Female developmental environment delays development of male honeybee (Apis mellifera)

Background Nutrition and cell size play an important role in the determination of caste differentiation in queen and worker of honeybees (Apis mellifera), whereas the haploid genome dominates the differentiation of drones. However, the effects of female developmental environment on the development of males remain unclear. In this study, young drone larvae were transferred into worker cells (WCs) or remained in drone cells (DCs) to rear drones. The drone larvae were also grafted into queen cells (QCs) for 48 h and then transplanted into drone cells until emerging. Morphological indexes and reproductive organs of these three types of newly emerged drones were measured. Newly emerged drones and third instar drone larvae from WCs, DCs and QCs were sequenced by RNA sequencing (RNA-Seq). Results The amount of food remaining in cells of the QC and WC groups was significantly different to that in the DC group at the early larval stage. Morphological results showed that newly emerged DC drones had bigger body sizes and more well-developed reproductive tissues than WC and QC drones, whereas the reproductive tissues of QC drones were larger than those of WC drones. Additionally, whole body gene expression results showed a clear difference among three groups. At larval stage there were 889, 1761 and 1927 significantly differentially expressed genes (DEGs) in WC/DC, QC/DC and WC/QC comparisons, respectively. The number of DEGs decreased in adult drones of these three comparisons [678 (WC/DC), 338 (QC/DC) and 518 (WC/QC)]. A high number of DEGs were involved in sex differentiation, growth, olfaction, vision, mammalian target of rapamycin (mTOR), Wnt signaling pathways, and other processes. Conclusions This study demonstrated that the developmental environment of honeybee females can delay male development, which may serve as a model for understanding the regulation of sex differentiation and male development in social insects by environmental factors.

Nutrition induced direct fitness for workers in a primitively eusocial wasp

Eusocial insects are characterised by a reproductive caste differentiation such that egg-laying is restricted to a small number of queens. The majority of the colony members function as non-reproducing workers and gain indirect fitness by rearing the offspring of the reproductives. In primitively eusocial species, some workers can also get direct fitness by sneaking in some eggs in the presence of the queens, replacing the queens and becoming new queens, or initiating new nests. Here we aimed to understand the factors that permit some workers to gain direct fitness and alter the number of workers doing so, using the primitively eusocial wasp Ropalidia marginata. We transplanted 12 naturally occurring colonies into large laboratory cages where there was adequate space for the workers to leave their natal nests and initiate new colonies. We compared six control colonies that we provided with ad libitum food placed near the nest to six test colonies in which we hand-fed the wasps in addition to the ad libitum food. Wasps in test colonies consumed more food, showed more aggression, replaced their queens, and initiated new nests significantly more often than those in control colonies. When considering all 12 colonies, the rates of queen replacement and nest foundation were significantly positively correlated with food consumption rates. The additional nutrition gained by hand-fed wasps appears to help workers in test colonies to develop their ovaries and lay eggs, implying that they are nutritionally castrated in control colonies.

Regulation of host phenotypic plasticity by gut symbiont communities in the Eastern Subterranean termite (Reticulitermes flavipes Kollar)

Termites are eusocial insects that host a range of prokaryotic and eukaryotic gut symbionts and can differentiate into a range of caste phenotypes. Soldier caste differentiation from termite workers follows two successive molts (worker-presoldier-soldier) that are driven at the endocrine level by juvenile hormone (JH). While physiological and eusocial mechanisms tied to JH signaling have been studied, the role of gut symbionts in the caste differentiation process is poorly understood. Here, we used the JH analog-methoprene in combination with the antibiotic kanamycin to manipulate caste differentiation and gut bacterial loads in Reticulitermes flavipes termites via four bioassay treatments: kanamycin, methoprene, kanamycin+methoprene, and an untreated (negative) control. Bioassay results demonstrated a significantly higher number of presoldiers in the methoprene, highest mortality in kanamycin+methoprene, and significantly reduced protist numbers in all treatments except the untreated control. Bacterial 16S rRNA gene sequencing provided alpha and beta diversity results that mirrored bioassay findings. From ANCOM analysis, we found that several bacterial genera were differentially abundant among treatments. Finally, follow-up experiments showed that if methoprene and kanamycin or untreated termites are placed together, zero or rescued presoldier initiation (respectively) occurs. These findings reveal that endogenous JH selects for symbiont compositions required to successfully complete presoldier differentiation. However, if the gut is voided before the influx of JH, it cannot select for the necessary symbionts that are crucial for molting. Based on these results we are able to provide a novel example of linkages between gut microbial communities and host phenotypic plasticity.

Extent and complexity of RNA processing in the development of honey bee queen and worker castes revealed by Nanopore direct RNA sequencing

The distinct honey bee (Apis mellifera) worker and queen castes have become a model for the study of genomic mechanisms of phenotypic plasticity. Prior studies have explored differences in gene expression and methylation during development of the two castes, but thus far no study has performed a genome-wide analysis of differences in RNA processing. To address this here we performed a Nanopore-based direct RNA sequencing with exceptionally long reads to compare the mRNA transcripts between honey bee queen and workers at three points during their larval development. We found thousands of significantly differentially expressed isoforms (DEIs) between queen and worker larvae. Most DEIs contained alternative splicing, and many of them contained at least two types of alternative splicing patterns, indicating complex RNA processing in honey bee caste differentiation. We found a negative correlation between poly(A) length and DEI expression, suggesting that poly(A) tails participate in the regulation of isoform expression. Hundreds of isoforms uniquely expressed in either queens or workers during their larval development, and isoforms were expressed at different points in queen and worker larval development demonstrating a dynamic relationship between isoform expression and developmental mechanisms. These findings show the full complexity of RNA processing and transcript expression in honey bee phenotypic plasticity.

Caste-specific phenotypic plasticity of Asian weaver ants: Revealing the allometric and non-allometric component of female caste system of Oecophylla smaragdina (Hymenoptera: Formicidae) by using geometric morphometrics

In eusocial insects, particularly in ants, caste differentiation is extremely complicated when we rely on traditional taxonomy. In most species, the worker caste does not display any distinct morphological characters neither the caste’s central division according to their morphological size variations. We used a landmark-based geometric morphometric approach to quantify the morphological characteristics of female caste systems (queen, major and minor worker ant) of Oecophylla smaragdina. Our findings suggested that each caste has its unique shape and size. Especially in the worker caste, apart from the size variations, we can use the shape as a prominent tool for distinguishing between them. The O. smaragdina exhibits a triphasic allometry pattern. Studying the allometry and non-allometry components of each caste system revealed a highly complex size and shape relationship in the female caste systems. From the allometric and non-allometric analysis, we concluded that the major worker ants showed a closer relationship with the queen than the minor worker ant. This outcome demonstrated that Asian weaver ant exhibits complex shape variations related to size and is correlated to their functional modular characters. This research sheds new light on caste systems’ taxonomic uncertainties for eusocial hymenopteran groups, especially ants.

10-hydroxy-2E-decenoic acid (10HDA) does not promote caste differentiation in Melipona scutellaris stingless bees

In bees from genus Melipona, differential feeding is not enough to fully explain female polyphenism. In these bees, there is a hypothesis that in addition to the environmental component (food), a genetic component is also involved in caste differentiation. This mechanism has not yet been fully elucidated and may involve epigenetic and metabolic regulation. Here, we verified that the genes encoding histone deacetylases HDAC1 and HDAC4 and histone acetyltransferase KAT2A were expressed at all stages of Melipona scutellaris, with fluctuations between developmental stages and castes. In larvae, the HDAC genes showed the same profile of Juvenile Hormone titers—previous reported—whereas the HAT gene exhibited the opposite profile. We also investigated the larvae and larval food metabolomes, but we did not identify the putative queen-fate inducing compounds, geraniol and 10-hydroxy-2E-decenoic acid (10HDA). Finally, we demonstrated that the histone deacetylase inhibitor 10HDA—the major lipid component of royal jelly and hence a putative regulator of honeybee caste differentiation—was unable to promote differentiation in queens in Melipona scutellaris. Our results suggest that epigenetic and hormonal regulations may act synergistically to drive caste differentiation in Melipona and that 10HDA is not a caste-differentiation factor in Melipona scutellaris.

Caste-biased Expression of fem and Amdsx Genes in Apis mellifera ligustica (Hymenoptera: Apidae)

Sex determination and caste differentiation are two crucial processes for morphology building in honeybees. It is unclear whether there is an interaction between these two processes. Here, we investigated the expression of fem and Amdsx genes between female castes of honeybees. We found that the expression of fem and Amdsx is significantly higher in queens than in workers, and this expression was positively regulated by juvenile hormone (JH). Our results suggest that sex-determining genes fem and Amdsx are also involved in honeybee caste differentiation.

Honeybee (Apis mellifera) Maternal Effect Causes Alternation of DNA Methylation Regulating Queen Development

Queen-worker caste dimorphism is a typical trait for honeybees (Apis mellifera). We previously showed a maternal effect on caste differentiation and queen development, where queens emerged from queen-cell eggs (QE) had higher quality than queens developed from worker cell eggs (WE). In this study, newly-emerged queens were reared from QE, WE, and 2-day worker larvae (2L). The thorax size and DNA methylation levels of queens were measured. We found that queens emerging from QE had significantly larger thorax length and width than WE and 2L. Epigenetic analysis showed that QE/2L comparison had the most different methylated genes (DMGs, 612) followed by WE/2L (473), and QE/WE (371). Interestingly, a great number of DMGs (42) were in genes belonging to mTOR, MAPK, Wnt, Notch, Hedgehog, FoxO, and Hippo signaling pathways that are involved in regulating caste differentiation, reproduction and longevity. This study proved that honeybee maternal effect causes epigenetic alteration regulating caste differentiation and queen development.

Among the shapeshifters: parasite-induced morphologies in ants (Hymenoptera, Formicidae) and their relevance within the EcoEvoDevo framework

As social insects, ants represent extremely interaction-rich biological systems shaped by tightly integrated social structures and constant mutual exchange with a multitude of internal and external environmental factors. Due to this high level of ecological interconnection, ant colonies can harbour a diverse array of parasites and pathogens, many of which are known to interfere with the delicate processes of ontogeny and caste differentiation and induce phenotypic changes in their hosts. Despite their often striking nature, parasite-induced changes to host development and morphology have hitherto been largely overlooked in the context of ecological evolutionary developmental biology (EcoEvoDevo). Parasitogenic morphologies in ants can, however, serve as “natural experiments” that may shed light on mechanisms and pathways relevant to host development, plasticity or robustness under environmental perturbations, colony-level effects and caste evolution. By assessing case studies of parasites causing morphological changes in their ant hosts, from the eighteenth century to current research, this review article presents a first overview of relevant host and parasite taxa. Hypotheses about the underlying developmental and evolutionary mechanisms, and open questions for further research are discussed. This will contribute towards highlighting the importance of parasites of social insects for both biological theory and empirical research and facilitate future interdisciplinary work at the interface of myrmecology, parasitology, and the EcoEvoDevo framework.