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Microorganisms often live in symbiosis with their hosts, and some are considered mutualists, where all species involved benefit from the interaction. How free-living microorganisms have evolved to become mutualists is unclear. Here we report an experimental system in which non-symbiotic Escherichia coli evolves into an insect mutualist. The stinkbug Plautia stali is typically associated with its essential gut symbiont, Pantoea sp., which colonizes a specialized symbiotic organ. When sterilized newborn nymphs were infected with E. coli rather than Pantoea sp., only a few insects survived, in which E. coli exhibited specific localization to the symbiotic organ and vertical transmission to the offspring. Through transgenerational maintenance with P. stali, several hypermutating E. coli lines independently evolved to support the host’s high adult emergence and improved body colour; these were called ‘mutualistic’ E. coli. These mutants exhibited slower bacterial growth, smaller size, loss of flagellar motility and lack of an extracellular matrix. Transcriptomic and genomic analyses of ‘mutualistic’ E. coli lines revealed independent mutations that disrupted the carbon catabolite repression global transcriptional regulator system. Each mutation reproduced the mutualistic phenotypes when introduced into wild-type E. coli, confirming that single carbon catabolite repression mutations can make E. coli an insect mutualist. These findings provide an experimental system for future work on host–microbe symbioses and may explain why microbial mutualisms are omnipresent in nature.

Eusocial insects, such as ants and honeybees, are characterized by the sophisticated division of labor among workers. In addition to age polyethism, under fluctuating environmental conditions, division of labor is maintained by task shifting of workers from one task to another if there are insufficient workers performing a certain task. Although several studies show that workers can perform task-shifting in many hymenopteran species, the physiological state of task-shifted workers is largely unknown without a few species. In this study, we examined the relationship between the ovarian development of workers and their task shifting using the queenless ant Diacamma cf. indicum from Japan. Our results showed that most of the nurses maintained their reproductive potential—as evidenced by their well-developed ovaries—even when they had shifted to the foraging task. In contrast, the foragers—whether they had shifted to the nursing task or not—had regressed ovaries. Combined with our previous study, the reproductive potential of workers mediates different effect of task experience on task shifting between nurses and foragers. Furthermore, our study highlights the importance of physiological heterogeneity underlying self-organized systems in insect societies.

Caste development in social insects requires the coordination of molting and metamorphosis during postembryonic development. In termites, i.e., hemimetabolous eusocial insects, caste fate is determined during postembryonic development. However, it is not fully understood how the mechanisms of molting/metamorphosis are regulated in the course of differentiation between reproductive and sterile castes. In termites, only reproductives derived from alates are imagos and other sterile castes (including developmentally-terminal soldier caste) are basically juveniles or nymphs. Furthermore, supplementary reproductives that appear when the original queens and kings die or become senescent, exhibit larval features such as winglessness, and are called neotenics. Therefore, the question of whether neotenics are larvae or imagos is still under debate. In this study, by inducing female neotenic differentiation in a damp-wood termite Hodotermopsis sjostedti, morphological investigations together with juvenile hormone (JH) quantification and expression/functional analyses of genes responsible for molting and/or metamorphosis were carried out. JH titer and expression of one of the downstream genes (Kr-h1) were shown to be temporarily lowered, but increased just prior to the molt into neotenics, while consistently lowered in imaginal molt (i.e., alate differentiation). In contrast, ecdysone-related genes (EcR and E93) were upregulated at both neotenic and alate differentiation, suggesting that the heterochronic actions of ecdysone and JH lead the neotenic differentiation. Moreover, expression analyses, supported by reverse genetic experiments, showed that EcR and E93 were specifically upregulated in genital sternites (EcR and E93) and ovaries (E93) and required for the development of imaginal characters. These results suggest that the resultant mosaic phenotype of female neotenics is due to modular responses of different body parts to hormonal actions.

Cephalopods have acquired numerous novelties and expanded their habitats to various marine environments as highly agile predators. Among cephalopod novelties, multiple arms are used for complex behaviors, including prey capture. Suckers on arms are innovative features for realizing these arm functions. In addition, tentacles in Decapodiformes (squids and cuttlefishes) are arms specialized in prey capture and tentacular suckers show unique morphologies. However, little is known about the developmental process of sucker formation that should differ between tentacles and other arms. In this study, therefore, sucker formation processes on second arms and tentacles were observed and compared in a bigfin reef squid, Sepioteuthis lessoniana, to reveal the developmental processes forming the unique sucker morphologies, especially in tentacles. Morphological and histological observations of suckers during embryogenesis showed that, in second arms, the sucker-producing area appeared at the most distal part. At the most proximal side of the sucker-producing area, new sucker buds were isolated by invagination of the epithelial tissue. At the proximal arm parts, suckers with functional structures were observed. In tentacles, although the basic sucker formation pattern was similar to that in second arms, sucker formation started at earlier embryonic stages and the number of suckers was drastically increased compared to that in second arms. In addition, although 4 sucker rows were observed at the tentacular club, i.e., the thickest part of a tentacle, our observations suggested that 2 sets of 2 sucker rows are compressed to form the 4 rows. Therefore, the sucker-formation processes are temporally and spatially different between arms and tentacles. In addition, S. lessoniana shows conserved and unique patterns of sucker formation in comparison with previously described species, suggesting that sucker formation patterns were diversified among Decapodiformes lineages. This article is protected by copyright. All rights reserved.

Among over 20,000 species of Annelida, only two branching species with a highly modified body-pattern are known until now: the Syllidae Syllis ramosa McIntosh, 1879, and Ramisyllis multicaudata Glasby et al. (Zoological Journal of the Linnean Society, 164, 481–497, 2012). Both have unusual ramified bodies with one head and multiple branches and live inside the canals of host sponges. Using an integrative approach (combining morphology, internal anatomy, ecology, phylogeny, genetic divergence, and the complete mitochondrial genome), we describe a new branching species from Japan, Ramisyllis kingghidorahi n. sp., inhabiting an undescribed species of Petrosia (Porifera: Demospongiae) from shallow waters. We compare the new species with its closest relative, R. multicaudata; emend the diagnosis of Ramisyllis; and discuss previous reports of S. ramosa. This study suggests a much higher diversity of branching syllids than currently known. Finally, we discuss possible explanations for the feeding behaviour in the new species in relation to its highly ciliated wall of the digestive tubes (especially at the distal branches and anus), and provide a hypothesis for the evolution of branching body patterns as the result of an adaptation to the host sponge labyrinthic canal system.

Significance Gene duplication is a major source of evolutionary innovation and is associated with the increases in biological complexity and adaptive radiation. Termites are model social organisms characterized by a sophisticated caste system. We analyzed the genome of the Japanese subterranean termite, an ecologically and economically important insect acting as a destructive pest. The analyses revealed the significance of gene duplication in social evolution. Gene duplication associated with caste-biased gene expression was prevalent in the termite genome. Many of the duplicated genes were related to social functions, such as chemical communication, social immunity, and defense, and they were often expressed in caste-specific organs. We propose that gene duplication facilitates social evolution through regulatory diversification leading to caste-biased expression and functional specialization.

Echinoderms constitute an animal phylum characterized by the pentaradial body plan. During the development from bilateral larvae to pentaradial adults, the formation of the multiple of five hydrocoel lobes, i.e., the buddings from the mesodermal coelom, is the firstly emerging pentameral character. The developmental mechanism underlying the hydrocoel-lobe formation should be revealed to understand the evolutionary process of this unique and highly derived body plan of echinoderms, although the morphogenetic mechanisms of hydrocoel lobes are largely uninvestigated. In this study, using the sea cucumber Apostichopus japonicus, in which hydrocoel is easily observable, the developmental process of hydrocoel lobes was described in detail, focusing on cell proliferation and rearrangement. Cell proliferation was not specifically distributed in the growing tips of the hydrocoel lobes, and inhibition of cell proliferation did not affect lobe formation. During lobe formation, the epithelium of the hydrocoel lobes was firstly thickened and then transformed into a simple epithelium, suggesting that tissue expansion via tissue remodeling contributes to the hydrocoel-lobe formation.

Significance Gene duplication is a major source of evolutionary innovation and is associated with the increases in biological complexity and adaptive radiation. Termites are model social organisms characterized by a sophisticated caste system. We analyzed the genome of the Japanese subterranean termite, an ecologically and economically important insect acting as a destructive pest. The analyses revealed the significance of gene duplication in social evolution. Gene duplication associated with caste-biased gene expression was prevalent in the termite genome. Many of the duplicated genes were related to social functions, such as chemical communication, social immunity, and defense, and they were often expressed in caste-specific organs. We propose that gene duplication facilitates social evolution through regulatory diversification leading to caste-biased expression and functional specialization., Termites are model social organisms characterized by a polyphenic caste system. Subterranean termites (Rhinotermitidae) are ecologically and economically important species, including acting as destructive pests. Rhinotermitidae occupies an important evolutionary position within the clade representing a transitional taxon between the higher (Termitidae) and lower (other families) termites. Here, we report the genome, transcriptome, and methylome of the Japanese subterranean termite Reticulitermes speratus. Our analyses highlight the significance of gene duplication in social evolution in this termite. Gene duplication associated with caste-biased gene expression was prevalent in the R. speratus genome. The duplicated genes comprised diverse categories related to social functions, including lipocalins (chemical communication), cellulases (wood digestion and social interaction), lysozymes (social immunity), geranylgeranyl diphosphate synthase (social defense), and a novel class of termite lineage–specific genes with unknown functions. Paralogous genes were often observed in tandem in the genome, but their expression patterns were highly variable, exhibiting caste biases. Some of the assayed duplicated genes were expressed in caste-specific organs, such as the accessory glands of the queen ovary and the frontal glands of soldier heads. We propose that gene duplication facilitates social evolution through regulatory diversification, leading to caste-biased expression and subfunctionalization and/or neofunctionalization conferring caste-specialized functions.

Although many animals that perform sexual reproduction exhibit sexual dimorphism, individuals with intersex traits between the traits of males and females appear in some species, depending on environmental factors. Ptychognathus ishii, a varunid crab, exhibits distinctive sexual dimorphism in the morphology of its abdomen, chelipeds and setal tufts on the chelipeds. In this study, however, we report for the first time that intersex individuals with intermediate characters between those of males and females were occasionally found in wild populations. Morphological features of intersex individuals are described. Their taxonomic positions are identified based on DNA sequences of part of the mitochondrial cytochrome c oxidase I (COI) gene. It was shown that the intersexuality was induced by entoniscid parasites, because all intersex individuals were parasitized by entoniscid isopods, identified as Entionella sp. The apparent correlation between parasitism and morphological anomalies suggests that the parasitic isopods affect physiological conditions, leading to the feminization of male hosts.

Four new species of annelids

SummaryTermites are model social organisms characterized by a polyphenic caste system. Subterranean termites (Rhinotermitidae) are ecologically and economically important species, including acting as destructive pests. Rhinotermitidae occupies an important evolutionary position within the clade representing an intermediate taxon between the higher (Termitidae) and lower (other families) termites. Here, we report the genome, transcriptome and methylome of the Japanese subterranean termite Reticulitermes speratus. The analyses highlight the significance of gene duplication in social evolution in this termite. Gene duplication associated with caste-biased gene expression is prevalent in the R. speratus genome. Such duplicated genes encompass diverse categories related to social functions, including lipocalins (chemical communication), cellulases (wood digestion and social interaction), lysozymes (social immunity), geranylgeranyl diphosphate synthase (social defense) and a novel class of termite lineage-specific genes with unknown functions. Paralogous genes were often observed in tandem in the genome, but the expression patterns were highly variable, exhibiting caste biases. Some duplicated genes assayed were expressed in caste-specific organs, such as the accessory glands of the queen ovary and frontal glands in soldier heads. We propose that gene duplication facilitates social evolution through regulatory diversification leading to caste-biased expression and subfunctionalization and/or neofunctionalization that confers caste-specialized functions.Significance StatementTermites are model social organisms characterized by a sophisticated caste system, where distinct castes arise from the same genome. Our genomics data of Japanese subterranean termite provides insights into the evolution of the social system, highlighting the significance of gene duplication. Gene duplication associated with caste-biased gene expression is prevalent in the termite genome. Many of the duplicated genes were related to social functions, such as chemical communication, social immunity and defense, and they often expressed in caste-specific organs. We propose that gene duplication facilitates social evolution through regulatory diversification leading to caste-biased expression and functional specialization. In addition, since subterranean termites are ecologically and economically important species including destructive pests in the world, our genomics data serves as a foundation for these studies.

Although many animals that perform sexual reproduction exhibit sexual dimorphism, individuals with intersex traits between the traits of males and females appear in some species, depending on environmental factors.

In bryozoans (phylum Bryozoa), representative colonial animals mostly found in marine environments, some species possess different types of individuals (heterozooids) specialized in different functions such as defense or structural support for their colonies. Among them, the best-known heterozooids are the avicularia, known to function as defenders. The differentiation processes of heterozooids, including avicularia, should be important keys to understand the evolutionary significance of bryozoans. However, the developmental process of avicularium formation remains to be fully understood. In this study, therefore, in order to understand the detailed developmental process and timing of avicularium formation, extensive observations were carried out in a bryozoan species, Bugulina californica (Cheilostomata, Bugulidae), that possesses adventitious avicularia, by performing stereomicroscopy on live materials, in addition to scanning electron microscopy and histological observations. The whole process can be divided into seven stages based on developmental events. Especially notably, at the earlier stages, there are three major budding events that produce proliferating cell masses corresponding to primordial tissues: (1) budding of the peduncle cushion at the outer margin of the distal part of a young autozooid, (2) budding of the head-part primordium from the peduncle cushion, and (3) budding of the polypide inside the head part. Experimental control of temperature showed that 20°C would be the best to maintain B. californica colonies.

In bryozoans (phylum Bryozoa), representative colonial animals mostly found in marine environments, some species possess different types of individuals (heterozooids) specialized in different functions such as defense or structural support for their colonies. Among them, the best-known heterozooids are the avicularia, known to function as defenders. The differentiation processes of heterozooids, including avicularia, should be important keys to understand the evolutionary significance of bryozoans. However, the developmental process of avicularium formation remains to be fully understood. In this study, therefore, in order to understand the detailed developmental process and timing of avicularium formation, extensive observations were carried out in a bryozoan species

Background Morphological novelties have been acquired through evolutionary processes and related to the adaptation of new life-history strategies with new functions of the bodyparts. Cephalopod molluscs such as octopuses, squids and cuttlefishes possess unique morphological characteristics. Among those novel morphologies, in particular, suckers arranged along the oral side of each arm possess multiple functions, such as capturing prey and locomotion, so that the sucker morphology is diversified among species, depending on their ecological niche. However, the detailed developmental process of sucker formation has remained unclear, although it is known that new suckers are formed or added during both embryonic and postembryonic development. In the present study, therefore, focusing on two cuttlefish species, Sepia esculenta and S. lycidas, in which the sucker morphology is relatively simple, morphological and histological observations were carried out during embryonic and postembryonic development to elucidate the developmental process of sucker formation and to compare them among other cephalopod species. Results The observations in both species clearly showed that the newly formed suckers were added on the oral side of the most distal tip of each arm during embryonic and postembryonic development. On the oral side of the arm tip, the epithelial tissue became swollen to form a ridge along the proximal-distal axis (sucker field ridge). Next to the sucker field ridge, there were small dome-shaped bulges that are presumed to be the sucker buds. Toward the proximal direction, the buds became functional suckers, in which the inner tissues differentiated to form the complex sucker structures. During postembryonic development, on both sides of the sucker field ridge, epithelial tissues extended to form a sheath, covering the ridge for protection of undifferentiated suckers. Conclusions The developmental process of sucker formation, in which sucker buds are generated from a ridge structure (sucker field ridge) on the oral side at the distal-most arm tip, was shared in both cuttlefish species, although some minor heterochronic shifts of the developmental events were detected between the two species.

Eusocial insects exhibit reproductive division of labor, in which only a fraction of colony members differentiate into reproductives. In termites, reproductives of both sexes are present in a colony and constantly engaged in reproduction. It has been suggested that the sex ratio of reproductives is maintained by social interactions. The presence of reproductives is known to inhibit the additional differentiation of same-sex reproductives, while it promotes the differentiation of opposite-sex reproductives. In this study, using the damp-wood termite Hodotermopsis sjostedti, physiological effects of male/female reproductives on the differentiation of supplementary reproductives (neotenics) were examined. The results showed that the only male-neotenic condition, i.e., the presence of male neotenics in the absence of female neotenics, accelerated the neotenic differentiation from female workers (i.e., pseudergates). Under this condition, the rise of juvenile hormone (JH) titer was repressed in females, and the application of a JH analog inhibited the female neotenic differentiation, indicating that the low JH titer leads to rapid differentiation. Thus, the only male-neotenic condition that actively promotes reproductive differentiation by manipulating physiological condition of females is suggested to be a mechanism underlying sexual asymmetry in reproductive function, which may lead the female-biased sex allocation of reproductives.

In cooperative societies such as those of ants, honey bees, and termites, the number of reproductives is often regulated by social interactions. In many termite species, helper individuals (i.e., larvae or workers) can potentially differentiate into a “neotenic” reproductive caste in the absence of reproductives. In some termite species, multiple neotenics coexist within a nest, often with female-biased sex ratios. However, although the presence of female neotenics can suppress neotenic differentiation of female workers, it is largely unknown how male neotenics affect the differentiation of female neotenics. Here, we show that male and female neotenics regulate the neotenic differentiation in a sex-specific manner in the damp-wood termite Hodotermopsis sjostedti, whose colonies are often headed by multiple male and female neotenics in the field. Our rearing experiments showed that the presence of female neotenics suppressed differentiation of female neotenics from fourth- to seventh-larvae, i.e., pseudergates (called as “workers” in this study), whereas male neotenics promoted the differentiation of female neotenics. Moreover, the results of rearing experiments that restricted physical contact between neotenics and workers suggested that these effects were not mediated by volatile chemicals, but rather by direct contact. We found that the male neotenics were frequently groomed by female workers, suggesting that these interactions promote the differentiation of female neotenics. Our results represent an empirical evidence that the neotenic differentiation from female and male workers is regulated by direct physical contact with the preexisting neotenics.

Background Generally in termites, alates differentiate through multiple nymphal instars which gradually develop wing buds. However, in a dampwood termite, Hodotermopsis sjostedti, alates molt directly from a single nymphal instar with short wing buds. In this study, to examine the mechanism underlying the wing formation during the alate differentiation in H. sjostedti, histological and morphological observations were carried out on the developmental process of wing formation during the nymphal instar, in comparison with those in Zootermopsis nevadensis, which has two nymphal instars. Furthermore, the expression patterns of genes that are thought to be responsible for wing formation, i.e., wing-patterning genes and genes encoding hormone-related factors, were quantified during alate differentiation and compared between the two species. Results The results showed that, in H. sjostedti, wings were formed in a complicatedly folded shape, not only inside the wing buds as seen in Z. nevadensis, but also under the dorsal thoracic cuticle, where the wing tips shifted toward the median thoracic part. Accordingly, the wing expansion pattern also differed from that in Z. nevadensis. Furthermore, the results of real-time qRT-PCR on overall expression profiles of wing-patterning genes and hormone-related genes suggest that the single nymphal instar in H. sjostedti well resembles to the second nymphal instar in Z. nevadensis. In particular, significant upregulation of vestigial (vg) and downregulation of Krüppel homolog 1 (Kr-h1) that were observed at the second nymphal instar in Z. nevadensis apparently occurred during the single nymphal instar in H. sjostedti. Conclusion The developmental events for wing formation are compacted into a single nymphal instar in H. sjostedti, and as a result, the unique wing formation is seen to compensate for the spatial restriction inside small wing buds, leading to the completion of functional wings. Electronic supplementary material The online version of this article (10.1186/s13227-019-0123-8) contains supplementary material, which is available to authorized users.

Some polychaete species in the family Syllidae exhibit distinctive life cycles, in which a posterior part of the body of an individual detaches as a reproductive individual called a "stolon". This type of reproductive mode is known as stolonization or schizogamy. Although a number of observations have been reported, and techniques using molecular markers have recently been applied to characterize this phenomenon, little is known about the developmental and physiological mechanisms underlying stolonization. In the present study

Eusocial insects organize their colonies based on division of labor and caste differentiation, in which caste-specific morphologies are sculpted during postembryonic development. In the differentiation between reproductive and sterile castes, characteristics related to mating and reproduction are developed in reproductives, and degenerated in neuters, although little is known about the developmental regulations during the differentiation. In some species of termites, a sensory protrusion at the posterior end of the abdomen, the stylus, is known to disappear in female reproductives. In the present study, we performed anatomical and histological analyses in the damp-wood termite Hodotermopsis sjostedti to elucidate the developmental process underlying the disappearance of the stylus during neotenic and alate differentiation. Although it was first hypothesized that styli were hidden beneath the enlarged seventh sternite, our observation results found out that the styli were completely lost in reproductive females. Further histological observations revealed that the stylus disappearance was not accomplished by degeneration process; rather, styli were separated from the abdomen and discarded with the exoskeleton (exuviae) during the molt into the reproductive caste. This phenomenon in which live tissues are discarded through developmental processes is suggested to be a case of abscission, as seen in plant leaves.

In social insects, interactions among colony members trigger caste differentiation with morphological modifications. In termite soldier differentiation, the mandible size considerably increases through two moltings (via the presoldier stage) under the control of juvenile hormone (JH). Regulatory genes are predicted to provide patterning information that induces the mandible-specific cell proliferation. To identify factors responsible for the mandibular enlargement, expression analyses of 18 candidate genes were carried out in the termite Hodotermopsis sjostedti. Among those, dachshund (dac), which identifies the intermediate domain along the proximodistal appendage axis, showed mandible-specific upregulation prior to the molt into presoldiers, which can explain the pattern of cell proliferation for the mandibular elongation. Knockdown of dac by RNAi reduced the mandibular length and distorted its morphology. Furthermore, the epistatic relationships among Methoprene tolerant, Insulin receptor, Deformed (Dfd) and dac were revealed by combined RNAi and qRT-PCR analyses, suggesting that dac is regulated by Dfd, downstream of the JH and insulin signaling pathways. Thus, caste-specific morphogenesis is controlled by interactions between the factors that provide spatial information and physiological status.

In social insects, interactions among colony members trigger caste differentiation with morphological modifications. In termite soldier differentiation, the mandible size considerably increases through two moltings (via the presoldier stage) under the control of juvenile hormone (JH). Regulatory genes are predicted to provide patterning information that induces the mandible-specific cell proliferation. To identify factors responsible for the mandibular enlargement, expression analyses of 18 candidate genes were carried out in the termite

Some polychaete species in the family Syllidae exhibit distinctive life cycles, in which a posterior part of the body of an individual detaches as a reproductive individual called a “stolon”. This type of reproductive mode is known as stolonization or schizogamy. Although a number of observations have been reported, and techniques using molecular markers have recently been applied to characterize this phenomenon, little is known about the developmental and physiological mechanisms underlying stolonization. In the present study, Megasyllis nipponica, a common syllid species distributed throughout Japan, is proposed as a model to reveal the developmental and physiological mechanism of stolonization, and the rearing system to maintain it in laboratory conditions is described. This species was repeatedly sampled around Hokkaido, where more dense populations were found from August to October. The animals were maintained in the laboratory under stable long-day condition (20°C, 16L:8D), and fed mainly with spinach powder. Stolonization processes, spawning, embryonic and postembryonic development were observed and documented, and the required period of time for each developmental stage was recorded. The complete generation time was around two months under the rearing condition. The information provided is valuable to maintain this and other syllid species in the laboratory, and hence contributes to the establishment of new evolutionary and developmental research lines in this group of annelids.