Jessica Dittmer

Host tissues as microhabitats for Wolbachia and quantitative insights into the bacterial community in terrestrial isopods

Animal–bacterial symbioses are highly dynamic in terms of multipartite interactions, both between the host and its symbionts as well as between the different bacteria constituting the symbiotic community. These interactions will be reflected by the titres of the individual bacterial taxa, for example via host regulation of bacterial loads or competition for resources between symbionts. Moreover, different host tissues represent heterogeneous microhabitats for bacteria, meaning that host‐associated bacteria might establish tissue‐specific bacterial communities. Wolbachia are widespread endosymbiotic bacteria, infecting a large number of arthropods and filarial nematodes. However, relatively little is known regarding direct interactions between Wolbachia and other bacteria. This study represents the first quantitative investigation of tissue‐specific Wolbachia–microbiota interactions in the terrestrial isopod Armadillidium vulgare. To this end, we obtained a more complete picture of the Wolbachia distribution patterns across all major host tissues, integrating all three feminizing Wolbachia strains (wVulM, wVulC, wVulP) identified to date in this host. Interestingly, the different Wolbachia strains exhibited strain‐specific tissue distribution patterns, with wVulM reaching lower titres in most tissues. These patterns were consistent across different host genetic backgrounds and might reflect different co‐evolutionary histories between the Wolbachia strains and A. vulgare. Moreover, Wolbachia‐infected females carried higher total bacterial loads in several, but not all, tissues, irrespective of the Wolbachia strain. Taken together, this quantitative approach indicates that Wolbachia is part of a potentially more diverse bacterial community, as exemplified by the presence of highly abundant bacterial taxa in the midgut caeca of several A. vulgare populations.

Phylogenomics of “Candidatus Hepatoplasma crinochetorum,” a Lineage of Mollicutes Associated with Noninsect Arthropods

Bacterial gut communities of arthropods are highly diverse and tightly related to host feeding habits. However, our understanding of the origin and role of the symbionts is often hindered by the lack of genetic information. “Candidatus Hepatoplasma crinochetorum” is a Mollicutes symbiont found in the midgut glands of terrestrial isopods. The only available nucleotide sequence for this symbiont is a partial 16S rRNA gene sequence. Here, we present the 657,101 bp assembled genome of Candidatus Hepatoplasma crinochetorum isolated from the terrestrial isopod Armadillidium vulgare. While previous 16S rRNA gene-based analyses have provided inconclusive results regarding the phylogenetic position of Candidatus Hepatoplasma crinochetorum within Mollicutes, we performed a phylogenomic analysis of 127 Mollicutes orthologous genes which confidently branches the species as a sister group to the Hominis group of Mycoplasma. Several genome properties of Candidatus Hepatoplasma crinochetorum are also highlighted compared with other Mollicutes genomes, including adjacent tryptophan tRNA genes, which further our understanding of the evolutionary dynamics of these genes in Mollicutes, and the presence of a probably inactivated CRISPR/Cas system, which constitutes a testimony of past interactions between Candidatus Hepatoplasma crinochetorum and mobile genetic elements, despite their current lack in this streamlined genome. Overall, the availability of the complete genome sequence of Candidatus Hepatoplasma crinochetorum paves the way for further investigation of its ecology and evolution.

The Terrestrial Isopod Microbiome: An All-in-One Toolbox for Animal–Microbe Interactions of Ecological Relevance

Bacterial symbionts represent essential drivers of arthropod ecology and evolution, influencing host traits such as nutrition, reproduction, immunity, and speciation. However, the majority of work on arthropod microbiota has been conducted in insects and more studies in non-model species across different ecological niches will be needed to complete our understanding of host–microbiota interactions. In this review, we present terrestrial isopod crustaceans as an emerging model organism to investigate symbiotic associations with potential relevance to ecosystem functioning. Terrestrial isopods comprise a group of crustaceans that have evolved a terrestrial lifestyle and represent keystone species in terrestrial ecosystems, contributing to the decomposition of organic matter and regulating the microbial food web. Since their nutrition is based on plant detritus, it has long been suspected that bacterial symbionts located in the digestive tissues might play an important role in host nutrition via the provisioning of digestive enzymes, thereby enabling the utilization of recalcitrant food compounds (e.g., cellulose or lignins). If this were the case, then (i) the acquisition of these bacteria might have been an important evolutionary prerequisite for the colonization of land by isopods, and (ii) these bacterial symbionts would directly mediate the role of their hosts in ecosystem functioning. Several bacterial symbionts have indeed been discovered in the midgut caeca of terrestrial isopods and some of them might be specific to this group of animals (i.e., Candidatus Hepatoplasma crinochetorum, Candidatus Hepatincola porcellionum, and Rhabdochlamydia porcellionis), while others are well-known intracellular pathogens (Rickettsiella spp.) or reproductive parasites (Wolbachia sp.). Moreover, a recent investigation of the microbiota in Armadillidium vulgare has revealed that this species harbors a highly diverse bacterial community which varies between host populations, suggesting an important share of environmental microbes in the host-associated microbiota. In this review, we synthesize our current knowledge on the terrestrial isopod microbiome and identify future directions to (i) fully understand the functional roles of particular bacteria (both intracellular or intestinal symbionts and environmental gut passengers), and (ii) whether and how the host-associated microbiota could influence the performance of terrestrial isopods as keystone species in soil ecosystems.

Host origin and tissue microhabitat shaping the microbiota of the terrestrial isopod Armadillidium vulgare

We present the first in-depth investigation of the host-associated microbiota of the terrestrial isopod crustacean Armadillidium vulgare. This species is an important decomposer of organic matter in terrestrial ecosystems and a major model organism for arthropod-Wolbachia symbioses due to its well-characterized association with feminizing Wolbachia 16S rRNA gene pyrotags were used to characterize its bacterial microbiota at multiple levels: (i) in individuals from laboratory lineages and field populations and (ii) in various host tissues. This integrative approach allowed us to reveal an unexpectedly high bacterial diversity, placing this species in the same league as termites in terms of symbiotic diversity. Interestingly, both animal groups belong to the same ecological guild in terrestrial ecosystems. While Wolbachia represented the predominant taxon in infected individuals, it was not the only major player. Together, the most abundant taxa represented a large scope of symbiotic interactions, including bacterial pathogens, a reproductive parasite (Wolbachia) and potential nutritional symbionts. Furthermore, we demonstrate that individuals from different populations harboured distinct bacterial communities, indicating a strong link between the host-associated microbiota and environmental bacteria, possibly due to terrestrial isopod nutritional ecology. Overall, this work highlights the need for more studies of host-microbiota interactions and bacterial diversity in non-insect arthropods.

Influence of changing plant food sources on the gut microbiota of Saltmarsh Detritivores

Changes in agricultural land-use of saltmarshes along the German North Sea coast have favoured the succession of the marsh grass Elytrigia atherica over the long-established Spartina anglica. Consequently, E. atherica represents a potential food source of increasing importance for plant-feeding soil detritivores. Considering the importance of this ecological guild for decomposition processes and nutrient cycling, we focussed on two sympatric saltmarsh soil macrodetritivores and their associated gut microbiota to investigate how the digestive processes of these species may be affected by changing plant food sources. Using genetic fingerprints of partial 16S rRNA gene sequences, we analysed composition and diversity of the bacterial gut community in a diplopod and an amphipod crustacean in relation to different feeding regimes representing the natural vegetation changes. Effects of syntopy on the host-specific gut microbiota were also taken into account by feeding the two detritivore species either independently or on the same plant sample. Bacterial community composition was influenced by both the host species and the available plant food sources, but the latter had a stronger effect on microbial community structure. Furthermore, bacterial diversity was highest after feeding on a mixture of both plant species, regardless of the host species. The gut microbiota of these two detritivores can thus be expected to change along with the on-going succession at the plant community level in this environment. Cloning and sequencing of bacterial 16S rRNA gene fragments further indicated a host-related effect since the two detritivores differed in terms of predominant bacterial taxa: diplopods harboured mainly representatives of the phyla Bacteroidetes and Gammaproteobacteria. In contrast, the genus Vibrio was found for the amphipod host across all feeding conditions.

Disentangling a Holobiont – Recent Advances and Perspectives in Nasonia Wasps

The parasitoid wasp genus Nasonia (Hymenoptera: Chalcidoidea) is a well-established model organism for insect development, evolutionary genetics, speciation, and symbiosis. The host-microbiota assemblage which constitutes the Nasonia holobiont (a host together with all of its associated microbes) consists of viruses, two heritable bacterial symbionts and a bacterial community dominated in abundance by a few taxa in the gut. In the wild, all four Nasonia species are systematically infected with the obligate intracellular bacterium Wolbachia and can additionally be co-infected with Arsenophonus nasoniae. These two reproductive parasites have different transmission modes and host manipulations (cytoplasmic incompatibility vs. male-killing, respectively). Pioneering studies on Wolbachia in Nasonia demonstrated that closely related Nasonia species harbor multiple and mutually incompatible Wolbachia strains, resulting in strong symbiont-mediated reproductive barriers that evolved early in the speciation process. Moreover, research on host-symbiont interactions and speciation has recently broadened from its historical focus on heritable symbionts to the entire microbial community. In this context, each Nasonia species hosts a distinguishable community of gut bacteria that experiences a temporal succession during host development and members of this bacterial community cause strong hybrid lethality during larval development. In this review, we present the Nasonia species complex as a model system to experimentally investigate questions regarding: (i) the impact of different microbes, including (but not limited to) heritable endosymbionts, on the extended phenotype of the holobiont, (ii) the establishment and regulation of a species-specific microbiota, (iii) the role of the microbiota in speciation, and (iv) the resilience and adaptability of the microbiota in wild populations subjected to different environmental pressures. We discuss the potential for easy microbiota manipulations in Nasonia as a promising experimental approach to address these fundamental aspects.

Variation of parasite load and immune parameters in two species of New Zealand shore crabs.

While parasites are likely to encounter several potential intermediate hosts in natural communities, a parasite’s actual range of compatible hosts is limited by numerous biological factors ranging from behaviour to immunology. In crustaceans, two major components of immunity are haemocytes and the prophenoloxidase system involved in the melanisation of foreign particles. Here, we analysed metazoan parasite prevalence and loads in the two sympatric crab species Hemigrapsus crenulatus and Macrophthalmus hirtipes at two sites. In parallel, we analysed the variation in haemocyte concentration and amount of circulating phenoloxidase (PO) in the haemolymph of the same individuals in an attempt to (a) explain differences in parasite prevalence and loads in the two species at two sites and (b) assess the impact of parasites on these immune parameters. M. hirtipes harboured more parasites but also exhibited higher haemocyte concentrations than H. crenulatus independent of the study site. Thus, higher investment in haemocyte production for M. hirtipes does not seem to result in higher resistance to parasites. Analyses of variation in immune parameters for the two crab species between the two sites that differed in parasite prevalence showed common trends. (a) In general, haemocyte concentrations were higher at the site experiencing higher parasitic pressure while circulating PO activity was lower and (b) haemocyte concentrations were influenced by microphallid trematode metacercariae in individuals from the site with higher parasitic pressure. We suggest that the higher haemocyte concentrations observed in both crab species exposed to higher parasitic pressure may represent an adaptive response to the impact of parasites on this immune parameter.

Feminizing Wolbachia influence microbiota composition in the terrestrial isopod Armadillidium vulgare

Wolbachia are widespread heritable endosymbionts of arthropods notorious for their profound effects on host fitness as well as for providing protection against viruses and eukaryotic parasites, indicating that they can interact with other microorganisms sharing the same host environment. Using the terrestrial isopod crustacean Armadillidium vulgare, its highly diverse microbiota (>200 bacterial genera) and its three feminizing Wolbachia strains (wVulC, wVulM, wVulP) as a model system, the present study demonstrates that Wolbachia can even influence the composition of a diverse bacterial community under both laboratory and natural conditions. While host origin is the major determinant of the taxonomic composition of the microbiota in A. vulgare, Wolbachia infection affected both the presence and, more importantly, the abundance of many bacterial taxa within each host population, possibly due to competitive interactions. Moreover, different Wolbachia strains had different impacts on microbiota composition. As such, infection with wVulC affected a higher number of taxa than infection with wVulM, possibly due to intrinsic differences in virulence and titer between these two strains. In conclusion, this study shows that heritable endosymbionts such as Wolbachia can act as biotic factors shaping the microbiota of arthropods, with as yet unknown consequences on host fitness.

Phenotypic shift in Wolbachia virulence towards its native host across serial horizontal passages

Vertical transmission mode is predicted to decrease the virulence of symbionts. However, Wolbachia, a widespread vertically transmitted endosymbiont, exhibits both negative and beneficial effects on arthropod fitness. This ‘Jekyll and Hyde’ behaviour, as well as its ability to live transiently outside host cells and to establish new infections via horizontal transmission, may reflect the capacity of Wolbachia to exhibit various phenotypes depending on the prevailing environmental constraints. To study the ability of Wolbachia to readily cope with new constraints, we forced this endosymbiont to spread only via horizontal transmission. To achieve this, we performed serial horizontal transfers of haemolymph from Wolbachia-infected to naive individuals of the isopod Armadillidium vulgare. Across passages, we observed phenotypic changes in the symbiotic relationship: (i) The Wolbachia titre increased in both haemolymph and nerve cord but remained stable in ovaries; (ii) Wolbachia infection was benign at the beginning of the experiment, but highly virulent, killing most hosts after only a few passages. Such a phenotypic shift after recurrent horizontal passages demonstrates that Wolbachia can rapidly change its virulence when facing new environmental constraints. We thoroughly discuss the potential mechanism(s) underlying this phenotypic change, which are likely to be crucial for the ongoing radiation of Wolbachia in arthropods.

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.