isopod progeny begin ontogenetic development within the marsupium in water, but conclude development within the marsupium in air. Our results for progeny growth until hatching from the marsupium provide evidence for the limiting effects of oxygen concentration and for a potentially adaptive solution. Inclusion of air within the marsupium compensates for initially constrained growth in water through catch-up growth, and it may explain how terrestrial isopods adapted to short- and long-term changes in oxygen concentration., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2017. Published by The Company of Biologists Ltd.)

Shredders play a central role in the breakdown of leaf material in aquatic systems. These organisms and the ecological function they provide may, however, be affected by chemical stressors either as a consequence of direct waterborne exposure or through alterations in food-quality (indirect pathway). To unravel the biological relevance of these effect pathways, we applied a 2×2-factorial test design. Leaf material was microbially colonized for 10days in absence or presence of the fungicide epoxiconazole (15μg/L) and subsequently fed to the shredder Asellus aquaticus under exposure to epoxiconazole (15μg/L) or in fungicide-free medium over a 28-day period (n=40). Both effect pathways caused alterations in asselids' food processing, physiological fitness, and growth, although not always statistically significantly: assimilation either increased or remained at a similar level relative to the control suggesting compensatory behavior of A. aquaticus to cope with the enhanced energy demand for detoxification processes and decreased nutritional quality of the food. The latter was driven by lowered microbial biomasses and the altered composition of fatty acids associated with the leaf material. Even with increased assimilation, direct and indirect effects caused decreases in the growth and lipid (fatty acid) content of A. aquaticus with relative effect sizes between 10 and 40%. Moreover, the concentrations of two essential polyunsaturated fatty acids (i.e., arachidonic acid and eicosapentaenoic acid) were non-significantly reduced (up to ~15%) in asselids. This effect was, however, independent of the exposure pathway. Although waterborne effects were generally stronger than the diet-related effects, results suggest impaired functioning of A. aquaticus via both effect pathways., (Copyright © 2016 Elsevier B.V. All rights reserved.)

A test system to investigate the biomagnification of organic chemicals in the terrestrial isopod Porcellio scaber was developed and validated. Adult isopods were fed on alder leaf powder (Alnus glutinosa) spiked with [ 14 C]hexachlorobenzene (HCB). Test animals, sampled regularly during the uptake (16 d) and depuration phases (16 d), were analyzed, and the kinetics of tissue concentrations were determined. Uptake (k 1 ) and depuration rates (k 2 ) were calculated to estimate kinetic biomagnification factors (BMFs). In addition, the effect of coprophagy on the uptake and accumulation of HCB as well as the tissue distribution of HCB in P. scaber was investigated. The test system was shown to be suitable for investigations into the terrestrial bioaccumulation of chemicals. Coprophagy had no effect on the bioaccumulation of HCB in P. scaber. The hepatopancreas was identified as the main target tissue for HCB accumulation. The low BMF of 0.057 resulted from an assimilation efficiency (α) of 31.42%, a low uptake rate k 1 (0.009 d -1 ), and a high depuration rate k 2 (0.164 d -1 ). The results indicate that the terrestrial bioaccumulation of organic chemicals in P. scaber might not represent a worst-case scenario for biomagnification, limiting the value of the test system for the regulatory assessment of organic chemicals. Environ Toxicol Chem 2016;35:2867-2873. © 2016 The Authors. Environmental Toxicology and Chemistry Published by Wiley Periodicals, Inc. on behalf of SETAC., (© 2016 The Authors. Environmental Toxicology and Chemistry Published by Wiley Periodicals, Inc. on behalf of SETAC.)

We assessed the bioavailability of Ag from Ag nanoparticles (NPs), stabilized with polyvinylpyrrolidone (PVP), to terrestrial isopods which were exposed to 10, 100 and 1000 μg Ag NPs/g of dry food. Different Ag species were determined in the NP suspension that was fed to isopods: (i) total Ag by atomic absorption spectroscopy, (ii) the sum of Ag-PVP complexes and free Ag + by anodic stripping voltammetry at the bismuth-film electrode, and (iii) free Ag + by ion-selective potentiometry. The amounts of Ag species in the consumed food were compared to the masses of Ag accumulated in the isopod digestive glands. Our results show that all three Ag species (Ag NPs, Ag-PVP complexes and free Ag + ) could be the source of bioaccumulated Ag, but to various degrees depending on the exposure concentration and transformations in the digestive system. We provide a proof that (i) Ag NPs dissolve and Ag-PVP complexes dissociate in the isopod digestive tract; (ii) the concentration of free Ag + in the suspension offered to the test organisms is not the only measure of bioavailable Ag. The type of NP stabilizer along with the NP transformations in the digestive system needs to be considered in the creation of new computational models of the nanomaterial fate., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Chemical composition of organic matter (OM) is a key driver for detritus consumption by macroinvertebrates and polyunsaturated fatty acid (PUFA) content is considered a candidate indicator of food palatability. Since traditionally used complex natural OM covaries in many quality attributes, it remains uncertain whether benthic invertebrates developed an actual preference for PUFA-rich food. Therefore we aimed to test the influence of the PUFA linoleic acid on OM consumption by aquatic macroinvertebrates using standardized surrogate substrates (decomposition and consumption tablet, DECOTAB) with added linoleic acid (PUFA) in comparison to consumption of DECOTAB containing only cellulose (Standard) or ground macrophytes (Plant). In microcosms, we observed a higher consumption rate of PUFA DECOTAB in comparison to Standard DECOTAB in two functionally distinct invertebrate species (Lumbriculus variegatus and Asellus aquaticus). This effect appeared to be overruled in the field due to unknown sources of natural variation. Although we observed higher consumption rates in species-rich ditches compared to species-poor ditches, consumption rates were comparable for all three types of DECOTAB deployed. Upon reduced food quality and palatability, results presented here hint that PUFA like linoleic acid may be a key OM attribute driving the performance of benthic macroinvertebrates and inherent functioning of aquatic ecosystems.

The field of stoichiogenomics aims at understanding the influence of nutrient limitations on the elemental composition of the genome, transcriptome, and proteome. The 20 amino acids and the 4 nt differ in the number of nutrients they contain, such as nitrogen (N). Thus, N limitation shall theoretically select for changes in the composition of proteins or RNAs through preferential use of N-poor amino acids or nucleotides, which will decrease the N-budget of an organism. While these N-saving mechanisms have been evidenced in microorganisms, they remain controversial in multicellular eukaryotes. In this study, we used 13 surface and subterranean isopod species pairs that face strongly contrasted N limitations, either in terms of quantity or quality. We combined in situ nutrient quantification and transcriptome sequencing to test if N limitation selected for N-savings through changes in the expression and composition of the transcriptome and proteome. No evidence of N-savings was found in the total N-budget of transcriptomes or proteomes or in the average protein N-cost. Nevertheless, subterranean species evolving in N-depleted habitats displayed lower N-usage at their third codon positions. To test if this convergent compositional change was driven by natural selection, we developed a method to detect the strand-asymmetric signature that stoichiogenomic selection should leave in the substitution pattern. No such signature was evidenced, indicating that the observed stoichiogenomic-like patterns were attributable to nonadaptive processes. The absence of stoichiogenomic signal despite strong N limitation within a powerful phylogenetic framework casts doubt on the existence of stoichiogenomic mechanisms in metazoans., (© The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Nanoparticles (NPs) are increasingly being used, and they present the risk of being introduced into food webs. Numerous studies have been conducted to evaluate the toxicological effects of NPs in the aquatic and freshwater environments and their transfer to upper-level trophic organisms. However, information on the transfer and consequent effects of NPs on soil invertebrates is still limited. In this study, we assessed the transfer of quantum dots (QDs) through a three-species terrestrial food chain that consisted of the yeast Saccharomyces cerevisiae, the collembolan Folsomia candida, and the pill bug Armadillidium vulgare, as well as their biodistribution in vital organs using fluorescence analytical techniques. To visualize QD incorporation and biodistribution in F. candida, longitudinal and transversal sections were observed after short-term (3 d) and long-term (12 d) feeding with QD-treated yeast. QDs were located only in the intestine of F. candida and excreted within 1-2 d. QDs were also transferred to the pill bug by feeding, and remained in its intestine. This study showed the transfer of NPs through a model terrestrial food chain and indicated the potential hazards of released NPs for organisms at different trophic levels., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Unlabelled: This work aimed at characterizing the metabolome of the isopod Porcellionides pruinosus and at assessing its variations over 14 days under laboratory culture conditions and upon exposure to the contaminant metal Nickel (Ni). The spectral profiles obtained by (1)H NMR spectroscopy were thoroughly assigned and subjected to multivariate analysis in order to highlight consistent changes. Over 50 metabolites could be identified, providing considerable new knowledge on the metabolome of these model organisms. Several metabolites changed non-linearly with Ni dose and exposure time, showing distinct variation patterns for initial (4 days) and later time points (7 and 14 days). In particular, at day 4, several amino acids were increased and sugars were decreased (compared to controls), whereas these variations were inverted for longer exposure, possibly reflecting earlier and more intensive moulting. Other variations, namely in betaines and choline-containing compounds, were suggested to relate with osmoregulation and detoxification mechanisms. Ni also had a marked effect on several nucleotides (increased upon exposure) and a moderate impact on lipids (decreased upon exposure). Overall, this study has provided new information on the Ni-induced metabolic adaptations of the P. pruinosus isopod, paving the way for improved mechanistic understanding of how these model organisms handle soil contamination., Significance: This study provided, for the first time to our knowledge, a detailed picture of the NMR-detectable metabolome of terrestrial isopods and of its fluctuations in time and upon exposure to the contaminant metal Nickel. Several time- and dose-dependent changes were highlighted, providing mechanistic insight into how these important model organisms handle Ni contamination.

Woodlice efficiently sequester copper (Cu) in 'cuprosomes' within hepatopancreatic 'S' cells. Binuclear 'B' cells in the hepatopancreas form iron (Fe) deposits; these cells apparently undergo an apocrine secretory diurnal cycle linked to nocturnal feeding. Synchrotron-based µ-focus X-ray spectroscopy undertaken on thin sections was used to characterize the ligands binding Cu and Fe in S and B cells of Oniscus asellus (Isopoda). Main findings were: (i) morphometry confirmed a diurnal B-cell apocrine cycle; (ii) X-ray fluorescence (XRF) mapping indicated that Cu was co-distributed with sulfur (mainly in S cells), and Fe was co-distributed with phosphate (mainly in B cells); (iii) XRF mapping revealed an intimate morphological relationship between the basal regions of adjacent S and B cells; (iv) molecular modelling and Fourier transform analyses indicated that Cu in the reduced Cu(+) state is mainly coordinated to thiol-rich ligands (Cu-S bond length 2.3 Å) in both cell types, while Fe in the oxidized Fe(3+) state is predominantly oxygen coordinated (estimated Fe-O bond length of approx. 2 Å), with an outer shell of Fe scatterers at approximately 3.05 Å; and (v) no significant differences occur in Cu or Fe speciation at key nodes in the apocrine cycle. Findings imply that S and B cells form integrated unit-pairs; a functional role for secretions from these cellular units in the digestion of recalcitrant dietary components is hypothesized., (© 2016 The Authors.)

Biomolecular profiling with Fourier-Transform InfraRed Microscopy was performed to distinguish the Zn(2+)-mediated effects on the crustacean (Porcellio scaber) digestive glands from the ones elicited by the ZnO nanoparticles (NPs). The exposure to ZnO NPs or ZnCl2 (1500 and 4000 µg Zn/g of dry food) activated different types of metabolic pathways: some were found in the case of both substances, some only in the case of ZnCl2, and some only upon exposure to ZnO NPs. Both the ZnO NPs and the ZnCl2 increased the protein (∼1312 cm(-1); 1720-1485 cm(-1)/3000-2830 cm(-1)) and RNA concentration (∼1115 cm(-1)). At the highest exposure concentration of ZnCl2, where the effects occurred also at the organismal level, some additional changes were found that were not detected upon the ZnO NP exposure. These included changed carbohydrate (most likely glycogen) concentrations (∼1043 cm(-1)) and the desaturation of cell membrane lipids (∼3014 cm(-1)). The activation of novel metabolic pathways, as evidenced by changed proteins' structure (at 1274 cm(-1)), was found only in the case of ZnO NPs. This proves that Zn(2+) are not the only inducers of the response to ZnO NPs. Low bioavailable fraction of Zn(2+) in the digestive glands exposed to ZnO NPs further supports the role of particles in the ZnO NP-generated effects. This study provides the evidence that ZnO NPs induce their own metabolic responses in the subtoxic range.

The acanthocephalan parasite Acanthocephalus dirus infects the freshwater isopod Caecidotea intermedius as an intermediate host before completing its life cycle in a fish. Transmission to the definitive host occurs after the parasite has reached the cystacanth stage, and development into this stage is associated with changes in several behavioral and physiological traits of the host. Given the potential importance of host energy availability to trait modification, we examined the relationship between cystacanth-stage infection and energy storage of adult isopods. Six samples of infected and uninfected male C. intermedius were collected from a population in March, April, and May during which time cystacanth-stage A. dirus dominate infections and modification of behavior and physiology occurs in nature. Biochemical assays revealed that infected male C. intermedius contained more glycogen and more lipid than uninfected males and that this difference was present throughout the sampling period, which represents the entire adult phase of the host's life. Additional analysis revealed that infected and uninfected males differed in their pattern of allocation to each energy source and that host lipid levels were negatively correlated with parasite intensity. We propose that the typical pattern of allocation and storage of host energy appears to be disrupted by A. dirus infection and that the changes are more likely to favor the parasite than the host.

The effects of Ag-NPs and AgNO3 on the isopod Porcellionides pruinosus were determined upon soil and dietary exposures. Isopods avoided Ag in soil, with EC50 values of ∼16.0 and 14.0 mg Ag/kg for Ag-NPs and AgNO3, respectively. Feeding inhibition tests in soil showed EC50s for effects on consumption ratio of 127 and 56.7 mg Ag/kg, respectively. Although similar EC50s for effects on biomass were observed for nanoparticulate and ionic Ag (114 and 120 mg Ag/kg dry soil, respectively), at higher concentrations greater biomass loss was found for AgNO3. Upon dietary exposure, AgNO3 was more toxic, with EC50 for effects on biomass change being >1500 and 233 mg Ag/kg for Ag-NPs and AgNO3, respectively. The difference in toxicity between Ag-NPs and AgNO3 could not be explained from Ag body concentrations. This suggests that the relation between toxicity and bioavailability of Ag-NPs differs from that of ionic Ag in soils., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The aim of the research, carried out on three species of terrestrial isopods - Armadillidium granulatum, Halophiloscia hirsuta and Trichoniscus alexandrae - is to bring a first consistent contribution to the knowledge of the ultrastructural organization of the testis follicles. The testis follicles are seat of a remarkable dynamic activity of their cell components (somatic cells and germ cells) that results in a continuous variation, related to the trend of spermatogenesis, of their morphology, organization and of the relationships between the two cell populations. The somatic cells, known in literature as follicular cells, nurse cells or Sertoli cells, are arranged at the periphery of the follicle to form an epithelial layer of variable thickness resting on a thin basal lamina in turn surrounded by a discontinuous network of muscle cells. In A. granulatum and H. hirsuta, two types of Sertoli cells are present: a first type, the nurse cells, envelop the spermatids in cavities within their cytoplasm and through their secretion activity play a fundamental role in the formation of the spermatophores; moreover, they phagocytizes the residual cytoplasm of spermatids. A second type of Sertoli cells shows features that leave clearly identify its supporting role to the spermatophores in formation. In T. alexandrae, instead, only one type of Sertoli cells, the nurse cell, is present, whose features are widely superimposable to those observed in the other two species. Moreover, two septa of Sertoli cells depart from the periphery of the testis follicle to constitute an articulated compartmentalization of the follicle itself, probably targeted to realize at its inside a series of microenvironments functionally diversified in order to meets the needs of the different stages of the spermatogenic cycle., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The heavy metals bioaccumulation capability in Armadillidium vulgare feeded with chestnut leaves contaminated with various sublethal concentrations of Cd and Pb, was evaluated under laboratory conditions. The metal concentration found in the hepatopancreas of treated animals, as measured by Inductively Coupled Plasma Mass Spectrometry (ICP-MS), affected the expression and localization of MT and HSP70 as shown by immunohistochemical and western blotting analysis. The Cd content of the animals treated with the various concentrations of the metal has been always higher than that of chestnut leaves contaminated. The accumulation of Pb was, instead, always modest compared to the content of the chestnut leaves. The immunohistochemical investigation in hepatopancreas tissue of animals treated with increasing concentrations of Cd and Pb, by using the anti-MT and anti-HSP70 antibodies, has provided a response clearly positive even if differentiated in relation to the metal and concentration tested. In particular, a positive response to anti-MT antibody was detected in B and S cells nuclei and S cells cytoplasm; the localization of HSP70 was particularly intense at the cell surface. Western blotting analysis showed significant up-regulation of the expression (about 2.6 fold) of HSP70 proteins in the hepatopancreas of animals exposed to highest Pb concentrations respect to control. Moreover, samples exposed to higher Cd and Pb concentrations showed a higher expression of MT (3.2 fold and 4 fold respectively) compared to control. In summary, our data beyond to clearly demonstrate for the first time the expression of MT in terrestrial isopods, suggest that A. vulgare would be a suitable organism for assessing Cd and Pb exposure in environments threatened by metal pollution as suggested by the modulation of the biomarkers MT and HSP70., (Copyright © 2015 Elsevier Inc. All rights reserved.)

This study tests the potential for using Armadillo officinalis as a bioindicator of exposure to and activation of benzene metabolic pathways using an in vivo model. A. officinalis specimens collected in a natural reserve were divided into a control and three test groups exposed to 2.00, 5.32 or 9.09 µg/m(3) benzene for 24h. Three independent tests were performed to assess model reproducibility. Animals were dissected to obtain three pooled tissue samples per group: hepatopancreas (HEP), other organs and tissues (OOT), and exoskeleton (EXO). Muconic acid (MA), S-phenylmercapturic acid (S-PMA), two human metabolites of benzene, and changes in mtDNA copy number, a human biomarker of benzene exposure, were determined in each sample; benzene was determined only in EXO. MA was measured by high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection, S-PMA by triple quadrupole mass spectrometer liquid chromatography with electro spray ionization (LC-MS-ESI-TQD), mtDNA by real-time quantitative PCR and end-point PCR, and benzene by quadrupole mass spectrometer head-space gas chromatography (HSGC-MS). MA and S-PMA levels rose both in HEP and OOT; EXO exhibited increasing benzene concentrations; and mtDNA copy number rose in HEP but not in OOT samples. Overall, our findings demonstrate that A. officinalis is a sensitive bioindicator of air benzene exposure and show for the first time its ability to reproduce human metabolic dynamics., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The aim of this study was to determine whether citric acid adsorbed onto cobalt ferrite (CoFe2O4) nanoparticles (NPs) influences the bioavailability of their constituents Co and Fe. Dissolution of Co and Fe was assessed by two measures: (i) in aqueous suspension using chemical analysis, prior to application onto the food of test organisms; and (ii) in vivo, measuring the bioavailability in the model terrestrial invertebrate (Porcellio scaber, Isopoda, Crustacea). The isopods were exposed to citric-acid-adsorbed CoFe2O4 NPs for 2 weeks, and tissue accumulation of Co and Fe was assessed. This was compared to pristine CoFe2O4 NPs, and CoCl2 and Fe(III) salts as positive controls. The combined data shows that citric acid enhances free metal ion concentration from CoFe2O4 NPs in aqueous suspension, although in vivo, very similar amounts of assimilated Co were found in isopods exposed to both types of NPs. Therefore, evaluation of the dissolution in suspension by chemical means is not a good predictor of metal assimilation of this model organism; body assimilation of Co and Fe is rather governed by the physiological capacity of P. scaber for the uptake of these metals. Moreover, we propose that citric acid, due to its chelating properties, may hinder the uptake of Co that dissolves from citric-acid-adsorbed CoFe2O4 NPs, if citric acid is present in sufficient quantity., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Supralittoral and high intertidal coastal zones are exposed to pollution from both marine and terrestrial sources and undergo higher deposition rates than the subtidal zone. It is therefore important to identify organisms for this section of the coastal area that can be tolerant to contaminants. The aim of this study was to determine if supralittoral isopods of the genus Ligia can be used as biomonitors, since they are abundant and widely distributed. For this purpose, concentrations of trace elements were determined in Ligia isopods in toto from 26 locations across the Gulf of California and Pacific coast of the Baja California peninsula, which were collected during the summers of 2009 and 2010. The concentrations of trace elements followed the order of; Zn≥Cu>As>Cd>Pb>Hg. Elevated concentrations of copper (up to 1010 μg/g) were detected in Ligia from Santa Rosalía (SRo), a locality where industrial mining of copper has historically occurred. Industrial and municipal sewage discharges appear to have contributed to the high concentrations of zinc (326 μg/g) and lead (144 μg/g) found in organisms from Guaymas location. The high mercury concentration in organisms from Mazatlán (M) (2.01 μg/g) was associated with a thermoelectric plant. Natural sources of metals were also detected; coastal upwelling appears to be associated with high cadmium concentrations in Ligia from Punta Baja (PB) (256 μg/g) in the Pacific coast, whereas hydrothermal vents may have contributed to high concentrations of arsenic at Ensenada (E) (61 μg/g). Our results suggest that Ligia isopods reflect the natural and anthropogenic inputs of trace metals in the environment and could potentially be used as biomonitor organisms of the intertidal rocky shores of the Gulf of California and Pacific coast., (Copyright © 2014 Elsevier Inc. All rights reserved.)

This study was designed to compare cadmium (Cd) and lead (Pb) bioaccumulation in three species of oniscidean isopods - Armadillidium granulatum Brandt, Armadillidium vulgare (Latreille) and Porcellio laevis Latreille which were exposed for three weeks to a contaminated diet, and to determine the morphological and ultrastructural changes in hepatopancreas. Metal accumulation, determined by Inductively Coupled Plasma Mass Spectrometry (ICP-MS), was linearly associated with the exposed concentration and was a function of the metal and the species tested. All three species accumulated lower levels of Pb than Cd. A. vulgare accumulated the largest concentration of Pb, especially at the higher doses, whereas P. laevis showed the greatest Cd accumulation, and the highest Cd concentration was lethal for all exposed species. The highest concentrations of Pb and Cd induced significant changes both in the general morphology of tubules and in the ultrastructural organization of epithelial cells in hepatopancreas. Some Pb/Cd induced alterations include: brush border disorganization; reduction of the basal labyrinth formed by the plasma membrane; condensation of some cytoplasm areas and of chromatin; rough endoplasmic reticulum and mitochondrial alterations; increase of secondary lysosomes and of type B granules in S cells. Some of the ultrastructural changes observed overlap with those induced by prolonged starvation, whereas others can be useful biomarkers of heavy metal toxicity. This study has confirmed that in terrestrial isopods, the accumulation of the different metals occurs in a species-specific manner; therefore ecological monitoring and assessment studies should consider each species individually. The research has confirmed that in the terrestrial isopods the accumulation of the different metals occurs in a species-specific way; therefore each species should first be evaluated in view of its employ in biomonitoring programs., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Plastic pollution is an emerging global threat for marine wildlife. Many species of birds, reptiles, and fishes are directly impaired by plastics as they can get entangled in ropes and drown or they can ingest plastic fragments which, in turn, may clog their stomachs and guts. Microplastics of less than 1 mm can be ingested by small invertebrates, but their fate in the digestive organs and their effects on the animals are yet not well understood. We embedded fluorescent microplastics in artificial agarose-based food and offered the food to marine isopods, Idotea emarginata. The isopods did not distinguish between food with and food without microplastics. Upon ingestion, the microplastics were present in the stomach and in the gut but not in the tubules of the midgut gland which is the principal organ of enzyme-secretion and nutrient resorption. The feces contained the same concentration of microplastics as the food which indicates that no accumulation of microplastics happens during the gut passage. Long-term bioassays of 6 weeks showed no distinct effects of continuous microplastic consumption on mortality, growth, and intermolt duration. I. emarginata are able to prevent intrusion of particles even smaller than 1 μm into the midgut gland which is facilitated by the complex structure of the stomach including a fine filter system. It separates the midgut gland tubules from the stomach and allows only the passage of fluids and chyme. Our results indicate that microplastics, as administered in the experiments, do not clog the digestive organs of isopods and do not have adverse effects on their life history parameters.

The analgesic activity of Porcellio laevis Latreille, Rhizoma Corydalis, and Radix Cynanchi Paniculati have been reported in recent years. A new formula named Jia-Yuan-Qing pill (JYQP) is therefore created by combining the three herbs at 9:7:7 ratio according to traditional Chinese theories. The present study aims to evaluate the effect of JYQP as a novel painkiller in various models. Acute toxicity test was applied to evaluate the safety of JYQP. Acetic-acid-induced writhing, hot plate test, formalin test, and naloxone-pretreated writhing test were employed to elaborate the analgesic activity of JYQP and its possible mechanism. A bone cancer pain mouse model was performed to further assess the effect of JYQP in relieving cancer pain. Test on naloxone-precipitated withdrawal symptoms was conduct to examine the physical dependence of mice on JYQP. Data revealed that JYQP reduced writhing and stretching induced by acetic acid; however, this effect could not be blocked by naloxone. JYQP specifically suppressed the phase II reaction time in formalin-treated mice; meanwhile, no analgesic effect of JYQP in hot plate test was observed, indicating that JYQP exerts analgesic activity against inflammatory pain rather than neurogenic pain. Furthermore, JYQP could successfully relieve bone cancer pain in mice. No physical dependence could be observed upon long-term administration in mice. Collectively, our present results provide experimental evidence in supporting clinical use of JYQP as an effective and safe agent for pain treatment.

The effects of soil pH on the toxicity of ZnO nanoparticles (NPs) to the terrestrial isopod Porcellionides pruinosus were evaluated. Isopods were exposed to a natural soil amended with CaCO3 to reach 3 different pH(CaCl2) levels (4.5, 6.2, and 7.3) and to standard LUFA 2.2 soil (pH 5.5) spiked with ZnO NPs (30 nm), non-nano ZnO (200 nm), and ionic Zn as ZnCl₂. Toxicity was expressed based on total Zn concentration in soil, as well as total Zn and free Zn²⁺ ion concentrations in porewater. Compared with ZnO-spiked soils, the ZnCl₂-spiked soils had lower pH and higher porewater Ca²⁺ and Zn levels. Isopod survival did not differ between Zn forms and soils, but survival was higher for isopods exposed to ZnO NPs at pH 4.5. Median effect concentrations (EC50s) for biomass change showed similar trends for all Zn forms in all soils, with higher values at intermediate pH. Median lethal concentration (LC50) and EC50 values based on porewater Zn or free Zn ion concentrations were much lower for ZnO than for ionic zinc. Zn body concentrations increased in a dose-related manner, but no effect of soil pH was found. It is suggested not only that dissolved or free Zn in porewater contributed to uptake and toxicity, but also that oral uptake (i.e., ingestion of soil particles) could be an important additional route of exposure., (© 2013 SETAC.)