Pill Bug Armadillidium Vulgare Research Articles

Metagenome-assembled genomes of three Hepatoplasmataceae provide insights into isopod-mollicute symbiosis.

The digestive organs of terrestrial isopods harbour bacteria of the recently proposed mollicute family Hepatoplasmataceae. The only complete genome available so far for Hepatoplasmataceae is that of 'Candidatus Hepatoplasma crinochetorum'. The scarcity of genome sequences has hampered our understanding of the symbiotic relationship between isopods and mollicutes. Here, we present four complete metagenome-assembled genomes (MAGs) of uncultured Hepatoplasmataceae members identified from shotgun sequencing data of isopods. We propose genomospecies names for three MAGs that show substantial sequence divergence from any previously known Hepatoplamsataceae members: 'Candidatus Tyloplasma litorale' identified from the semiterrestrial isopod Tylos granuliferus, 'Candidatus Hepatoplasma vulgare' identified from the common pill bug Armadillidium vulgare, and 'Candidatus Hepatoplasma scabrum' identified from the common rough woodlouse Porcellio scaber. Phylogenomic analysis of 155 mollicutes confirmed that Hepatoplasmataceae is a sister clade of Metamycoplasmataceae in the order Mycoplasmoidales. The 16S ribosomal RNA gene sequences and phylogenomic analysis showed that 'Candidatus Tyloplasma litorale' and other semiterrestrial isopod-associated mollicutes represent the placeholder genus 'g_Bg2' in the r214 release of the Genome Taxonomy Database, warranting their assignment to a novel genus. Our analysis also revealed that Hepatoplasmataceae lack major metabolic pathways but has a likely intact type IIA CRISPR-Cas9 machinery. Although the localization of the Hepatoplasmatacae members have not been verified microscopically in this study, these genomic characteristics are compatible with the idea that these mollicutes have an ectosymbiotic lifestyle with high nutritional dependence on their host, as has been demonstrated for other members of the family. We could not find evidence that Hepatoplasmataceae encode polysaccharide-degrading enzymes that aid host digestion. If they are to provide nutritional benefits, it may be through extra-copy nucleases, peptidases, and a patatin-like lipase. Exploration of potential host-symbiont interaction-associated genes revealed large, repetitive open reading frames harbouring beta-sandwich domains, possibly involved with host cell adhesion. Overall, genomic analyses suggest that isopod-mollicute symbiosis is not characterized by carbohydrate degradation, and we speculate on their potential role as defensive symbionts through spatial competition with pathogens to prevent infection.

Male Feminization of the Common Pillbug Armadillidium vulgare by Wolbachia bacteria

Wolbachia pipientis (Wolbachia) is a maternally inherited endosymbiotic bacterium that infects over 50% of arthropods, including pillbugs, and acts as a reproductive parasite in the host. In the common terrestrial pillbug Armadillidium vulgare (A. vulgare), Wolbachia alters the sex ratio of offspring through a phenomenon called feminization, where genetic males develop into reproductive females. Previous studies have focused on the presence or absence of Wolbachia as a sex ratio distorter in laboratory cultured and natural populations mainly from sites in Europe and Japan. Our three-year study is the first to evaluate the effects of the Wolbachia sex ratio distorter in cultured A. vulgare offspring in North America. We asked whether Wolbachia bacteria feminize A. vulgare isopod male offspring from infected mothers and if this effect can be detected in F1 offspring by comparing the male/female offspring ratios. If so, the F1 offspring ratio should show a higher number of females than males compared to the offspring of uninfected mothers. Over three years, pillbug offspring were cultured from pregnant A. vulgare females and developed into adults. We determined the Wolbachia status of mothers and counted the ratios of male and female F1 progeny to determine feminization effects. In each year sampled, significantly more female offspring were born to Wolbachia-infected mothers than those from uninfected mothers. These ratio differences suggest that the Wolbachia infection status of mothers directly impacts the A. vulgare population through the production of reproductive feminized males, which in turn provides an advantage for further Wolbachia transmission.

Social aggregation in the terrestrial isopod Armadillidium vulgare

Social aggregative behaviour may have evolved in marine isopods to aid hunting, and help the animals protect themselves from predation. This social behaviour could have assisted some isopod species to make the evolutionary transition from marine to terrestrial environments, as aggregative huddling of isopods on land reduces desiccation when out of water. To determine if the common pill-bug Armadillidium vulgare demonstrates social behaviour, groups of animals were filmed for 60 min in an arena that contained a binary choice of two shelters. This experiment was directly repeated with the same specimens three more times in the same day and this series of four experimental runs was replicated a total of 10 times with a different group of animals for each of the 10 series. In most cases, the majority of specimens chose one of the two identical shelters at random, and congregated under this winning shelter. The amount of animals that congregated under the winning shelter, however, significantly decreased over the four consecutive runs, whereas the amount of animals that congregated under the losing shelter significantly increased with prior experience. These data indicate that A. vulgare is a social animal but this social behaviour can decrease with exposure to the novel environment. This attenuation in social behaviour may serve to decrease the drive to join the larger of the two groups in an area where a danger from predation has not been encountered and allow experienced animals to explore new terrain.

Influence of saprophages (Isopoda, Diplopoda) on leaf litter decomposition under different levels of humidification and chemical loading

The paper presents a study about the influence of two saprophage groups (Isopoda, Diplopoda) on leaf litter decomposition under different levels of humidification and chemical stress. Because of their worldwide distribution, we focused on the common pillbug Armadillidium vulgare (Latreille, 1804) (Isopoda, Armadillidiidae), and the common millipede species Rossiulus kessleri (Lohmander, 1927) (Julida, Julidae). The function of environment creation by the given saprophages, as destructors of dead plant matter, supporting such ecosystem services as soil fertility improvement and nutrients’ turnover, is highlighted. To conduct the experiment, the animals were collected manually and using pitfall trapping. In order to bring the experimental conditions closer to the natural, the individuals were not sexed. The maximum consumption of leaf litter by woodlice was recorded in the conditions with adequate moisture (0.5 mL of distilled water per box) and amounted to 2.52 mg/10 individuals per day, which exceeds its consumption with low and increased moisture, respectively, by 1.82 and 1.24 times. As for the effect of interaction, the consumption of maple litter with optimal moisture (4.77 mg/10 individuals per day) was the greatest. The largest absolute difference between broad-leaved tree species in the average weight of leaf litter consumed by woodlice was between maple leaf litter and oak leaf litter, the minimum – between robinia leaf litter and oak leaf litter. According to the results of the obtained data (Diplopoda), it can be stated that there is a statistically significant effect of chemical stress and discrepancy of the average zinc content in the object of study (in Diplopoda and their faecal pellets). We found that the diet provided did not affect the distribution of zinc in Diplopoda under conditions of chemical stress. According to the results of pairwise comparisons, we determined that the zinc content in the Diplopoda clearly differs for control and almost every concentration of zinc sulfate solution – 0.03 and 0.15 g/L, the samples of which do not form a homogeneous group. The species composition, abundance and distribution in space of soil invertebrates are informative indicators which reflect the ecological state of soils, intensity in development of soil horizons as well as intensity of processes occurring in them.

Lignocellulose degradation at the holobiont level: teamwork in a keystone soil invertebrate

BackgroundWoodlice 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.ResultsTo 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.ConclusionsOur results provide new insights into the role of the microbiome in the evolution of terrestrial isopods and their adaptive radiation in terrestrial habitats.

Birth of a W sex chromosome by horizontal transfer of Wolbachia bacterial symbiont genome

Sex determination is a fundamental developmental pathway governing male and female differentiation, with profound implications for morphology, reproductive strategies, and behavior. In animals, sex differences between males and females are generally determined by genetic factors carried by sex chromosomes. Sex chromosomes are remarkably variable in origin and can differ even between closely related species, indicating that transitions occur frequently and independently in different groups of organisms. The evolutionary causes underlying sex chromosome turnover are poorly understood, however. Here we provide evidence indicating that Wolbachia bacterial endosymbionts triggered the evolution of new sex chromosomes in the common pillbug Armadillidium vulgare We identified a 3-Mb insert of a feminizing Wolbachia genome that was recently transferred into the pillbug nuclear genome. The Wolbachia insert shows perfect linkage to the female sex, occurs in a male genetic background (i.e., lacking the ancestral W female sex chromosome), and is hemizygous. Our results support the conclusion that the Wolbachia insert is now acting as a female sex-determining region in pillbugs, and that the chromosome carrying the insert is a new W sex chromosome. Thus, bacteria-to-animal horizontal genome transfer represents a remarkable mechanism underpinning the birth of sex chromosomes. We conclude that sex ratio distorters, such as Wolbachia endosymbionts, can be powerful agents of evolutionary transitions in sex determination systems in animals.

In vivo visual evaluation of nanoparticle transfer in a three-species terrestrial food chain

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.

Large gene overlaps and tRNA processing in the compact mitochondrial genome of the crustacean Armadillidium vulgare

A faithful expression of the mitochondrial DNA is crucial for cell survival. Animal mitochondrial DNA (mtDNA) presents a highly compact gene organization. The typical 16.5 kbp animal mtDNA encodes 13 proteins, 2 rRNAs and 22 tRNAs. In the backyard pillbug Armadillidium vulgare, the rather small 13.9 kbp mtDNA encodes the same set of proteins and rRNAs as compared to animal kingdom mtDNA, but seems to harbor an incomplete set of tRNA genes. Here, we first confirm the expression of 13 tRNA genes in this mtDNA. Then we show the extensive repair of a truncated tRNA, the expression of tRNA involved in large gene overlaps and of tRNA genes partially or fully integrated within protein-coding genes in either direct or opposite orientation. Under selective pressure, overlaps between genes have been likely favored for strong genome size reduction. Our study underlines the existence of unknown biochemical mechanisms for the complete gene expression of A. vulgare mtDNA, and of co-evolutionary processes to keep overlapping genes functional in a compacted mitochondrial genome.

Feminizing Wolbachia: a transcriptomics approach with insights on the immune response genes in Armadillidium vulgare

BackgroundWolbachia are vertically transmitted bacteria known to be the most widespread endosymbiont in arthropods. They induce various alterations of the reproduction of their host, including feminization of genetic males in isopod crustaceans. In the pill bug Armadillidium vulgare, the presence of Wolbachia is also associated with detrimental effects on host fertility and lifespan. Deleterious effects have been demonstrated on hemocyte density, phenoloxidase activity, and natural hemolymph septicemia, suggesting that infected individuals could have defective immune capacities. Since nothing is known about the molecular mechanisms involved in Wolbachia-A. vulgare interactions and its secondary immunocompetence modulation, we developed a transcriptomics strategy and compared A. vulgare gene expression between Wolbachia-infected animals (i.e., “symbiotic” animals) and uninfected ones (i.e., “asymbiotic” animals) as well as between animals challenged or not challenged by a pathogenic bacteria.ResultsSince very little genetic data is available on A. vulgare, we produced several EST libraries and generated a total of 28 606 ESTs. Analyses of these ESTs revealed that immune processes were over-represented in most experimental conditions (responses to a symbiont and to a pathogen). Considering canonical crustacean immune pathways, these genes encode antimicrobial peptides or are involved in pathogen recognition, detoxification, and autophagy. By RT-qPCR, we demonstrated a general trend towards gene under-expression in symbiotic whole animals and ovaries whereas the same gene set tends to be over-expressed in symbiotic immune tissues.ConclusionThis study allowed us to generate the first reference transcriptome ever obtained in the Isopoda group and to identify genes involved in the major known crustacean immune pathways encompassing cellular and humoral responses. Expression of immune-related genes revealed a modulation of host immunity when females are infected by Wolbachia, including in ovaries, the crucial tissue for the Wolbachia route of transmission.

Effects of parasitic sex‐ratio distorters on host genetic structure in the Armadillidium vulgare–Wolbachia association

In the pill bug Armadillidium vulgare (Crustacea, Oniscidea), Wolbachia facilitates its spread through vertical transmission via the eggs by inducing feminization of genetic males. The spread of feminizing Wolbachia within and across populations is therefore expected to influence mitochondrial DNA (mtDNA) genetic structure by hitchhiking. To test this hypothesis, we analysed nuclear and mtDNA genetic structure, and Wolbachia prevalence in 13 populations of the pill bug host. Wolbachia prevalence (ranging from 0% to 100% of sampled females) was highly variable among populations. All three Wolbachia strains previously observed in A.vulgare were present (wVulC, wVulM and wVulP) with wVulC being the most prevalent (nine of 13 populations). The host showed a genetic structure on five microsatellite loci that is compatible with isolation by distance. The strong genetic structure observed on host mtDNA was correlated with Wolbachia prevalence: three mitotypes were in strong linkage disequilibrium with the three strains of Wolbachia. Neutrality tests showed that the mtDNA polymorphism is not neutral, and we thus suggest that this unusual pattern of mtDNA polymorphism found in A.vulgare was due to Wolbachia.

2A2-G02 オカダンゴムシの交替性転向反応を模擬するニューラルネットワークの構築とその実機実験

This paper explains how to construct the micom car that imitates turn alternation behavior of the Pill-bug Armadillidium Vulgare. The micom car is controlled by artificial neural networks (ANNs). ANNs are designed by the back propagation through the proposed simulator that mimicking the minimal simulation of the Sussex approach.

オカダンゴムシの交替性転向反応とその逃避行動としての意味

The pill bug Armadillidium vulgare shows clear turn alternation behavior. As the mechanism underlying the behavior, the bilateral asymmetrical leg movement (BALM) hypothesis has been proposed. Pill bugs, however, showed a longer following when in a zigzag pathway than in a straight pathway, suggesting the presence of an innate turning pattern in their neural system. On the other hand, they showed clear thigmotaxis: they walked a similar distance along a single side-wall as along a straight pathway. The adaptive significance of this behavior was analyzed in an artificial arena. The turn alternation was more frequent in an escaping situation than in voluntary walking, and also the walking speed was faster when escaping than voluntary walking. These results suggest that the turn alternation of the pill bug functions as a behavior for effective escape from natural enemies.

Evidence for a new feminizing Wolbachia strain in the isopod Armadillidium vulgare: evolutionary implications.

Wolbachia are intracellular maternally inherited alpha-Proteobacteria infecting a wide range of arthropods. In the common pill bug Armadillidium vulgare, the known Wolbachia strain is responsible for feminization of genetic males. We have investigated Wolbachia diversity in 20 populations of A. vulgare from west and east Europe, north Africa and north America. A new Wolbachia strain (wVulM) was identified through the variability of the wsp gene, distantly related to that previously known (wVulC) in this host species. No individual with multiple infections was detected. Inoculation experiments indicated that the new wVulM bacterial strain also induces feminization in A. vulgare. However, the wVulC strain showed a higher transmission rate than the wVulM strain and was the most geographically widespread Wolbachia in A. vulgare populations. Mitochondrial 16SrDNA gene sequencing was conducted in Wolbachia-infected individuals, revealing the occurrence of four host lineages. The comparison of bacterial strains and their respective host mitochondrial phylogenies failed to show concordance, indicating horizontal transmission of the Wolbachia strains within populations of A. vulgare.

Conflict between feminizing sex ratio distorters and an autosomal masculinizing gene in the terrestrial isopod Armadillidium vulgare Latr.

Female sex determination in the pill bug Armadillidium vulgare is frequently under the control of feminizing parasitic sex factors (PSF). One of these PSF is an intracytoplasmic Wolbachia-like bacterium (F), while the other (f) is suspected of being an F-bacterial DNA sequence unstably integrated into the host genome. In most wild populations harboring PSF, all individuals are genetic males (ZZ), and female phenotypes occur only due to the presence of PSF which overrides the male determinant carried by the Z chromosome (females are thus ZZ +F or ZZ +f neo-females). Here we report the effects of the conflict between these PSF and a dominant autosomal masculinizing gene (M) on phenotypes. The M gene is able to override the feminizing effect of the f sex factor and, consequently, male sex may be restored. However, M is unable to restore male sex when competing with the F bacteria. It seems that the main effect of M is to delay the expression of F bacteria slightly, inducing intersex phenotypes. Most of these intersexes are functional females, able to transmit the masculinizing gene. The frequency of M and its effects on the sex ratio in wild populations are discussed.

Regeneration of antennae and its control of molting in the terrestrial isopod, Armadillidium vulgare

AbstractThe interactions of regeneration and pregnancy controls on the duration of different stages of molting in the pillbug Armadillidium vulgare were determined by removing the antennae during different stages of the molt cycle of the nonovigerous and ovigerous animals. Removal of one or two antennae from the males during the beginning of stage C (intermolt period) of their molting cycle resulted in regeneration and shortening of stage C and thereby in acceleration of ecdysis. Further, removal of both antennae induced the animals to ecdyse within 1–4 days of each other, thus providing a convenient method of synchronizing the molt cycle. Amputation of both antennae from males in late stage C also resulted in regeneration; however, the duration of stage C was lengthened, resulting in a delay in ecdysis. Amputation of both antennae from males and nonovigerous females in the middle of preecdysis (stage D) resulted in molting without regeneration and on schedule like the control animals. However, the antennaless animals regenerated their antennae, shortened their intermolt period (stage C), and ecdysed precociously. From these results, we conclude that amputation, before the critical period in the molt cycle of the pillbugs (during stage C), prevents the release of ecdysteroids and, consequently, generalized apolysis until regeneration is completed, whereupon this block is removed and generalized apolysis and further events of molting occur. Once ecdysteroid release has been initiated to cause generalized apolysis, as in stage D, amputation does not prevent the further events of molting. Amputation of both antennae during the beginning of the intermolt period in the ovigerous females, although permitting regeneration, did not immediately stimulate ecdysis as in the nonovigerous animals; instead, it occurred only after release of the larvae from the brood pouch. When compared to the control ovigerous animals, the overall length of stage C was shortened. The results obtained from the antennal amputations during different physiological stages of the pillbug show that of all the stages of the molt cycle, stage C is labile, and its duration is affected by regeneration and pregnancy.

A Parataeniella (Trichomycetes, Eccrinales) in an Isopod

SUMMARYA new species of the fungus order Eccrinales (Trichomycetes), Parataeniella armadillidii, is described and illustrated, attached to the chitinous lining of the hindgut of the pillbugs Armadillidium vulgare and A. nasatum (Isopoda) in Kansas. This is the first species of Parataeniella reported outside of Europe. A significant feature of the genus is the conversion of the entire unbranched and nonseptate thallus into a single sporangium producing numerous uninucleate spores. In this respect Parataeniella resembles the ectocommensal Trichomycete genus Amoebidium (Amoebidiales), but Parataeniella produces no amoeboid cells and has other reproductive features characteristic of the Eccrinales.

A Parataeniella (Trichomycetes, Eccrinales) in an Isopod

SUMMARYA new species of the fungus order Eccrinales (Trichomycetes), Parataeniella armadillidii, is described and illustrated, attached to the chitinous lining of the hindgut of the pillbugs Armadillidium vulgare and A. nasatum (Isopoda) in Kansas. This is the first species of Parataeniella reported outside of Europe. A significant feature of the genus is the conversion of the entire unbranched and nonseptate thallus into a single sporangium producing numerous uninucleate spores. In this respect Parataeniella resembles the ectocommensal Trichomycete genus Amoebidium (Amoebidiales), but Parataeniella produces no amoeboid cells and has other reproductive features characteristic of the Eccrinales.