The ontogeny of soldiers in Prorhinotermes simplex (Isoptera, Rhinotermitidae)

The ontogeny of the soldier caste in Prorhinotermes simplex is studied with emphasis on the instar origin of soldiers, the duration of the presoldier stage, and changes in external anatomy during the soldier morphogenesis. The soldiers of different instar origin are compared morphometrically and by analysis of the soldier defensive secretion. The soldiers were found to develop from larvae of the second up to the eighth instar, via a short (13–17 days) presoldier stage. The early soldier instars were found exclusively in incipient colonies while the mature colony contains late instar soldiers only. The abrupt change in the external anatomy occurs in two steps. During the larva-presoldier moult, the head increases in length but only slightly in width. During the presoldier-soldier moult, both the length and width of the head increase markedly and the typical cordate shape is attained; the relative size of the pronotum increases considerably. The long falcate mandibles develop mainly during the larva-presoldier moult. One antennal segment is added during these two moults. With increasing instar age of soldiers a relative increase of the pronotum and the head size in its posterior region was observed. No functional differences in external anatomy were found among the six soldier instars. The composition of the frontal gland secretion is similar in the six soldier instars, an extraordinarily high amount of a defensive substance, (E)-1-nitropentadecene, was detected together with (Z,E)-α-farnesene, presumably an alarm substance. The strategy of soldier production is discussed. We conclude that the observed patterns of soldier production in P. simplex correspond well to its ecological strategy of an advanced single-site nester able to migrate and form foraging groups. Received 16 August 2005; revised 21 November and 6 February 2006; accepted 16 February 2006.     

Effects of heat shock and ambient temperature on female soldier production in a polyembryonic parasitic wasp

Larval castes are well known in both sexes of polyembryonic parasitic wasps, in which sterile soldier larvae are clonally produced from a single egg and thousands of reproductive larvae develop into adults. The proportion of soldier larvae in Copidosoma floridanum is first determined genetically in both sexes but subsequently increases in females as a result of environmental disturbances, such as the presence of competitive parasitoids. Because temperature is the most influential environmental factor affecting ectotherms, we hypothesize that an increase in ambient temperature also affects the production of female soldier larvae. Conversely, we hypothesize that there would be no effect of temperature on the production of male soldier larvae. In the present study, we show that heat stress experienced by C. floridanum at an early stage in host development increases the number of female soldier larvae produced during the last instar stage of the host. We also show that rearing at extreme (high and low) temperatures induces an increase in the number of female soldiers produced. The effect of heat stress and ambient temperature on soldier production is found only in females, as is the response to heterospecific competitors. By contrast, no responses are detected in males for either heat stress or ambient temperature. The adaptive effect of these phenomena is not known, although the results of the present study imply the existence of a common physiological cascade that causes an increase in the number of soldier larvae in females.     

Colony demographics of rare soldier-polymorphic worker caste systems in <Emphasis Type="Italic">Pheidole</Emphasis> ants (Hymenoptera,Formicidae)

Nearly all species in the ant genus Pheidole have dimorphic workers, with distinct small minors and larger soldiers. The size range of both castes is typically narrow. Just seven described species are soldier-polymorphic, with a broad soldier size range. Here, we characterize worker caste allocation and demography in the soldier-polymorphic P. obtusospinosa, P. rhea, and P. tepicana, and the dimorphic P. spadonia for comparison. The head allometry of soldiers in soldier-polymorphic species is strongly positive and that of dimorphic species is negative. Among soldier-polymorphic species, the soldier castes differ from each other in the degree of positive allometry. In addition, they differ in the number of size modes: P. obtusospinosa and P. rhea have two and P. tepicana has one. During colony ontogeny, P. obtusospinosa first has one mode and develops the second mode much later, while P. rhea produces multiple modes throughout. We also characterize worker caste systems based on the biomass allocation. For all three soldier-polymorphic species, the majority of soldiers are small soldiers. Pheidole obtusospinosa and P. rhea allocate roughly equal biomass to the two soldier classes, while P. tepicana allocates little to supersoldiers based on both biomass and caste ratio. These findings illustrate the interplay among caste ratios, biomass allocation, size frequency distributions, and allometry in the evolution of different worker caste systems. We conclude that soldier-polymorphic species may have evolved convergently in response to broad-scale factors, but differences among them suggest selection pressures in small-scale environments have been different.     

Preliminary studies on the control of caste formation in the harvester termiteHodotermes mossambicus (Hagen)

The control of caste formation in the harvester termiteHodotermes mossambicus (Hagen) was investigated.Normal laboratory colonies contained a primary reproductive pair, between 6.59 and 24.98% workers, 0.39 and 3.45% soldiers and 74.64 and 92.06% larvae. Supplementary reproductives did not develop in colonies deprived of their king and queen. Continuous removal of soldiers stimulated soldier production. We have suggested that a pheromone produced by the soldiers plays a major role in controlling the soldier population of a colony. A similar phermone system appreared to control the production of workers.Evidence was also obtained that the primary reproductive pair influenced the production of workers and soldiers.     

Lack of kin discrimination in the eusocial aphid pseudoregmabambucicola (Homoptera: Aphididae)

The significance of kin discrimination for the evolution of sociality is theoretically important. The behavioural response ofPseudoregma bambucicola soldiers to aphids from other colonies was studied to examine the possibility of context-specific kin discrimination. When non-kin aphids were artificially introduced into experimental colonies, they were never killed or removed by soldiers. Field experiments revealed that the soldiers frequently attacked distantly related species such asAstegopteryx bambucifoliae andCeratoglyphina styracicola, but rarely conspecific aphids or congeneric species such asP. koshunensis andP. alexanderi. Another experiment showed that the soldiers showed altruistic defence even when coexisting with non-kin reproductives. These results agree with and extend the results from previous studies of kin discrimination in social aphids. The absence of kin discrimination suggests that some other factors besides higher genetic relatedness may play a prominent role in the evolutionary maintenance of soldier behaviour in aphids.     

Soldier determination in Pheidole bicarinata: Inhibition by adult soldiers

In the ant Pheidole bicarinata, adult soldiers suppress the induction of soldier developments in larvae that have been treated with methoprene, as well as in acetone-treated controls. Induction of soldier development is inhibited in methoprene-treated larvae when the adult soldiers are present as nurses, when they are present in the colony but minor workers serve as nurses, and when they are held in a minor worker-permeable cage to prevent them from contacting larvae. Suppression of soldier determination can be overridden by high doses of methoprene applied during the juvenile hormone-sensitive period for soldier determination. The degree of suppression increases with increasing contact between larvae and adult soldiers. The most likely mechanism of inhibition is a contact soldier pheromone. On the basis of our results, we expand our model of soldier determination to accommodate the fact that the threshold titre of juvenile hormone necessary to induce soldier development changes in the presence of soldiers. A threshold that is sensitive to the presence of adult soldiers provides a mechanism whereby the worker caste ratio in colonies can be regulated.     

Buying time for colony mates: the anti-predatory function of soldiers in the eusocial aphid Ceratovacuna japonica (Homoptera, Hormaphidinae)

Eusocial aphids produce sterile individuals (soldiers) that specialize in colony protection. Killing predators is often considered the main function of soldiers. In this study, we tested the effect of harassment by soldiers of a eusocial aphid, Ceratovacuna japonica (Homoptera, Hormaphidinae), on predation by this species’ natural enemy, the larvae of Atkinsonia ignipicta (Lepidoptera, Stathmopodidae). We experimentally introduced some aphids and a predator to petri dishes and compared the survivorship of first-instar reproductives in the presence and absence of soldiers. We showed that soldiers can reduce the rate of predation on their colony mates without killing the predators. When predators encountered soldiers, they did not attempt to prey on them. Instead, they evaded them and often started to make a nest by spinning silken threads. The soldiers, in contrast, waved their forelegs and attacked the predator, and they sometimes succeeded in grasping the predator’s body. Because the predator used its mandibles to remove any soldier that succeeded in grasping its body, the soldier did not kill the predator. The reduction of predation was apparently caused by the delay of predation resulting from the harassment behavior of the soldiers. In eusocial aphids, a defensive strategy that delays predation may buy the soldiers’ colony mates time to reproduce or to escape from the predator.     

Social structure and the defensive role of soldiers in a eusocial bamboo aphid,Pseudoregma bambucicola (Homoptera: Aphididae): A test of the defence-optimization hypothesis

A complete understanding of the evolution of sociality in aphids requires a detailed knowledge of the patterns of soldier investment in their ecology. The eusocial bamboo aphidPseudoregma bambucicola has a morphologically specialized first-instar soldier caste. The proportion of soldiers was positively correlated with colony size. Within a colony, soldiers were evenly distributed among subcolonies; within each subcolony, however, their distribution was biased toward peripheries which were exposed to many predators. Field experiments introducing natural enemies such asEupeodes confrater (Diptera: Syrphidae) andSynonycha grandis (Coleoptera: Coccinellidae) revealed that the survival rate of these predators was negatively correlated with the density of soldiers, suggesting that soldiers can more or less defend their colonies by killing or removing a range of natural enemies. Observations suggest that large mature colonies attract more predators than newly established small colonies and that, within a colony, the predators attack each subcolony regardless of its position on bamboo shoots. This implies the presence of a positive correlation between colony size and predation risk. Thus, the investment in soldiers seems to reflect the attacking pattern of predators within a colony. These results agree with the defence-optimization hypothesis in soldier investment ofP. bambucicola colonies.     

Physical contact between soldier and worker is essential in soldier self-regulation of Coptotermes formosanus (Isoptera, Rhinotermitidae)

In a termite society, the soldier proportion is regulated at a species-specific level with a seasonal fluctuation. A feedback mechanism in soldier proportion regulation is well recognized, while the proximate means by which the presence of soldiers inhibits the new soldier formation from workers remains mostly elusive. In the present study, some possible means for this inhibition were tested with the Formosan subterranean termite, Coptotermes formosanus Shiraki. The results showed that neither soldier-originated volatiles nor non-volatile chemicals without physical contact were responsible for the inhibition of new soldier formation. The essentiality of physical contact between workers and soldiers in soldier regulation was established. However, by physical contacts, as the soldier-replacement experiment showed, workers could not individually recognize or “count” soldiers to maintain a set soldier proportion. The possible feedback mechanisms for soldier regulation are discussed.     

Seasonal extension of the soldier instar as a route to increased defence investment in the social aphid Pemphigus spyrothecae

Abstract.  1. The possibility that the duration of the soldier instar could be extended over the course of the galling season was examined in Pemphigus spyrothecae Passerini (Hemiptera: Aphididae). In this species, the individuals of one morph of the dimorphic first instar are non-sterile specialised defenders known as soldiers.
2. In the face of a declining birth rate, the proportion of first-instar soldiers to first-instar non-soldiers was found to increase. This provides strong evidence of extension of the soldier instar over time.
3. Instar extension provides a mechanism for post-natal investment in soldiers and allows individuals with insufficient time to mature to remain in the defensive stage so that they may continue to make an indirect contribution to their clone's reproductive success.     

Soldier differentiation during embryogenesis of a social aphid, Pseudoregma bambucicola

To understand the developmental process of aphid soldier differentiation, we investigated the morphological characters of normal nymphs, soldier nymphs and developing embryos of Pseudoregma bambucicola. Results of morphometric analyses showed that normal and soldier nymphs formed discrete clusters on the basis of several morphological characters, although a small number of intermediate individuals, termed ‘intercaste nymphs’, were present. In late embryonic stages, normal and soldier embryos were morphologically distinguishable. The earlier the embryonic stage, the smaller the morphological differences between them. In early embryos less than 1000 µm in length, normal and soldier embryos were not morphologically distinguishable, suggesting that the onset of soldier differentiation occurs at an early embryonic stage. Throughout embryonic development, morphological differentiation of the soldier caste proceeded gradually. Notably, several morphological characters of soldiers grew remarkably upon larviposition. Observation of embryonic leg cuticle revealed a characteristic folding structure, indicating that some morphological traits of the soldier are exaggerated upon larviposition through expansion of the folded cuticle. We suggest that morphological differentiation of the soldier caste in P. bambucicola comprises two phases: gradual growth during embryogenesis and rapid growth upon larviposition.     

Morphometric changes during soldier differentiation of the damp-wood termite Hodotermopsis japonica (Isoptera, Termopsidae)

Summary. To clarify the allometric development of body parts accompanying soldier differentiation in termites, we measured 16 body parts of soldiers, presoldiers, pseudergates (workers), nymphs and larvae of the damp-wood termite Hodotermopsis japonica. Principal component analysis (PCA) was performed using these parameters, which revealed that differentiation into soldiers differed distinctly from development into adult (reproductive) individuals. In particular, the anterior body parts enlarged during development of soldiers. Similarly, elongation of the apical portion of both mandibles was noted during soldier differentiation. X-ray analysis of mandibles revealed sclerotization of the soldier mandibles after differentiation into terminal soldiers. These morphological changes during soldier differentiation are associated with changes in their roles within the colony. Through soldier differentiation, the morphology of this caste of termite becomes functionally suited for attacking predators, and unsuitable for feeding on wood using their mandibles. Based on these data, we suggest that there must be some morphogenetic factors leading caste specific morphology such as soldier mandibles.     

Soldier presence suppresses presoldier differentiation through a rapid decrease of JH in the termite Reticulitermes speratus

The regulation of caste differentiation is essential to insect eusociality. Termite soldiers are sterile and cannot eat by themselves because they have specialized mouth morphology. Almost all termite species have a soldier caste, and the soldier ratio per colony is maintained at a low level, probably by elaborate regulatory mechanisms. Although the soldier presence is considered to negatively affect soldier differentiation in all examined species, the detailed mechanism remains unclear. Presoldier differentiation can be induced artificially by juvenile hormone (JH) application to workers, showing that JH is a key factor underlying the regulation of soldier differentiation. In this study, to elucidate physiological changes in workers because of the soldier presence during the molt into presoldiers, JH III applications and JH titer quantifications were carried out in the rhinotermitid termite Reticulitermes speratus. Firstly, the effects of soldier presence before the molt into presoldiers induced by JH III application to workers were investigated. The rates of presoldier molt induced by the treatments with soldiers were significantly lower than those without soldiers. Secondly, worker JH titers in the presence or absence of soldiers were quantified by LC-MS on day 0, 5, 10, and 15 after JH application. Results indicated that the worker JH titers (endogenous + applied JH III) in the presence of soldiers were significantly lower than those without soldiers on day 5 after the JH treatment. On days 10 and 15, such soldier effects were not observed. Finally, the effective duration of soldier presence after the JH application was elucidated. A 4 day period of co-existence with soldiers suppressed presoldier differentiation, suggesting that the soldier presence rapidly decreased the JH titer in other colony members (i.e., workers), resulting in the inhibition of presoldier production.     

Soldier‐biased gene expression in a subterranean termite implies functional specialization of the defensive caste

In a termite colony, reproduction is typically monopolized by a small number of sexuals that are supported by reproductively altruistic soldiers and workers. We expect caste differentiation to be associated with clear‐cut differences in gene expression, and for these differences to reflect caste function and development. Here, we use RNA‐Sequencing to compare the gene expression profiles of sexual nymphs and two non‐reproductive helper castes (i.e., workers and soldiers) of the Eastern subterranean termite Reticulitermes flavipes. We found that of n = 93 genes that are strictly expressed as a function of caste, a majority (78%) show a soldier‐specific pattern. This conspicuous soldier‐bias in genome‐wide expression suggests that this defensively specialized caste is functionally well‐differentiated from both the reproductive and the other non‐reproductive caste of this species, despite a shared developmental program with workers. Gene ontology analysis supports the notion of functional specialization by soldiers, as soldier‐biased gene sets are enriched for novel biological processes. Whether this pattern reflects ancient or more recent bouts of selection for caste novelty at the gene‐regulatory level is not known, but because soldiers are sterile and thus have no direct fitness, any selection for novelty must have been mediated indirectly, through reproducing relatives.     

The influence of juvenile hormone analogue on soldier differentiation in the higher termite,Macrotermes michaelseni*

ABSTRACT. The influence of topically-applied or vapour-administered juvenile hormone analogue (ZR-515, methoprene (JHA)), on soldier differentiation in a higher termite species, Macrotermes michaelseni (Sjostedt) (Isoptera; Macro-termitinae) was examined. It was found that even male third larvae, which under natural conditions develop exclusively into major workers, could be induced by JHA to differentiate into presoldiers. Intermediate forms between workers and soldiers were produced from some of the treated individuals. Administered in vapour form, JHA induced a higher proportion of presoldiers and a lower proportion of intermediate forms than it did when administered topically. The rate of presoldier formation was found to be JHA-dose-dependent. Biometric analysis showed that male presoldiers have the largest head capsule size, but the smallest mandibular index. The possible mechanism of soldier induction is discussed.     

A Test of Genital Allometry Using Two Damselfly Species does not Produce Hypoallometric Patterns

It is widely admitted that sexual selection is the responsible force behind genital traits. However, the particular mechanisms of genital evolution are still debated. Recently, studies of genital static allometry in insects have been used to elucidate such mechanisms. Insect genital traits are often reported to show negative allometry (i.e., a slope < 1), which has generated a number of ideas on how genital traits are selected. However, many studies that have inferred selection mechanisms have omitted consideration of the function of genital traits, used unreliable indicators of body size, and only rarely included female genitalia in their analysis. We investigated whether negative allometry operates for genitalia in two damselfly species (Protoneura cara and Ischnura denticollis). Damselflies are suitable for genital allometry tests as their genital function and body size indicators (wing length and head width) are relatively well known and established. First, we show that the aedeagus is used to physically remove sperm from both sperm storage organs (bursa and spermatheca) and that wing length and head width correlate positively with other morphological traits for the two study species. Second, we estimated genital allometry by measuring aedeagal length, vaginal length, bursal volume, and spermathecal volume. Our results indicate no consistent allometric pattern. Allometry for aedeagal length and vaginal width was not the same. Thus, there was no support for a negative allometric relationship. We urge researchers investigating allometry to look directly at how genitalia function rather than inferring function from allometric relationships only.     

Phenotypically plastic traits regulate caste formation and soldier function in polyembryonic wasps

Polyembryonic encyrtid wasps are parasitoids that have evolved a clonal form of embryogenesis and a caste system where some progeny become reproducing wasps whereas others develop into a sterile soldier caste. Theory based on the biology of Copidosoma floridanum predicts that the primary role of soldier larvae is to mediate conflict over sex ratio, which also favours female‐biased soldier production. Other data, however, suggest that female‐biased soldier production reflects a developmental constraint. Here, we assessed whether female‐biased soldier function by polyembryonic wasps reflects sex‐specific adaptation or constraint by conducting comparative studies with Copidosoma bakeri, a species that produces clutch sizes similar to C. floridanum yet rarely produces broods associated with sex ratio conflict. Our results indicate that the oviposition behaviour of adults, development of progeny and function of soldier larvae differ greatly between C. bakeri and C. floridanum. These findings indicate that caste formation and soldier function in polyembryonic encyrtid wasps are regulated by phenotypically plastic traits. Our results further suggest that the primary function of the soldier caste in some species is defence of host resources from competitors whereas in others it is the resolution of sex ratio conflict.     

Social aphids use their antennae to perceive density cue for soldier production

In social insects, local interactions among colony members facilitate information transfer, and allow the whole colony to regulate division of labor and task allocation in an integrated and coordinated manner. In particular, regulation of caste differentiation in response to external cues is important for sustaining social insect colonies. The social aphid Tuberaphis styraci exhibits a caste polyphenism, producing second‐instar soldiers and non‐soldiers. Previous studies using an artificial diet rearing system identified high aphid density as a crucial cue for soldier production, which acts on embryos in the maternal body and newborn first‐instar nymphs to induce soldier differentiation. While direct contact stimuli from live non‐soldiers were suggested to mediate the density effect, how the aphids perceive the stimuli has been unknown. Here we investigated how antennal removal of adult females affects the soldier production in T. styraci. Under a high density condition, intact females produced the highest percentage of soldiers, females deprived of both antennae produced the lowest percentage of soldiers, and females deprived of one antenna exhibited an intermediate percentage of soldiers. Scanning electron microscopic observations of the aphids revealed the existence of sensory organs for chemoreception and tactile sensation on the antennae of the mother aphids. These results indicate that T. styraci females use their antennae to perceive soldier‐inducing density cue, suggesting that maternal perception of density cue is involved in regulation over caste phenotype of their offspring.     

The metabolic enzyme phosphoglucose isomerase (Pgi) affects the outcome of intra‐specific competition in a polyembryonic wasp

1. Polyembryonic parasitoid wasps in the family Encyrtidae (Hymenoptera) have evolved a caste system consisting of morphologically and functionally distinct larvae called soldiers and reproductives. 2. Two selective pressures are thought to underlie the evolution of the soldier caste: defence against competitors and resolution of the sex ratio conflict. Previous studies also indicate that soldier development time strongly affects the outcome of intra‐specific competition in the polyembryonic encyrtid Copidosoma floridanum Ashmead. This study builds on prior findings by showing that alleles of the metabolic enzyme phosphoglucose isomerase (Pgi) differentially affect soldier development time and the outcome of competition. 3. Soldier larvae with the Pgi alleles 100 or 120 emerged on average 65 h post‐parasitism, whereas soldier larvae with a third allele, 54, emerged at 67 h. In turn, C. floridanum broods homozygous for the 100 and 120 alleles outcompete broods homozygous for the 54 allele. 4. Pgi allelic diversity may be maintained through a life‐history trade‐off affecting female brood sizes with homozygous broods bearing the developmentally disadvantageous 54 allele producing more adult females than broods bearing alternate common alleles.