Inter-clonal competition over queen succession imposes a cost of parthenogenesis on termite colonies.

In social insect colonies, selfish behaviour due to intracolonial conflict among members can result in colony-level costs despite close relatedness. In certain termite species, queens use asexual reproduction for within-colony queen succession but rely on sexual reproduction for worker and alate production, resulting in multiple half-clones of a single primary queen competing for personal reproduction. Our study demonstrates that competition over asexual queen succession among different clone types leads to the overproduction of parthenogenetic offspring, resulting in the production of dysfunctional parthenogenetic alates. By genotyping the queens of 23 field colonies of Reticulitermes speratus, we found that clone variation in the queen population reduces as colonies develop. Field sampling of alates and primary reproductives of incipient colonies showed that overproduced parthenogenetic offspring develop into alates that have significantly smaller body sizes and much lower survivorship than sexually produced alates. Our results indicate that while the production of earlier and more parthenogenetic eggs is advantageous for winning the competition for personal reproduction, it comes at a great cost to the colony. Thus, this study highlights the evolutionary interplay between individual-level and colony-level selection on parthenogenesis by queens.

Maternal determination of soldier proportion and paternal determination of soldier sex ratio in hybrid Reticulitermes (Isoptera: Rhinotermitidae) termite colonies.

Altruistic caste, including worker and soldier (derived from worker), plays a critical role in the ecological success of social insects. The proportion of soldiers, soldier sex ratios, and the number of workers vary significantly between species, and also within species, depending on colony developmental stage and environmental factors. However, it is unknown whether there are sex-linked effects from parents on controlling the caste fate or not. Here, we compared soldier sex ratios, soldier proportions, and population size among a four mating types of Reticulitermes amamianus (Ra) and R. speratus (Rs) (male × female, mRa × fRa, mRa × fRs, mRs × fRa, mRs × fRs) and demonstrate that the soldier sex ratio and worker population size of hybrid colonies skew to colonies of king's species, while the soldier proportion skew to queen's species. The survival rate of offspring resulting from interspecies hybridization was significantly higher for mRa × fRs than for mRs × fRa. The results of this study demonstrate the asymmetric influence of kings and queens on caste determination and colony growth, which can contribute to our better understanding of parental influence on the colony dynamics of social insects.

Cell type specific polyploidization in the royal fat body of termite queens.

Tissue-specific endopolyploidy is widespread among plants and animals and its role in organ development and function has long been investigated. In insects, the fat body cells of sexually mature females produce substantial amounts of egg yolk precursor proteins (vitellogenins) and exhibit high polyploid levels, which is considered crucial for boosting egg production. Termites are social insects with a reproductive division of labor, and the fat bodies of mature termite queens exhibit higher ploidy levels than those of other females. The fat bodies of mature termite queens are known to be histologically and cytologically specialized in protein synthesis. However, the relationship between such modifications and polyploidization remains unknown. In this study, we investigated the relationship among cell type, queen maturation, and ploidy levels in the fat body of the termite Reticulitermes speratus. We first confirmed that the termite fat body consists of two types of cells, that is, adipocytes, metabolically active cells, and urocytes, urate-storing cells. Our ploidy analysis using flow cytometry has shown that the fat bodies of actively reproducing queens had more polyploid cells than those of newly emerged and pre-reproductive queens, regardless of the queen phenotype (adult or neotenic type). Using image-based analysis, we found that not urocytes, but adipocytes became polyploid during queen differentiation and subsequent sexual maturation. These results suggest that polyploidization in the termite queen fat body is associated with sexual maturation and is regulated in a cell type-specific manner. Our study findings have provided novel insights into the development of insect fat bodies and provide a basis for future studies to understand the functional importance of polyploidy in the fat bodies of termite queens.

The royal food of termites shows king and queen specificity.

Society in eusocial insects is based on the reproductive division of labor, with a small number of reproductive individuals supported by a large number of nonreproductive individuals. Because inclusive fitness of all colony members depends on the survival and fertility of reproductive members, sterile members provide royals with special treatment. Here, we show that termite kings and queens each receive special food of a different composition from workers. Sequential analysis of feeding processes demonstrated that workers exhibit discriminative trophallaxis, indicating their decision-making capacity in allocating food to the kings and queens. Liquid chromatography tandem-mass spectrometry analyses of the stomodeal food and midgut contents revealed king- and queen-specific compounds, including diacylglycerols and short-chain peptides. Desorption electrospray ionization mass spectrometry imaging analyses of 13C-labeled termites identified phosphatidylinositol and acetyl-l-carnitine in the royal food. Comparison of the digestive tract structure showed remarkable differences in the volume ratio of the midgut-to-hindgut among castes, indicating that digestive division of labor underlies reproductive division of labor. Our demonstration of king- and queen-specific foods in termites provides insight into the nutritional system that underpins the extraordinary reproduction and longevity of royals in eusocial insects.

A method for estimating colony size using queen fecundity in termites under field conditions.

Colony size in social insects is one of the most important factors in shaping their self-organized system. It affects a wide variety of traits such as foraging and defense strategies, social immune responses, the degree of polymorphism, and reproductive output. However, colony size estimation of subterranean termites in the field has been challenging, due to their extremely cryptic biology and multiple site-nesting behavior. Since natural selection favors workers that maximize the number of their siblings, the amount of egg production may reflect the number of workers in the colony. Here, we report a method for inferring colony size in the field using total egg production in each colony from a subterranean termite, Reticulitermes speratus. Our investigation of field colonies revealed that the body weight of queens reaches a peak and had the largest variance in June and July and accurately predicts the number of eggs laid by the queen per 24 h. Using laboratory-reared colonies, we found that the total egg production in each colony is proportional to the number of workers. We also estimated the colony size of 198 field colonies and found that the median and maximum colony size was 24,500 and 451,800 workers per colony. The method for inferring colony size presented here may also be applicable to termite species with a clear seasonality in egg production. The colony size estimate will contribute to understanding the life history strategies and social systems of termites.

Plastic brain structure changes associated with the division of labor and aging in termites.

Division of labor is a prominent feature of social insect societies, where different castes engage in different specialized tasks. As brain differences are associated with behavioral differences, brain anatomy may be linked to caste polymorphism. Here, we show that termite brain morphology changes markedly with caste differentiation and age in the termite, Reticulitermes speratus. Brain morphology was shown to be associated with reproductive division of labor, with reproductive individuals (alates and neotenic reproductives) having larger brains than nonreproductives (workers and soldiers). Micro-computed tomography (CT) imaging and dissection observations showed that the king's brain morphology changed markedly with shrinkage of the optic lobes during their long life in the dark. Behavioral experiments showed that mature primary kings lose visual function as a result of optic lobe shrinkage. These results suggested that termites restructure their nervous systems to perform necessary tasks as they undergo caste differentiation, and that they also show flexible changes in brain morphology even after the final molt. This study showed that brain morphology in social insects is linked to caste and aging, and that the evolution of the division of labor is underpinned by the development of diverse neural systems for specialized tasks.

Discovery of an underground chamber to protect kings and queens during winter in temperate termites.

Overwintering is a critical part of the annual cycle for species that live in temperate, polar, and alpine regions. Consequently, low-temperature biology is a key determinant of temperate species distribution. Termites are distributed predominantly in tropical regions, and a limited number of species are found in the temperate zone. Here, in the termite Reticulitermes speratus, we report the discovery of an underground chamber that protects kings and queens to survive the winter, which is separate from the one they used during the warmer breeding season. In the spring, the royals inhabited decayed logs on the ground, then moved to their underground chamber located in the roots of stumps in the fall. The winter minimum temperature measured in the royal chamber was higher than that in the logs on the ground. In overwintering termites, the kings and queens had higher cold tolerance than workers and soldiers. Air temperatures dropped below the critical temperature multiple times, as evidenced from the past 140 years of weather records in Kyoto. These results demonstrated the survival strategies of reproductives to overcome the environment at the latitudinal limits. This study helps further the understanding of the termite's seasonal phenology, long-term survivorship, and life cycle.

Heritable effects on caste determination and colony-level sex allocation in termites under field conditions.

The ecological success of social insects is attributed to the division of labor, where newly hatched offspring differentiate into either fertile progeny or functionally sterile worker castes. There is growing evidence for the heritable (genetic or epigenetic) effects on caste determination based on laboratory experiments. Here, we indirectly demonstrate that heritable factors have the principal role in caste determination and strongly affect colony-level production of both sexes of fertile dispersers (i.e., alates) in field colonies of the termite Reticulitermes speratus. An egg-fostering experiment suggests that the colony-dependent sex-specific caste fates were almost entirely determined before oviposition. Our investigation of field colonies revealed that such colony-dependent sex-specific caste fates result in the intercolonial variation in the numerical sex ratio of differentiated fertile offspring and, eventually, that of alates. This study contributes to better understanding the mechanisms underlying the division of labor and life-history traits in social insects.

King- and queen-specific degradation of uric acid contributes to reproduction in termites.

Caste-based reproductive division of labour in social insects is built on asymmetries in resource allocation within colonies. Kings and queens dominantly consume limited resources for reproduction, while non-reproductive castes such as workers and soldiers help reproductive castes. Studying the regulation of such asymmetries in resource allocation is crucial for understanding the maintenance of sociality in insects, although the molecular background is poorly understood. We focused on uric acid, which is reserved and used as a valuable nitrogen source in wood-eating termites. We found that king- and queen-specific degradation of uric acid contributes to reproduction in the subterranean termite Reticulitermes speratus. The urate oxidase gene (RsUAOX), which catalyses the first step of nitrogen recycling from stored uric acid, was highly expressed in mature kings and queens, and upregulated with differentiation into neotenic kings/queens. Suppression of uric acid degradation decreased the number of eggs laid per queen. Uric acid was shown to be provided by workers to reproductive castes. Our results suggest that the capacity to use nitrogen, which is essential for the protein synthesis required for reproduction, maintains colony cohesion expressed as the reproductive monopoly held by kings and queens.

Proteomic analysis of the heart in normal aging mice.

Aging induces pathological cardiovascular changes such as cardiac dysfunction and arteriosclerosis. With aging, heart cells, especially, become more susceptible to lethal damage. In this report, we tried to understand the precise mechanism of myocardial change resulting from aging by examining the heart proteome in aging mice using two-dimensional gel electrophoresis (2DE). The proteins were stained with fluorescence dyes (SYPRO Ruby and Pro-Q Diamond) and identified by subsequent MALDI-TOF-MS / MS. As a result, markedly altered levels of 14 proteins and 7 phosphoproteins were detected in the hearts of 3-, 7-, 11-, and 20-month-old mice. The functions of these identified proteins and phosphoproteins were energy metabolism, muscle contraction, glycolysis, and cytoskeletal support. Additionally, the results of Western blotting confirmed changes in the expression of FTH, CPNE5, and SUCLA2. These findings showed that aging modified the expression of proteins and phosphoproteins in the heart. We suggest that changes in the expression of these proteins are critical to the development of cardiac dysfunction resulting from aging. J. Med. Invest. 69 : 217-223, August, 2022.

First Polycipivirus and Unmapped RNA Virus Diversity in the Yellow Crazy Ant, Anoplolepis gracilipes.

The yellow crazy ant, Anoplolepis gracilipes is a widespread invasive ant that poses significant threats to local biodiversity. Yet, compared to other global invasive ant species such as the red imported fire ant (Solenopsis invicta) or the Argentine ant (Linepithema humile), little is known about the diversity of RNA viruses in the yellow crazy ant. In the current study, we generated a transcriptomic database for A. gracilipes using a high throughput sequencing approach to identify new RNA viruses and characterize their genomes. Four virus species assigned to Dicistroviridae, two to Iflaviridae, one to Polycipiviridae, and two unclassified Riboviria viruses were identified. Detailed genomic characterization was carried out on the polycipivirus and revealed that this virus comprises 11,644 nucleotides with six open reading frames. Phylogenetic analysis and pairwise amino acid identity comparison classified this virus into the genus Sopolycivirus under Polycipiviridae, which is tentatively named "Anoplolepis gracilipes virus 3 (AgrV-3)". Evolutionary analysis showed that AgrV-3 possesses a high level of genetic diversity and elevated mutation rate, combined with the common presence of multiple viral strains within single worker individuals, suggesting AgrV-3 likely evolves following the quasispecies model. A subsequent field survey placed the viral pathogen "hotspot" of A. gracilipes in the Southeast Asian region, a pattern consistent with the region being recognized as part of the ant's native range. Lastly, infection of multiple virus species seems prevalent across field colonies and may have been linked to the ant's social organization.

Enhanced heterozygosity from male meiotic chromosome chains is superseded by hybrid female asexuality in termites.

Although males are a ubiquitous feature of animals, they have been lost repeatedly in diverse lineages. The tendency for obligate asexuality to evolve is thought to be reduced in animals whose males play a critical role beyond the contribution of gametes, for example, via care of offspring or provision of nuptial gifts. To our knowledge, the evolution of obligate asexuality in such species is unknown. In some species that undergo frequent inbreeding, males are hypothesized to play a key role in maintaining genetic heterozygosity through the possession of neo-sex chromosomes, although empirical evidence for this is lacking. Because inbreeding is a key feature of the life cycle of termites, we investigated the potential role of males in promoting heterozygosity within populations through karyotyping and genome-wide single-nucleotide polymorphism analyses of the drywood termite Glyptotermes nakajimai We showed that males possess up to 15 out of 17 of their chromosomes as sex-linked (sex and neo-sex) chromosomes and that they maintain significantly higher levels of heterozygosity than do females. Furthermore, we showed that two obligately asexual lineages of this species-representing the only known all-female termite populations-arose independently via intraspecific hybridization between sexual lineages with differing diploid chromosome numbers. Importantly, these asexual females have markedly higher heterozygosity than their conspecific males and appear to have replaced the sexual lineages in some populations. Our results indicate that asexuality has enabled females to supplant a key role of males.

The lose-to-win strategy of the weak: intraspecific parasitism via egg abduction in a termite.

Predation by larger conspecifics poses a major threat to small juveniles in many animal species. However, in social insects, raids perpetrated by large colonies may provide smaller colonies with opportunities for parasitization. Herein, in the termite Reticulitermes speratus, we demonstrate that small incipient colonies parasitize large mature colonies through egg abduction when attacked by raiding conspecifics. We observed that the eggs of incipient colonies were brought into raiding colonies while their parents were killed during the attack. In this species, unmated females found new colonies with female-female (FF) cooperation, in addition to the typical monogamous colony foundation. Interestingly, the abducted eggs of FF pairs developed into nymphs (reproductive caste) in the raiding colonies, whereas the eggs of male-female (MF) pairs developed into workers (non-reproductive caste). Parthenogenetic eggs are known to be developmentally predisposed to becoming female reproductives owing to genomic imprinting in termites. This study demonstrates that the plundering of small colonies by larger conspecific colonies not only results in the extinction of the weaker colonies, but also serves as a strategy that incipient colonies use to obtain the reproductive position in large colonies by stealth. The results elucidate the diversity and complexity of inter-colonial interactions in social insects.

Improving invasive species management using predictive phenology models: an example from brown marmorated stink bug (Halyomorpha halys) in Japan.

BACKGROUND: In order to better understand the population dynamics of invasive species in their native range, we developed two predictive phenological models using the ubiquitous invasive insect pest, Halyomorpha halys (Stål) (Hemiptera: Pentatomidae), as the model organism. Our work establishes a zero-inflated negative binomial regression (ZINB) model, and a general additive mixed model (GAMM) based on 11 years of black light trap monitoring of H. halys at three locations in Japan. RESULTS: The ZINB model indicated that degree days (DD) have a significant effect on the trap catch of adult H. halys, and that precipitation has no effect. A dataset generated by 1000 simulations from the ZINB suggested that higher predicted trap catches equated to a lower probability of encountering a zero-count. The GAMM produced a cubic regression smooth curve which forecasts the seasonal phenology of H. halys as following a bell-shaped trend in Japan. Critical DD points during the field season in Japan included 261 DD for first H. halys adult detection and 1091 DD for peak activity. CONCLUSIONS: This study establishes the first models capable of forecasting native H. halys population dynamics based on DD. These robust models practically improve population forecasting of H. halys in the future and help fill gaps in knowledge pertaining to its native phenology, thus ultimately contributing to the progression of efficient management of this globally invasive species. © 2021 Society of Chemical Industry.

Oocyte resorption in termite queens: Seasonal dynamics and controlling factors.

Female insects can resorb their oocytes that could not be oviposited. Oocyte resorption is proposed to be an adaptive mechanism to optimize fitness in hostile environments, recouping resources that might otherwise be lost. Social insects have developed reproductive division of labor, wherein a small number of queens are devoted to egg production. Matured queens are highly specialized in reproduction and are largely dependent on nestmate workers for their nourishment. Therefore, oocyte resorption in the queens should be influenced by social factors such as the amount of available workforce, as well as external and abiotic factors. In this study, we investigated the seasonal dynamics and regulation factors of oocyte resorption in actively reproducing termite queens. We continuously collected the field-nests of the subterranean termite Reticulitermes speratus and demonstrated that queens frequently resorbed their oocytes in late summer, even though it is one of the most productive seasons in this species. On the other hand, our laboratory experiment showed that oocyte resorption itself was strongly induced regardless of the season. We also found that the rate of oocyte resorption was influenced by colony size (the number of attending workers). These results suggest that termite queens seasonally resorb their oocytes, yet oocyte resorption itself is regulated by social factors rather than by seasonal factors. Our study provides a unique insight into the regulation of reproduction in social insects.

Why and how do termite kings and queens live so long?

Lifespan varies greatly across the tree of life. Of the various explanations for this phenomenon, those that involve trade-offs between reproduction and longevity have gained considerable support. There is an important exception: social insect reproductives (queens and in termites, also kings) exhibit both high reproductive outputs and extraordinarily long lives. As both the ultimate and proximate mechanisms underlying the absence of the fecundity/longevity trade-off could shed light on the unexpected dynamics and molecular mechanisms of extended longevity, reproductives of social insects have attracted much attention in the field of ageing research. Here, we highlight current ecological and physiological studies on ageing and discuss the various possible evolutionary and molecular explanations of the extended lifespans of termite reproductives. We integrate these findings into a coherent framework revealing the evolution of longevity in these reproductives. Studies on termites may explain why and how ageing is shaped by natural selection. This article is part of the theme issue 'Ageing and sociality: why, when and how does sociality change ageing patterns?'

Proteomic analysis involved with synaptic plasticity improvement by GABAA receptor blockade in hippocampus of a mouse model of Alzheimer's disease.

GABAergic system plays a part in synaptic plasticity in the hippocampus. We had reported a long-term potentiation (LTP)-like facilitation in vivo, known as synaptic plasticity, through GABAA receptor blockade by bicuculline and the expression of proteins involved with this synaptic plasticity in mouse hippocampus. In the present study, we aimed to show improvement of impaired synaptic plasticity through GABAA receptor blockade and to clarify the molecular mechanisms involved with this improvement in the hippocampus of mice overexpressing human amyloid precursor protein with the E693Δ mutation (APPOSK-Tg) as an Alzheimer's disease model showing impaired synaptic plasticity. Electrophysiological study showed that the LTP-like facilitation expressed with application of bicuculline in vivo was significantly greater than impaired tetanic LTP in APPOSK-Tg mice, which was improved by bicuculline. Proteomic analysis showed that the expression of 11 proteins in the hippocampus was significantly changed 8 h after bicuculline application to APPOSK-Tg mice. The identified proteins could be functionally classified as chaperone, cytoskeletal protein, energy metabolism, metabolism, neuronal development, and synaptic component. Additionally, western blotting validated the changes in four proteins. We therefore propose that the improvement of impaired synaptic plasticity through GABAA receptor blockade could be mediated by the changed expression of these proteins.

Ongoing Coevolution of Wolbachia and a Widespread Invasive Ant, Anoplolepis gracilipes.

While Wolbachia are commonly found among arthropods, intraspecific infection rates can vary substantially across the geographic populations. Here we report nearly 100% prevalence of Wolbachia in the global populations of the yellow crazy ant, Anoplolepis gracilipes. To understand coevolutionary history between Wolbachia and A. gracilipes, we identified single nucleotide polymorphisms (SNPs) in Wolbachia from the ant across 12 geographical regions and compared the phylogeny of SNP-based Wolbachia to patterns of the ant's mitochondrial DNA (mtDNA) variation. Our results revealed a strong concordance between phylogenies of Wolbachia and host mtDNA, providing immediate evidence of co-divergence. Among eight identified SNP loci separating the genetic clusters of Wolbachia, seven loci are located in potential protein-coding genes, three of which being non-synonymous SNPs that may influence gene functions. We found a Wolbachia hypothetical protein gene with signature of positive selection. These findings jointly allow us to characterize Wolbachia-ant coevolution and also raise a question about mechanism(s) underlying maintenance of high prevalence of Wolbachia during the colonization of this invasive ant.

Age-Dependent Increase in Soldier Pheromone of the Termite Reticulitermes speratus.

Pheromone communication helps maintaining the sophisticated colony organization in social insects. In the termite Reticulitermes speratus, there are two functionally distinct soldier groups: royal guards and entrance guards. Royal guards protect kings and queens in the innermost part of the nest, whereas entrance guards prevent predators from intruding into the nest at the periphery. A recent study revealed that younger and older soldiers work as royal and entrance guards, respectively. This age-dependent distribution is thought to help workers to recognize where in the nest they are located. However, it is not known whether workers can discriminate the age of soldiers. Here, we show that the abundance of soldier pheromone changes with age and that workers discriminate a soldier's age by recognizing the pheromone abundance. Gas chromatography-mass spectrometry analysis revealed that the chemical profiles of extracts in three soldier groups of different ages (newly differentiated, royal guard, and entrance guard soldiers) are markedly different. Entrance guard soldiers have the most soldier pheromone among the three age classes. Furthermore, our bioassays suggested that the worker's movement from chamber to chamber is inhibited only when a soldier with less soldier pheromone is located at the chamber entrance. These results suggest that the soldier pheromone functions as a soldier age indicator and that workers change their behavior depending on the age of the soldier defending the chamber entrance. This study contributes to our understanding of the relationship between aging and pheromone communication in social insects.

Chemical identification of an aggregation pheromone in the termite Reticulitermes speratus.

Social behaviours in termites are regulated by sophisticated chemical communication systems. The majority of subterranean termites continuously forage for new wood resources to expand their nesting areas; an aggregation pheromone is presumed to regulate this process. However, the chemical components of this pheromone have never been determined. We identified the chemical properties of the aggregation pheromone that signals nestmate presence and induces arrest in the termite Reticulitermes speratus. The results of gas chromatography-mass spectrometry analyses and bioassays indicated that R. speratus worker release the pheromone to their nesting site. The pheromone consists of an aromatic compound (2-phenylundecane), cuticular hydrocarbons (pentacosane and heptacosane), fatty acids (palmitic acid and trans-vaccenic acid), and cholesterol; the pheromone induces long-term aggregation at new nesting and feeding sites. Although 2-phenylundecane alone attracted workers, the combination of all six compounds showed greater arrestant activity than 2-phenylundecane alone. This suggests that 2-phenylundecane functions as an attractant, whereas the remaining five components function as arrestants. Our results indicate that foraging worker termites produce a multi-component aggregation pheromone by combining a volatile hydrocarbon and non-volatile lipids with cuticular hydrocarbons. This pheromone enables rapid, long-lasting aggregation of termite workers, which contributes to efficient feeding and colonisation of new wood. Our work furthers the understanding of chemical communication systems underlying social assembly in social insects.