Eusocial Termites Research Articles

Pervasive relaxed selection in termite genomes.

Genetic changes that enabled the evolution of eusociality have long captivated biologists. More recently, attention has focussed on the consequences of eusociality on genome evolution. Studies have reported higher molecular evolutionary rates in eusocial hymenopteran insects compared with their solitary relatives. To investigate the genomic consequences of eusociality in termites, we analysed nine genomes, including newly sequenced genomes from three non-eusocial cockroaches. Using a phylogenomic approach, we found that termite genomes have experienced lower rates of synonymous substitutions than those of cockroaches, possibly as a result of longer generation times. We identified higher rates of non-synonymous substitutions in termite genomes than in cockroach genomes, and identified pervasive relaxed selection in the former (24-31% of the genes analysed) compared with the latter (2-4%). We infer that this is due to reductions in effective population size, rather than gene-specific effects (e.g. indirect selection of caste-biased genes). We found no obvious signature of increased genetic load in termites, and postulate efficient purging of deleterious alleles at the colony level. Additionally, we identified genomic adaptations that may underpin caste differentiation, such as genes involved in post-translational modifications. Our results provide insights into the evolution of termites and the genomic consequences of eusociality more broadly.

Different strategies between queens and workers against fungal pathogens in the termite Reticulitermes chinensis

AbstractSocial insects are prone to pathogen infection because of high exposure rates from social interactions. However, it remains unclear whether queens have enhanced pathogen resistance, because reproduction is largely confined to queens. Here, we used a natural host–pathogen system, the subterranean termite Reticulitermes chinensis and the entomopathogenic fungus Metarhizium anisopliae, to investigate the differences in allogrooming, locomotion, and immune gene expression between queens and workers against pathogen infection. We found that fungal infection significantly reduced survival in both queens and workers. Infected queens received significantly more grooming time from sanitary nestmates than infected workers, but they returned much less grooming time to sanitary nestmates than infected workers. Infection resulted in a reduction in the average locomotion speed and distance of queens but had no effect on worker locomotion. Infection resulted in upregulated expression of two immune genes (termicin and transferrin), two antioxidant genes (CAT and SOD), and phosphate genes CYP450 in queens but not in workers. Our results indicated that eusocial termites evolved strategies that prioritize the reproductive castes' welfare in defending against the pathogen infection to ensure continued reproduction and colony persistence.

Major changes in domain arrangements are associated with the evolution of termites.

Domains as functional protein units and their rearrangements along the phylogeny can shed light on the functional changes of proteomes associated with the evolution of complex traits like eusociality. This complex trait is associated with sterile soldiers and workers, and long-lived, highly fecund reproductives. Unlike in Hymenoptera (ants, bees, and wasps), the evolution of eusociality within Blattodea, where termites evolved from within cockroaches, was accompanied by a reduction in proteome size, raising the question of whether functional novelty was achieved with existing rather than novel proteins. To address this, we investigated the role of domain rearrangements during the evolution of termite eusociality. Analysing domain rearrangements in the proteomes of three solitary cockroaches and five eusocial termites, we inferred more than 5,000 rearrangements over the phylogeny of Blattodea. The 90 novel domain arrangements that emerged at the origin of termites were enriched for several functions related to longevity, such as protein homeostasis, DNA repair, mitochondrial activity, and nutrient sensing. Many domain rearrangements were related to changes in developmental pathways, important for the emergence of novel castes. Along with the elaboration of social complexity, including permanently sterile workers and larger, foraging colonies, we found 110 further domain arrangements with functions related to protein glycosylation and ion transport. We found an enrichment of caste-biased expression and splicing within rearranged genes, highlighting their importance for the evolution of castes. Furthermore, we found increased levels of DNA methylation among rearranged compared to non-rearranged genes suggesting fundamental differences in their regulation. Our findings indicate the importance of domain rearrangements in the generation of functional novelty necessary for termite eusociality to evolve.

Alarm communication predates eusociality in termites

Termites (Blattodea: Isoptera) have evolved specialized defensive strategies for colony protection. Alarm communication enables workers to escape threats while soldiers are recruited to the source of disturbance. Here, we study the vibroacoustic and chemical alarm communication in the wood roach Cryptocercus and in 20 termite species including seven of the nine termite families, all life-types, and all feeding and nesting habits. Our multidisciplinary approach shows that vibratory alarm signals represent an ethological synapomorphy of termites and Cryptocercus. In contrast, chemical alarms have evolved independently in several cockroach groups and at least twice in termites. Vibroacoustic alarm signaling patterns are the most complex in Neoisoptera, in which they are often combined with chemical signals. The alarm characters correlate to phylogenetic position, food type and hardness, foraging area size, and nesting habits. Overall, species of Neoisoptera have developed the most sophisticated communication system amongst termites, potentially contributing to their ecological success.

Same-sex Pairs Retain Their Reproductive Capacity as a Potential Opportunity for Individual Reproductive Success in Termites

In eusocial termites, successful pairing is an essential element of dispersal and distribution after the departure of alates from natal colonies. Two situations could arise during the pairing process: mixed-sex pairs and same-sex pairs. However, most previous studies focused on mixed-sex pairs, overlooking groups formed by same-sex pairings, especially potential fecundity (the total number of oocytes or ovarioles), oogenesis and the development stage of oocytes of females in female-female pairs, and spermatogenesis and testis development of males in male-male pairs. In this study, through experimentation, we investigated the reproductive ability of virgin dealates based on various pairing types as mentioned above. We found that the life spans of virgin dealates can cover 1 yr or even more when they establish a nest with a partner, which is more than 10-fold longer than the life span of individuals establishing a colony alone. After 1 yr of pairing, the potential fecundity of virgin same sex dealates did not degenerate significantly compared with newly emerged dealates, including the number of ovarioles, size of testis, oogenesis, and the development stage of the oocytes. Moreover, when individuals of same-sex pairings experimentally changed into mixed-sex pairs after 1 yr, the eggs produced in the colony hatched into larvae. These findings suggest that dealates which through same-sex pairs retain fecundity after 1 yr have more reproductive potential than dealates that failed to pair with heterosexuals, shedding light on the ecological significance of homosexual behaviors in terms of the successful extension and fecundity of eusocial termites.

Lifespan prolonging mechanisms and insulin upregulation without fat accumulation in long-lived reproductives of a higher termite

Kings and queens of eusocial termites can live for decades, while queens sustain a nearly maximal fertility. To investigate the molecular mechanisms underlying their long lifespan, we carried out transcriptomics, lipidomics and metabolomics in Macrotermes natalensis on sterile short-lived workers, long-lived kings and five stages spanning twenty years of adult queen maturation. Reproductives share gene expression differences from workers in agreement with a reduction of several aging-related processes, involving upregulation of DNA damage repair and mitochondrial functions. Anti-oxidant gene expression is downregulated, while peroxidability of membranes in queens decreases. Against expectations, we observed an upregulated gene expression in fat bodies of reproductives of several components of the IIS pathway, including an insulin-like peptide, Ilp9. This pattern does not lead to deleterious fat storage in physogastric queens, while simple sugars dominate in their hemolymph and large amounts of resources are allocated towards oogenesis. Our findings support the notion that all processes causing aging need to be addressed simultaneously in order to prevent it.

Termite eusociality and contrasting selective pressure on social and innate immunity

Termite eusociality and contrasting selective pressure on social and innate immunity

Extra-pair paternity in the wood-feeding cockroach Cryptocercus punctulatus Scudder: Social but not genetic monogamy.

Subsocial Cryptocercus cockroaches are the sister group to termites and considered to be socially monogamous. Because genetic monogamy is a suggested requirement for evolution of cooperative breeding/eusociality, particularly in hymenopterans, clarification of the mating biology of Cryptocercus would help illuminate evolutionary trends in eusocial insects. To investigate possible extra-pair paternity in C.punctulatus, microsatellite markers were used to analyse offspring parentage, the stored sperm in females and results of experimental manipulation of sperm competition. Extra-pair paternity was common in field-collected families, but a lack of maternal alleles in several nymphs suggests sampling error or adoption. Isolating prereproductive pairs and assaying subsequently produced nymphs confirmed that nymphs lacked alleles from the pair male in 40% of families, with extra-pair male(s) siring 27%-77% of nymphs. Sperm of extra-pair males was detected in the spermatheca of 51% of paired prereproductive females. Mate switching and surgical manipulation of male mating ability indicated a tendency towards last male sperm precedence. Overall, the results demonstrate that about half of young females are serially monogamous during their maturational year, but bond, overwinter and produce their only set of offspring in company of the last mated male (=pair male). Repeated mating by the pair male increases the number of nymphs sired, but because many females use stored sperm of previous copulatory partners to fertilize eggs, pair males extend parental care to unrelated nymphs. The results suggest that genetic monogamy either developed in the termite ancestor after splitting from the Cryptocercus lineage, or that genetic monogamy may not be a strict prerequisite for the evolution of termite eusociality.

Revising dating estimates and the antiquity of eusociality in termites using the fossilized birth–death process

AbstractDeciphering the timing and tempo of lineage diversification of organisms has greatly benefited from advances in Bayesian phylogenetic analyses using morphological data. Those advances, however, have not been used for termites despite a rich fossil record. Here, we estimate divergence times for living and fossil termites using the fossilized birth–death (FBD) process on a previously published morphological matrix expanded with two new fossils that we describe (see Appendices S1 and S2). Those fossils, based on soldier specimens, are the mid‐Cretaceous mastotermitid Milesitermes engeli gen. et sp. nov., and the Middle Eocene Reticulitermes grimaldii sp. nov. The latter is the oldest occurrence of a Rhinotermitidae soldier and the first termite soldier described from Baltic amber. Our dating estimates provide new stem‐ages and crown‐ages for termites, suggesting older ages than previously thought for several lineages. Importantly, crown‐Isoptera – and, therefore, eusociality – may have arisen approximately 200 Ma. We conclude with further directions to keep improving our understanding of the timing of differentiation in termites.

Biochemical, molecular, and morphological variations of flight muscles before and after dispersal flight in a eusocial termite, Reticulitermes chinensis.

Swarming behavior facilitates pair formation, and therefore mating, in many eusocial termites. However, the physiological adjustments and morphological transformations of the flight muscles involved in flying and flightless insect forms are still unclear. Here, we found that the dispersal flight of the eusocial termite Reticulitermes chinensis Snyder led to a gradual decrease in adenosine triphosphate supply from oxidative phosphorylation, as well as a reduction in the activities of critical mitochondrial respiratory enzymes from preflight to dealation. Correspondingly, using three-dimensional reconstruction and transmission electron microscopy (TEM), the flight muscles were found to be gradually deteriorated during this process. In particular, two tergo-pleural muscles (IItpm5 and III-tpm5) necessary to adjust the rotation of wings for wing shedding behavior were present only in flying alates. These findings suggest that flight muscle systems vary in function and morphology to facilitate the swarming flight procedure, which sheds light on the important role of swarming in successful extension and fecundity of eusocial termites.

Pathogenic Dynamics During Colony Ontogeny Reinforce Potential Drivers of Termite Eusociality: Mate Assistance and Biparental Care

As an ecologically dominant taxon, termites appear to be resilient to environmental stressors. However, swarming alates (winged-individuals) encounter a myriad of environmental pressures that drastically reduce the probability of colony foundation. Dispersing alates face high rates of predation, desiccation, nitrogen limitation, and risks of infection, among others. We propose that alates benefit from mate assistance and biparental care to overcome some of these challenges. We assessed whether the bacteria, Serratia marcescens (an ecologically relevant, gram-negative, facultative termite pathogen), negatively affected the growth of newly founded termite colonies. Additionally, we revealed the significance of the king’s presence in improving successful establishment of incipient colonies. Virgin queens of the dampwood termite, Zootermopsis angusticollis, were subjected to one of four treatments: naive (untreated), or injections with either sterile saline, heat-killed S. marcescens, or a sublethal does of live S. marcescens. Each queen was then paired with a naive, virgin king. The incipient colonies underwent censuses every four days for 80 days. We estimated survival rates and compared the onset of oviposition and hatching, overall egg production and larval hatching success, all as a function of queen treatment and the presence of a mate. We identified factors that, under pathogenic stress, influenced these fitness-related milestones. Queen infection significantly reduced the number of successfully established colonies. Moreover, both the presence of a king and his mass significantly influenced the queen’s survival, her onset of oviposition, overall egg production, and hatching success. We conclude that termite colonies incur significant fitness costs after a queen suffers an acute infection and that the presence of a king (and his stored resources) may help mitigate the negative effects of a queen’s infection. Pathogenic pressures, combined with the significant role of kings in colony success, appear to reinforce two-parent colony foundation, mate assistance, biparental care, and ultimately the overlap of generations, all of which have been considered preadaptations for eusociality. By studying the fitness consequences of pathogenic stress during the ontogeny of a termite colony, we can infer some of the conditions and pressures under which termite sociality likely emerged.

Transcriptomic and Functional Analyses of Phenotypic Plasticity in a Higher Termite, Macrotermes barneyi Light.

Eusocial termites have a complex caste system, which leads to the division of labor. Previous studies offered some insight into the caste differentiation in lower termites; however, few studies were focusing on the molecular mechanisms of higher termites with sophisticated societies. Comparative transcriptomic analyses of five immature castes of a higher termite, Macrotermes barneyi Light, suggest that phenotypic plasticity is modulated by an array of transcriptional changes, including differentially expressed genes (e.g., caste-biased genes Vtg and TnC), co-expression networks (e.g., genes associated with nymph reproduction), and alternative splicing (e.g., events related to muscle development in presoldiers). Transcriptional (RT-PCR and RT-qPCR) and functional (in vivo RNAi) validation studies reveal multiple molecular mechanisms contributing to the phenotypic plasticity in eusocial termites. Molecular mechanisms governing the phenotypic plasticity in M. barneyi could be a rule rather than an exception in the evolution of sociality.

Isocitrate dehydrogenase-mediated metabolic disorders disrupt active immunization against fungal pathogens in eusocial termites

Active immunization is a crucial colony-level pathogen defense to improve the survival of eusocial termites, which limits the lethal effect of biopesticides for termite control. Here, we report a possible strategy for RNAi-mediated metabolic disorders, which causes reduction in insect pathogen defense and might facilitate the foundation of nontoxic and sustainable pest control methods. Isocitrate dehydrogenase (IDH) was significantly upregulated by active immunization in termites, but its regulatory mechanism has yet to be well understood. We found that dsIDH-injected termites exhibited significantly reduced IDH at mRNA and protein levels and altered levels of isocitrate and NADH, indicating the impaired NAD+-IDH reaction. IDH-silenced termites displayed metabolic disorders, which was implied by significant changes in several metabolites from the carbohydrates and amino acids. When grooming toward fungus-exposed termites, IDH-silenced nestmates showed a significant upregulation of four apoptosis-related genes and caspase 3 activity, and hence the significantly increased rate of apoptosis. These physiological changes led to more apoptotic cell death in different body parts of the nestmates. In particular, more apoptotic cells increased disease susceptibility of the nestmates as revealed by growth of the significantly increased number of colony-forming units from dissected gut contents. Furthermore, the IDH-silenced nestmates exhibited lower antifungal activity and higher mortality, suggesting that IDH downregulation disrupted active immunization against fungal pathogens in termites. These findings illustrated the metabolic regulation of active immunization and provide a sound foundation for enhancing the impact of biopesticides by reducing the pathogen defense of hosts.

Caste-specific microRNA expression in termites: insights into soldier differentiation.

Eusocial insects have polyphenic caste systems in which each caste exhibits characteristic morphology and behaviour. In insects, caste systems arose independently in different lineages, such as Isoptera and Hymenoptera. Although partial molecular mechanisms for the development of eusociality in termites have been clarified by the functional analysis of genes and hormones, the contribution of microRNAs (miRNAs) to caste differentiation is unknown. To understand the role of miRNAs in termite caste polyphenism, we performed small RNA sequencing in a subterranean termite (Reticulitermes speratus) and identified the miRNAs that were specifically expressed in the soldier and worker castes. Of the 550 miRNAs annotated in the R.speratus genome, 74 were conserved in insects and 174 were conserved in other termite species. We found that eight miRNAs (mir-1, mir-125, mir-133, mir-2765, mir-87a and three termite-specific miRNAs) are differentially expressed (DE) in soldiers and workers of R.speratus. This differential expression was experimentally verified for five miRNAs by real-time quantitative PCR. Further, four of the eight DE miRNAs in soldier and worker termite castes were also differentially expressed in hymenopteran castes. The finding that Isoptera and Hymenoptera shared several DE miRNAs amongst castes suggests that these miRNAs evolved independently in these phylogenetically distinct lineages.

Reconstructed evolution of insulin receptors in insects reveals duplications in early insects and cockroaches.

Social insects show an extreme degree of phenotypic plasticity. In highly eusocial species, this manifests in the generation of distinct castes with extreme differences in both morphology and life span. The molecular basis of these differences is highly entangled and not fully understood, but several recent studies demonstrated that insulin/insulin-like growth factor signaling (IIS) is one of the key pathways. Here, we investigate the molecular evolution of insect insulin receptors (InRs), which are membrane-bound dimers that enable IIS by relaying extracellular signals to intracellular signaling cascades. Classic models of invertebrate IIS include only one InR gene, but some recent studies on less commonly studied insects have found two InRs, which act in an antagonistic manner to facilitate polyphenism in at least one documented case. We search 22 arthropod genomes and identify several InR copies and their evolutionary origin that were lacking from previous annotations. Phylogenetic analysis shows that the two insect InR genes date back at least 400 million years to a common ancestor of winged insects. Most notably, we also identified the evolutionary origin of a third InR copy that is unique to the clade of Blattodea, just before therein the eusocial termites evolved. One of the InR paralogs consistently shows caste-biased expression in all three termites, which strongly suggests a role in caste differentiation. These results have important ramifications for past and future InR inhibition/InR knockdown experiments in insects and they provide a set of key genes regulating life span and morphology in termite castes.

Remodeling of the juvenile hormone pathway through caste-biased gene expression and positive selection along a gradient of termite eusociality.

The evolution of division of labor between sterile and fertile individuals represents one of the major transitions in biological complexity. A fascinating gradient in eusociality evolved among the ancient hemimetabolous insects, ranging from noneusocial cockroaches through the primitively social lower termites-where workers retain the ability to reproduce-to the higher termites, characterized by lifetime commitment to worker sterility. Juvenile hormone (JH) is a prime candidate for the regulation of reproductive division of labor in termites, as it plays a key role in insect postembryonic development and reproduction. We compared the expression of JH pathway genes between workers and queens in two lower termites (Zootermopsis nevadensis and Cryptotermes secundus) and a higher termite (Macrotermes natalensis) to that of analogous nymphs and adult females of the noneusocial cockroach Blattella germanica. JH biosynthesis and metabolism genes ranged from reproductive female-biased expression in the cockroach to predominantly worker-biased expression in the lower termites. Remarkably, the expression profile of JH pathway genes sets the higher termite apart from the two lower termites, as well as the cockroach, indicating that JH signaling has undergone major changes in this eusocial termite. These changes go beyond mere shifts in gene expression between the different castes, as we find evidence for positive selection in several termite JH pathway genes. Thus, remodeling of the JH pathway may have played a major role in termite social evolution, representing a striking case of convergent molecular evolution between the termites and the distantly related social hymenoptera.

Hemimetabolous genomes reveal molecular basis of termite eusociality

Around 150 million years ago, eusocial termites evolved from within the cockroaches, 50 million years before eusocial Hymenoptera, such as bees and ants, appeared. Here, we report the 2-Gb genome of the German cockroach, Blattella germanica, and the 1.3-Gb genome of the drywood termite Cryptotermes secundus. We show evolutionary signatures of termite eusociality by comparing the genomes and transcriptomes of three termites and the cockroach against the background of 16 other eusocial and non-eusocial insects. Dramatic adaptive changes in genes underlying the production and perception of pheromones confirm the importance of chemical communication in the termites. These are accompanied by major changes in gene regulation and the molecular evolution of caste determination. Many of these results parallel molecular mechanisms of eusocial evolution in Hymenoptera. However, the specific solutions are remarkably different, thus revealing a striking case of convergence in one of the major evolutionary transitions in biological complexity.

Irreversible transfer of brood care duties and insights into the burden of caregiving in incipient subterranean termite colonies

1. In incipient termite colonies, biparental brood care rapidly shifts towards alloparental brood care. This transition was suggested to recapitulate the evolutionary trajectory from subsocial wood roach ancestors to eusociality in termites.2. Incipient colonies of the subterranean termite Coptotermes gestroi (Wasmann) were investigated to determine if this transition was irreversible and if the burden of brood care on the first alloparents could be measured. To do so, the initial size of the work force necessary for an incipient colony to survive once the brood care became alloparental was determined.3. The results of the study show that within 5 months after foundation, brood care duties were fully transferred to workers and the primary reproductives became irreversibly dependent on these workers for survival, reproduction, and colony growth.4. Once the brood care became strictly alloparental, the presence of a single worker was enough to maintain the survival of the king and queen, confirming that ‘reversed parental care’ was also achieved. However, major brood loss and suppressed egg‐laying activity from the queen was observed, suggesting that the burden of brood care was too high for a single worker to absorb. Therefore, once brood care has shifted to alloparents, a critical number of workers is necessary to prevent brood loss and initiate colony growth.5. As the initial cost of brood care is rapidly absorbed with colony growth in termites, the performance in brood care of the first few alloparents in a subsocial wood roach ancestor may have contributed to the emergence of eusociality in this clade.

Group-housed females promote production of asexual ootheca in American cockroaches

BackgroundFacultative parthenogenesis, seen in many animal phyla, is a reproductive strategy in which females are able to generate offspring when mating partners are unavailable. In some subsocial and eusocial insects, parthenogenesis is often more prevalent than sexual reproduction. However, little is known about how social cooperation is linked to the promotion of parthenogenesis. The domiciliary cockroach Periplaneta americana is well-suited to addressing this issue as this species belongs to the superfamily Blattoidea, which diverged into eusocial termites and shows facultative parthenogenesis.ResultsWe studied environmental factors that influence asexual production of ootheca using behavioral assays in P. americana. When more than three virgin females immediately after the imaginal molt were kept together in a small sealed container, they tended to produce egg cases (oothecae) via parthenogenesis earlier than did isolated females, resulting in apparent synchronization of ootheca production, even among females housed in different containers. In contrast, virgin females housed with genitalia-ablated males or group-housed females with antennae ablated did not significantly promote ootheca production compared to isolated females. Daily addition of the primary sex pheromone component to the container did not promote ootheca production in isolated females. Another line of study showed that grouped females make parthenogenesis more sustainable than previously known; a founder colony of 15 virgin females was sufficient to produce female progeny for a period of more than three years.ConclusionsGroup-housed females promote and stabilize asexual ootheca production compared to isolated females, and that this promotion is triggered by female-specific chemosensory signals (other than sex pheromone) primarily detected by antennae. Promotion of ootheca production between females is likely to be an early stage of social cooperation, reminiscent of the foundation and maintenance of a colony by female pairs in the eusocial termite Reticulitermes speratus.

Evidence for social parasitism of early insect societies by Cretaceous rove beetles.

The evolution of eusociality in ants and termites propelled both insect groups to their modern ecological dominance. Yet, eusociality also fostered the evolution of social parasitism—an adverse symbiosis, in which the superorganismal colonies formed by these insects are infiltrated by a profusion of invertebrate species that target nest resources. Predominant among these are the aleocharine rove beetles (Staphylinidae), a vast and ecologically diverse subfamily with numerous morphologically and behaviourally specialized socially parasitic lineages. Here, we report a fossil aleocharine, Mesosymbion compactus gen. et sp. nov., in Burmese amber (∼99 million years old), displaying specialized anatomy that is a hallmark of social parasites. Mesosymbion coexisted in the Burmese palaeofauna with stem-group ants and termites that provide the earliest indications of eusociality in both insect groups. We infer that the advent of eusociality led automatically and unavoidably to selection for social parasitism. The antiquity and adaptive flexibility of aleocharines made them among the first organisms to engage in this type of symbiosis.