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.     

Higher aggression towards closer relatives by soldier larvae in a polyembryonic wasp

In the polyembryonic wasp Copidosoma floridanum, females commonly lay one male and one female egg in a lepidopteran host. Both sexes proliferate clonally within the growing host larva. Distinct larval castes develop from each wasp egg, the majority being ‘reproductives’ plus some ‘soldiers’ which sacrifice reproduction and attack competitors. Maturing mixed sex broods are usually female biased, as expected when intra-brood mating is common. Pre-mating dispersal followed by outbreeding is expected to increase sexual conflict over brood sex ratios and result in greater soldier attack rates. Owing to sexually asymmetric relatedness, intra-brood conflicts are expected to be resolved primarily via female soldier attack. We observed soldier behaviour in vitro to test whether lower intra-brood relatedness (siblings from either within-strain or between-strain crosses were presented) increased inter-sexual aggression by female as well as male soldiers. As found in prior studies, females were more aggressive than males but, contrary to expectations and previous empirical observations, soldiers of both sexes showed more aggression towards more closely related embryos. We speculate that lower intra-brood relatedness indicates maternal outbreeding and may suggest a rarity of mating opportunities for reproductives maturing from the current brood, which may enhance the value of opposite sex brood-mates, or that higher aggression towards relatives may be a side-effect of mechanisms to discriminate heterospecific competitors.     

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.     

Interspecific competition between the polyembryonic wasp Copidosoma floridanum and the gregarious endoparasitoid Glyptapanteles pallipes

We examined interspecific competition between the egg-larval polyembryonic parasitoid Copidosoma floridanum (Hymenoptera: Encyrtidae) and the gregarious larval endoparasitoid Glyptapanteles pallipes (Hymenoptera: Braconidae). Approximately 72% of multiparasitized Acanthoplusia agnata (Lepidoptera: Noctuidae) produced C. floridanum adults, 14% produced G. pallipes adults, and 14% died without producing any parasitoids. Development was delayed and the rate of weight gain was reduced in multiparasitized hosts that produced C. floridanum compared with singly-parasitized hosts. In contrast, only weight was reduced in multiparasitized hosts that produced G. pallipes adults. Compared with single parasitism, the brood size of wasps emerging from multiparasitized hosts was reduced in both species. The percentage of hosts containing precocious larvae of C. floridanum, which are considered to be soldiers, did not increase in response to parasitization by G. pallipes. However, developmental cessation and death of G. pallipes eggs and larvae may be closely related to the number of coexisting precocious larvae.     

Reproductive biology and related host-parasitoid interactions between the gypsy moth,Lymantria dispar and Glyptapanteles flavicoxis,a gregarious endoparasitoid*

The mean total progeny produced by female Glyptapanteles flavicoxis was significantly greater when the parasitoid attacked 4th instar gypsy moth rather than younger larvae. The development of parasitoids within older instars was faster than that in younger hosts. The proportion of female progeny was not altered by the host instar parasitised. The total reproductive output per female G. flavicoxis increased as the number of host larvae increased. However, since the reproductive output was spread among more available hosts, the reproductive output per host declined as the number of host larvae increased. There was no trend in the relationship between total progeny produced or the proportion of female progeny and the number of matings of parental females.     

Reproductive schedule and factors affecting soldier production in the eusocial bamboo aphid Pseudoregma bambucicola (Homoptera, Aphididae)

Summary: The reproductive characteristics of the soldier-producing aphid Pseudoregma bambucicola were studied in Kagoshima, Southern Japan, to know the factors affecting soldier production of eusocial aphids. The soldier proportion in aphid colonies was highest from October to November. In some large colonies, soldiers were observed in all seasons except in July when colony size was relatively small. Multiple regression analysis showed that the colony size was a principal factor affecting soldier proportion throughout a year. Other social or environmental factors such as aphid composition, host plant conditions and predator abundance were not always significant. Rearing experiments revealed that large colonies (̿,000 individuals) produced soldiers in almost all seasons while small colonies (<1,000) never produced any soldiers. The caste-production schedule of adult females was examined in the field. When solitary females produced both castes, they usually produced normal nymphs first and then soldiers. Females from large colonies tended to produce more soldiers in the earlier period of their lifetime, whereas females from newly established small colonies produced no or only a few soldiers at later times. The average number of soldiers and normal nymphs produced consecutively by a single female was >10 and >20, respectively. Because they have a small number of ovarioles (<15 on average), females should alter caste production within the same ovarioles according to changes in environmental conditions. Artificial removal or introduction of predators and reduction of colony size did not affect soldier production over two successive generations, revealing maternal effects on soldier production. Females cannot shift caste production quickly in response to changes in predator abundance and colony size. This is probably due to early developmental determination of castes within the mother's body.     

Mutual interference in the mass-reared fruit fly parasitoid,Diachasmimorpha longicaudata (Hymenoptera: Braconidae)

Diachasmimorpha longicaudata is a solitary endoparasitoid produced in Mexico for the biological control of Anastrepha fruit flies (Diptera: Tephritidae). We studied the effect of mutual interference among conspecific foraging females to better understand the parasitoid-host dynamics established in the mass-rearing system of this species. We used a constant host availability of 60 third instar larvae of Anastrepha ludens (per oviposition unit type Petri dish) that were individually exposed for 3 h to 1, 2, 4, 8, 10, 14, and 20 D. longicaudata females, seven days old without oviposition experience. The following parameters were evaluated: total number of attacked hosts (i.e., larvae with at least one oviposition scar), number of attacked hosts per female, adult emergence percentage, and female proportion. We also performed a second assay at mass-rearing level, contrasting the current proportion used in mass-rearing (~?two host larvae per female) with the alternative treatments (5 and 7.5 host larvae per female). Results showed that the density of females foraging on a patch exhibits an inverse relationship to the number of hosts attacked by one female. The highest values for adult emergence (73.4%), patch exploitation (94.6%), and female proportion (0.86) were obtained in the treatments with 7.5, 6, and 3 host larvae per female, respectively. Under mass-rearing conditions, we found that a lower density of females per cage (~?five larvae per female) reduced superparasitism levels without affecting adult emergence and the proportion of females. This suggests that the mass-rearing efficiency of D. longicaudata could be improved by reducing the density of foraging females.

     

Reproductive Biology and Behavior of Rhabditis pellio, (Schneider) (Rhabditida: Rhabditidae)

Laboratory studies were conducted on the mating behavior of Rhabditis pellio males and females, which were maintained on a culture of Flavobacterium sp. bacteria isolated from earthworms. The mean time that elapsed between first contact of the sexes and their ultimate separation was 23.2 min. However, only 5.0 min were required for copulation (the interval during which male spicules were inserted into the female vagina). Three-day-old females that were permitted to mate once on their first day of adult life produced only one-third as many larvae as did females that were permitted unlimited mating. However, the longevity of females was found to decrease with an increase in the number of matings. Both males and females that were permitted to mate daily produced the greatest number of offspring when they were 4 days old. When the initiation of mating was delayed beyond their third day of life, the number of larvae produced by females decreased. In approximately one-half of the copulations, males failed to inseminate their female partners.     

Sperm limitation affects sex allocation in a parasitoid wasp Nasonia vitripennis

Insect reproduction is influenced by various external factors including temperature, a well-studied constraint. We investigated to what extent different levels of sperm limitation of males exposed to different heat stresses (34 and 36℃) afFect fem ales' offspring production and sex allocation in Nasonia vitripennis. In this haplodiploid parasitoid wasp attacking different species of pest flies, we investigated the effect of the quantity of sperm females received and stored in their spermatheca on their sperm use decisions, hence sex allocation, over successive ovipositions. In particular, we compared the sex allocation of females presenting three levels of sperm limitation (i.e.,mated with control, 34 ℃ heat-stressed or 36℃heat-stressed males) on each host they parasitized. To disentangle the potential reduction of sperm quality after a heat stress exposure from that of sperm quantity, we also explored the clutch size and sex ratio produced by fem ales that were partially sperm limited after copulating with multiply mated males. Independently of their sperm numbers, all types of fem ales produced a similar total number of offspring, but the more limited ones had fewer daughters. Sperm limitation further affected the distribution of daughters' production across time.In addition to constraints acting on female physiology, male fertility should therefore be considered in studies measuring reproductive outputs of insects submitted to heat stresses.     

Effect of differential host exposure on reproduction in the solitary ectoparasitoid Eupelmus vuilleti

We examined self-superparasitism in Eupelmus vuilleti (CRW.), a solitary ectoparasitoid of larvae of Callosobruchus maculatus (F.) which develop in cowpea seeds. The reproduction of isolated females was analysed in the presence of seeds containing 10 or 30 hosts for 24, 48, and 72 h. At the two host densities, female reproductive activity increased with host exposure time, but variations in offspring number per female were limited. The propensity of the females to self-superparasitise their hosts after long exposure times could explain these results. The offspring sex ratio was female biased but tended to 0.5 for longer exposure times. The absence of host renewal for a 3-day period was followed by an increase in self-superparasitism which reduced the parasitoid survival rate and by emergence of a lower number of females in the offspring. The offspring production was then analysed when females were provided with ten hosts a day or with 30 hosts every three days throughout their life span. The reduction of host renewal frequency did not affect male production but caused a reduction in the number of females emerging from the seeds. The importance of a high rate of self-superparasitism on the population biology of this parasitoid is discussed.     

Performance of Sclerodermus brevicornis,a parasitoid of invasive longhorn beetles,when reared on rice moth larvae

Biological control efficiency can be improved by developing effective mass‐rearing systems to produce large numbers of high‐quality parasitoids. This study explored an alternative host for rearing Sclerodermus brevicornis (Kieffer) (Hymenoptera: Bethylidae), a potential biocontrol agent for the suppression of exotic and invasive wood‐boring longhorn beetle (Coleoptera: Cerambycidae) populations in the European agroforestry ecosystems. We tested larvae of the rice moth, Corcyra cephalonica Stainton (Lepidoptera: Pyralidae), as host for the parasitoid. We quantified the probability and timing of host attack and parasitism as well as reproductive success, offspring production, and the characteristics of adult offspring. As S. brevicornis is a quasi‐social species (multiple females, communally produced offspring broods), we also explored the effects of varying the number of females to which individual hosts were presented, with the aim of determining the optimal female‐to‐host ratio. As time to host attack can be a limiting factor in S. brevicornis rearing protocols, we tested the use of adult females of another bethylid species, Goniozus legneri Gordh, to paralyse C. cephalonica larvae prior to presentation. We identified the conditions within our experiment that maximized offspring production per host and offspring production per adult female parasitoid. We found that C. cephalonica is suitable as a factitious host and, as it is considerably more straightforward for laboratory rearing than cerambycid species, it is a good candidate for adoption by future S. brevicornis mass‐rearing and release programmes.     

Bone-eating Osedax females and their 'harems' of dwarf males are recruited from a common larval pool

Extreme male dwarfism occurs in Osedax (Annelida: Siboglinidae), marine worms with sessile females that bore into submerged bones. Osedax are hypothesized to use environmental sex determination, in which undifferentiated larvae that settle on bones develop as females, and subsequent larvae that settle on females transform into dwarf males. This study addresses several hypotheses regarding possible recruitment sources for the males: (i) common larval pool--males and females are sampled from a common pool of larvae; (ii) neighbourhood--males are supplied by a limited number of neighbouring females; and (iii) arrhenotoky--males are primarily the sons of host females. Osedax rubiplumus were sampled from submerged whalebones located at 1820-m and 2893-m depths in Monterey Bay, California. Immature females typically did not host males, but mature females maintained male 'harems' that grew exponentially in the number of males as female size increased. Allozyme analysis of the females revealed binomial proportions of nuclear genotypes, an indication of random sexual mating. Analysis of mitochondrial DNA sequences from the male harems and their host females allowed us to reject the arrhenotoky and neighbourhood hypotheses for male recruitment. No significant partitioning of mitochondrial diversity existed between the male and female sexes, or between subsamples of worms collected at different depths or during different years (2002-2007). Mitochondrial sequence diversity was very high in these worms, suggesting that as many as 10(6) females contributed to a common larval pool from which the two sexes were randomly drawn.     

Presence of soldier larvae determines the outcome of competition in a polyembryonic wasp

Soldier‐producing polyembryonic waSPS are the only social animals that develop as parasites inside the bodies of other insects. Characterizing the kin composition of broods is central to understanding the evolution of the soldier caste in these unique social insects. Here we studied the role of soldiers in mediating the outcome of competition among clones of the polyembryonic wasp Copidosoma floridanum. Soldier‐producing female clones usually monopolized host resources, whereas soldierless male clones usually coexisted in hosts. Behavioural experiments further indicated that early‐emerging soldiers are specialized to combat intraspecific competitors and later‐emerging soldiers are specialized for defence against interspecific competitors. Taken together, our results point to intraspecific competition as a major selective force in the evolution of the soldier caste. Our data also present an evolutionary conundrum: given the benefit of soldiers, why are male clones functionally soldierless?     

Floral display, pollinator visitation and reproductive success in the dioecious perennial herb Wurmbea dioica (Liliaceae)

Floral traits that increase attractiveness to pollinators are predicted to evolve through selection on male function rather than on female function. To determine the importance of male-biased selection in dioecious Wurmbea dioica, we examined sexual dimorphism in flower size and number and the effects of these traits on pollinator visitation and reproductive success of male and female plants. Males produced more and larger flowers than did females. Bees and butterflies responded to this dimorphism and visited males more frequently than females, although flies did not differentiate between the sexes. Within sexes, insect pollinators made more visits to and visited more flowers on plants with many flowers. However, visits per flower did not vary with flower number, indicating that visitation was proportional to the number of flowers per plant. When flower number was experimentally held constant, visitation increased with flower size under sunny but not overcast conditions. Flower size but not number affected pollen removal per flower in males and deposition in females. In males, pollen removal increased with flower size 3 days after flowers opened, but not after 6 days when 98% of pollen was removed. Males with larger flowers therefore, may have higher fitness not because pollen removal is more complete, but because pollen is removed more rapidly providing opportunities to pre-empt ovules. In females, pollen deposition increased with flower size 3 days but not 6 days after flowers opened. At both times, deposition exceeded ovule production by four-fold or more, and for 2 years seed production was not limited by pollen. Flower size had no effect on seed production per plant and was negatively related to percent seed set, implying a tradeoff between allocation to attraction and reproductive success. This indicates that larger flower size in females is unlikely to increase fitness. In both sexes, gamete production was positively correlated with flower size. In males, greater pollen production would increase the advantage of large flowers, but in females more ovules may represent a resource cost. Selection to increase flower size and number in W. dioica has probably occurred through male rather than female function. Received: 15 June 1997 / Accepted: 12 February 1998     

Bionomics of Opius bellus (Hymenoptera: Braconidae), an endoparasitoid of Anastrepha fraterculus (Diptera: Tephritidae) in fruit-growing areas of Northwestern Argentina

Opius bellus is a neotropical larval-prepupal parasitoid known to attack the pestiferous fruit fly, Anastrepha fraterculus. Due to interest in the use of native parasitoids in forthcoming fruit fly biocontrol programmes in Argentina, O. bellus was colonised for the first time using laboratory-reared A. fraterculus larvae. A series of experiments were conducted to (1) best achieve an efficient parasitoid rearing by determining optimal larval host age, host:parasitoid ratio and host exposure time and (2) assess their potential as biological control agents by determining reproductive parameters. The most productive exposure regimen was: 7–9 d-old (early and middle third-instars) A. fraterculus larvae for 4 h at a 4:1 host:parasitoid ratio; this array of factors was sufficient to achieve the highest average adult emergence (48%) and an offspring sex ratio at equitable proportion. Increasing both host:parasitoid ratio further than 4:1 and the host exposure time beyond 4 h did not significantly enhance parasitoid female offspring yield. Females produced eggs for 29.5 ± 1.4 days. At 32 days of age, 50% of the females were still alive. The majority of the progeny were produced by females between 20 and 24 d-old. At 26°C, gross fecundity rate, net reproductive rate, intrinsic rate of increase and mean generation time were 20.7 ± 4.2 offspring/female, 9.6 ± 2.5 females/newborn females, 0.06 ± 0.01 females/female/day and 8.4 ± 0.2 days, respectively. The long lifespan and reproductive parameters suggest that this parasitoid species has suitable attributes for mass-rearing.     

Does aggregation behavior of codling moth larvae,Cydia pomonella,increase the risk of parasitism by Mastrus ridibundus?

Mastrus ridibundus is a specialist hymenopteran parasitoid that parasitizes last-instar larvae or prepupae of the codling moth, Cydia pomonella. Foraging females eavesdrop on an aggregation pheromone produced by cocooning larvae. We investigated whether larvae that cocoon in aggregation experience a greater rate of parasitism than larvae that cocoon in isolation. In wind tunnel experiments, 10 larvae in aggregations were more readily located by female M. ridibundus than 10 larvae well separated from each other. Similarly, aggregations of 30 larvae were more attractive to female M. ridibundus than those of 3 larvae. In cage experiments, larval cocooning in aggregation or isolation had no effect on the mean rate of parasitism and the mean number of eggs deposited per parasitized host. In Petri-dish experiments, the location of larvae within an aggregation significantly affected their rate of parasitism, with those in the center of an aggregation completely shielded from parasitism. Our data suggest that aggregation behavior by C. pomonella larvae does not appear to increase the rate of parasitism. The increased risk of aggregated larvae to be detected by M. ridibundus is likely offset by diluted parasitism risk and structural refugia effects that larvae in aggregation experience. As an egg-limited parasitoid, female M. ridibundus can parasitize on average only one larva in an aggregation, with the likelihood of parasitism for each larva being inversely proportional to the number of larvae in that aggregation.     

Costs and consequences of superparasitism in the polyembryonic parasitoid Copidosoma koehleri (Hymenoptera: Encyrtidae)

Abstract.  1. Polyembryonic wasps provide dramatic examples of intra-specific developmental conflict. In these parasitoids, each egg proliferates into a clonal lineage of genetically identical larvae. If more than one egg is laid in a host (superparasitism), individuals of different clones may compete for food resources.
2. In the polyembryonic encyrtid Copidosoma koehleri , one larva per clone can differentiate into a sterile soldier. It is shown that soldiers are always females, and that they attack intra-specific competitors.
3. Research hypotheses were that (a) clones that develop in superparasitised hosts suffer heavier mortality than clones that develop in singly parasitised hosts, and (b) female clones cause higher mortality to their competitors than male clones, hence larval survival is lower in superparasitised hosts that contain females than in male-only broods.
4. The potential frequency of superparasitism in C. koehleri was manipulated by varying parasitoid–host ratios and exposure durations.
5. As parasitoid densities and exposure durations increased, the frequency of superparasitism rose, brood sizes increased, but the number of hosts that completed development was reduced. The number of offspring per parasitoid female decreased with increasing parasitoid–host ratios. Offspring size and longevity were inversely correlated with brood size. As superparasitism rates increased, fewer all-male broods were produced. Male–female broods were female-biased, suggesting selective killing of males by female soldiers. All-female broods were significantly smaller than all-male broods at high parasitoid densities only, possibly reflecting aggression among soldiers of competing clones.
6. The results support the working hypotheses, and suggest that female larvae outcompete males in superparasitised hosts.     

Developmental patterns in the polyembryonic parasitoid wasp Copidosoma koehleri

Polyembryony is a unique mode of development in which multiple genetically identical embryos develop from a single egg. In some polyembryonic species a proportion of the embryos develop into soldier larvae, which attack competitors in the host. We studied the development of the polyembryonic wasp Copidosoma koehleri in its host Phthorimaea opercullela. We dissected hosts parasitized by either virgin or mated female wasps at 2day intervals from hatching to the final instars. We documented host mass and head width, the number and size of developing wasps and the presence of a soldier larva. Additionally, we kept a sample of parasitized hosts until emergence of wasps and measured the head width of emerging adults. We characterized wasp development in relation to host development. One half of the broods produced by mated wasps contained one soldier larva throughout development. This suggests that in C. koehleri each female brood produces a single soldier larva, and that the soldier probably survives and grows gradually during host development. Additionally, we found that female broods were larger than male broods during development and also upon emergence. Accordingly, body size was larger for males during development as well as upon emergence. These findings may extend the existing knowledge on polyembryonic development in general, and serve as a baseline for further experiments.