Patch Defence in the Parasitoid Wasp Trissolcus basalis (Insecta: Scelionidae): The Time Structure of Pairwise Contests,and the ‘Waiting Game’

Aggressive defence of host patches has been reported in many parasitoid wasps, but rarely examined in quantitative detail. One aspect of interest is that foraging female parasitoids do not simply consume resource patches, they invest offspring in them. Therefore, patch defence in parasitoids can involve not only resource defence prior to oviposition, but also postoviposition defence of offspring (maternal care). In this paper, the time-structure and sequence of pairwise agonistic contests between females of the parasitoid Trissolcus basalis (Wollaston) (Hymenoptera: Scelionidae) are analysed. Three main periods were evident in contests. In the first period, both females exploited the patch with no aggression. After the initiation of fighting, they entered a ‘contest period’, during which resident and intruder roles became clearly resolved. The resident then usually guarded the patch for up to several hours before leaving. This signalled the beginning of the third period, in which the intruder returned to superparasitise the patch. During the contest period, resident behaviour initially reflected the trade-off between exploiting fresh hosts, and defending those it had already parasitised from the intruder, which persistently returned to the patch to try and oviposit, with some success. However, when the patch became fully parasitised, both resident and intruder switched to a ‘waiting game’, in which they sat motionless for extended periods, the resident on the patch and the intruder at a distance. These stand-offs were punctuated by occasional aggressive patrolling by the resident, and cryptic returns to the palch by the intruder. This waiting game appears to be an informational war of attrition, suggesting a conceptual basis for modelling patch-leaving decisions using evolutionary game theory.     

Fine structure of the preocellar pit in Trissolcus basalis (Woll.) (Hymenoptera : Scelionidae)

The preocellar pit of both sexes of Trissolcus basalis (Hymenoptera : Scelionidae), a solitary egg parasitoid of Nezara viridula, is described. Externally, the opening is median, spherical, with an average diameter 12 μm Internally, the preocellar pit corresponds to a bell-shaped apodeme (27 μm deep) with smooth cuticular walls, except for the apical portion, which is rounded and roughly sculptured. Ultrastructural observations show that the preocellar apodeme apex is connected to the protocerebrum by a bundle of elongated epidermal cells, which are very rich in microtubules extending from the base to the cell apex. It is assumed that the apodeme and the epidermal cell bundle act as a suspension to support the brain. The taxonomic significance of this pit is recognized.     

Source of the host marking pheromone in the egg parasitoid Trissolcus basalis (Hymenoptera: Scelionidae)

After oviposition, Trissolcus basalis females always mark the host's surface, depositing host marking substances for herself and to warn other ovipositing females. The perception of these host marking substances, probably through the antennae, can induce the female to leave and seek healthy hosts. Parasitoid females exposed to conspecific parasitized egg masses left the host egg masses significantly more often than when exposed to non-parasitized egg masses. More egg mass leaving behavior also was observed when the egg masses were treated with Dufour's gland secretion but not when treated with secretion from the common oviducts. The common oviduct has a secretory epithelium that produces electron-dense vesicles, probably containing proteinaceous substances. The secretory cells of the accessory gland, Dufour's gland, contain electron-lucid vesicles, whose secretion appears to be a lipid similarly to that found in pheromone secreting glands. Ultrastructural and behavioral evidence suggests that Dufour's gland is the host marking pheromone source.     

Cold storage of the egg parasitoids Trissolcus basalis (Wollaston) and Telenomus podisi Ashmead (Hymenoptera: Scelionidae)

Adults of Trissolcus basalis and Telenomus podisi were stored either at 15 or 18 °C after their immature development had been completed at 18 or 25 °C. Longevity of the parasitoids in the storage temperatures was evaluated, as well as fecundity and longevity following their return to 25 °C after different periods in reproductive diapause. Temperature during immature development influenced female longevity and highest mean longevity was obtained for females that developed to the adult stage at 25 °C and then were stored at 15 °C (ca. 13 months for T. basalis and 10 months for Te. podisi). For adults of T. basalis that developed at 25 °C, storage periods of 120 or 180 days at 15 or 18 °C did not affect fecundity. The fecundity of T. basalis females that developed at 18 °C and were stored for 120 days at 15 or 18 °C was not affected; however, after remaining for 180 days, fecundity was reduced in ca. 30 and 50%, respectively. Storage of Te. podisi adults at 15 or 18 °C significantly reduced fecundity. It is concluded that adults of T. basalis can be stored in the adult stage at 15 or 18 °C between two soybean crop seasons for mass production purposes, aiming the biological control of stink bugs.     

Biological control and the species status of two host-associated populations of Trissolcus basalis (Wollaston) (Hymenoptera: Scelionidae).

Abstract Populations of the morphologically defined taxon Trissolcus basalis (Wollaston) from two major hosts (Nezara viridula [L.] and Agonoscelis rutila [F.]) were tested, using biological criteria, to determine whether they represented populations of one genetical species, or whether they represented populations of two host-associated sibling species. Individuals from both hosts exhibited no obvious courtship differences and they selected mates at random in mate-choice experiments. In addition, female wasps produced offspring of both sexes after mating with a male from either population. We conclude that the individuals of the two populations tested constitute a single genetical species.     

Mating Behavior of Trissolcus basalis (Wollaston) (Hymenoptera: Scelionidae): Potential for Outbreeding in a Predominantly Inbreeding Species

The mating behavior of the quasi-gregarious egg parasitoid Trissolcus basalis (Wollaston) was investigated under field conditions. Trissolcus basalis has female-biased sex ratios and is a protandrous species, with males emerging 1–2 days before females. Males competed aggressively for control of the egg mass, with one male assuming dominance and control of the egg mass, although changes in dominance occurred at least once on each egg mass observed. Typical mating behavior involved the dominant male mating his sisters immediately upon their emergence from the egg mass. These behaviors are characteristic of an inbreeding species that manifests local mate competition. However, several aspects of the mating behavior of T. basalis are inconsistent with that of an inbreeding species. Over 18% of emerging females were not mated by the dominant male upon emergence, 13% of females were not observed to be mated at all and may have left their natal site as virgins, 25% of females were mated multiple times and sometimes by multiple males, females remained near the natal site for up to several hours after emergence before emigrating, and males dispersed away from the natal site during female emergence. Trissolcus basalis may be a predominantly inbreeding species but its emergence and mating behavior suggest that low-frequency outbreeding is also likely to occur.     

Molecular phylogeny of Trissolcus species (Hymenoptera: Scelionidae)

Trissolcus species (Hymenoptera: Scelionidae) are the most promising biological control agents against sunn pest. The accurate identification of natural enemies is crucial in order to develop successful biological control programs. This paper presents phylogenetic analyses of Trissolcus species based on data sets consisting of 18S, 28S, ITS1, ITS2 and cytochrome oxidase subunit I (COI) genes. The most commonly used genetic loci in Trissolcus species identification is the cytochrome oxidase I gene (COI). Also restriction fragment length polymorphism analyses of PCR amplified COI gene have been developed to discriminate closely related species. We suggest that Trissolcus grandis Thompson and Trissolcus semistriatus Nees split significantly into two different genetic groups while there is another species diverging from T. semistriatus which is decsribed as ‘Trissolcus flavotibialis Kocak & Guz, n. sp.’     

The Effect of Seasonal Changes on Nezara viridula (L.) (Hemiptera: Pentatomidae) and Trissolcus basalis (Wollaston) (Hymenoptera: Scelionidae) in Hawaii

The effect of seasonal changes in temperature and photoperiod on the interaction of Nezara viridula (Hemiptera: Pentatomidae) and its egg parasitoid Trissolcus basalis (Hymenoptera: Scelionidae) was investigated in the laboratory. We found no evidence of reproductive diapause in N. viridula under simulated Hawaiian summer and winter conditions. Further, although “diapause” coloration was obtained in the laboratory, it was not correlated with reproductive status. Studies of the survival of T. basalis provided with honey under the same simulated conditions showed that under summer conditions, only 2.1% of the female and 13.5% of the male population were still alive by 60 days. When provided with N. viridula egg masses at 30 days, 79.4% of the eggs were parasitized in a 3-h period. Under winter conditions, 54.3, 28.3, and 14.5% of the females were alive at 30, 60, and 90 days after adult emergence. When provided with N. viridula egg masses at 30, 60, and 90 days for 3 h, 57.6, 32.8, and 47.1% of the eggs were successfully parasitized. These studies suggest the limiting factor in the interaction of T. basalis and N. viridula is not reproductive diapause, but instead the ability of T. basalis to survive summer conditions.     

Morphology of the antennal sex-gland in male Trissolcus basalis (Woll.) (Hymenoptera : Scelionidae), an egg parasitoid of the green stink bug,Nezara viridula (Hemiptera : Pentatomidae)

The 5th antennomere, beginning with the scape, of the male Scelionidae antenna has been referred to as a sex-segment due to its morphological and presumed sexual function. A study of the sex-segment of the parasitoid, Trissolcus basalis (Woll.) (Hymenoptera : Scelionidae) revealed a single ventral peg that is involved in releasing a secretion from an exocrine gland located in the antennomere. This is a “class 3” exocrine gland and the secretion appears to be partially proteinaceous since it is susceptible to protease digestion. Although the function of the sex-segment and gland remains unknown, the suggestion that it is involved in courtship is supported. Some sex-segment processes of other Scelionidae are also considered.     

Three new species of Trissolcus ashmead (Hymenoptera: Platygastroidea: Scelionidae) from Bulgaria

Identification key of 20 species of the genus Trissolcus registered in Bulgaria is given. Three species, T. fuscus sp. n., T. simplex sp. n. and T. nigricans sp. n. are described as new for science.     

Host Age Response in the Parasitoid Wasp Spalangia cameroni (Hymenoptera: Pteromalidae)

Female Spalangia cameroni produced more offspring from younger house fly pupae, both when given a choice of host ages and when not given a choice. Host age did not affect offspring survivorship. Offspring were larger when they had developed on younger hosts and the effect was independent of offspring sex. Having previously parasitized old hosts versus young hosts did not reduce a female's production of offspring in subsequent hosts. Females distinguished between young and old hosts both in the light and in the dark. Females did not distinguish between host ages prior to physical contact with the host but could distinguish by the time they first began exploring a host by tapping it with their antennae; thus, they could distinguish before drilling into a host.     

New genera and species of Scelionidae (Insecta: Proctotrupoidea) from Rovno amber

Two new genera and three new species of parasitic wasps from the family Scelionidae are described from the Late Eocene Rovno amber: Pseudotelea gracilis Kononova, gen. et sp. nov. (subfamily Scelioninae), Pseudidris striatus Kononova, gen. et sp. nov., and Ceratobaeoides cornutus Kononova, sp. nov. (subfamily Baeinae). The new taxa are compared with representatives of the Recent fauna.     

Ratio-dependent parasitism with Trissolcus basalis (Wollaston) (Hymenoptera: Scelionidae) on egg rafts of Nezara viridula (Linnaeus) (Hemiptera: Pentatomidae): effect of experimental variables and compatibility of 'ratio' and 'Holling' models

Abstract Egg rafts of Nezara viridula were exposed to the parasitoid wasp Trissolcus basalis in experimental arenas to establish the relationship of the rates of attack and parasitism to various combinations of arena size, parasitoid density, host density and parasitoid-to-host ratio. Arena sizes were varied in the ratio 1:9:63, with the largest having a search area of 1.44 m². Parasitoid and host densities were varied over a 27-fold range. The parasitoid-to-host ratios used were 1:1, 3:1 and 6:1. Finding time was related to a constant factor (flight propensity), rather than to the difficulty of finding (density of hosts). Initial attack rates were therefore related only to parasitoid numbers (or density), even at the lower densities and ratios. Parasitism rates (a function of attack rate per host) were thus also strongly related to parasitoid to host ratio, regardless of densities used and arena sizes. Even reducing host density, while keeping time and parasitoid density constant, increased the parasitism rate. A ratio model for parasitism rate was therefore compatible with the data but the more explicit Holling 'disc' equation was also compatible because handling time was sufficiently large to make it sensitive to the ratio of parasitoids to hosts for the densities used. We conclude that the two models would predict different results if the density of host egg rafts was in a range below one per square metre.     

Egg-parasitoids of the genus Trissolcus (Hymenoptera,Scelionidae, Telenominae) from the Palaearctic fauna (the flavipes morphological group). 1. New species of the genus Trissolcus

The Trissolcus flavipes species group is revised. A key to the species of this group is provided. In addition to the diagnostic characters, the localities are listed for each species. Eleven new species occurring in various Palaearctic areas are described. Some body structures are illustrated. The morphological characters of the new species are compared.     

First report of chromosomes of the parasitoid wasp Trissolcus basalis (Wollaston) (Hymenoptera: Platygastridae: Telenominae)

Here we present the first data on the chromosome complement of Trissolcus basalis gathered in conjunction with the ongoing whole genome sequencing efforts for this species. The cytogenetic investigation revealed a diploid karyotype with 20 chromosomes, comprising one pair of large metacentrics and nine pairs of acrocentrics that gradually decrease in length.     

Rickettsia Symbionts Cause Parthenogenetic Reproduction in the Parasitoid Wasp Pnigalio soemius (Hymenoptera: Eulophidae)

Bacteria in the genus Rickettsia are intracellular symbionts of disparate groups of organisms. Some Rickettsia strains infect vertebrate animals and plants, where they cause diseases, but most strains are vertically inherited symbionts of invertebrates. In insects Rickettsia symbionts are known to have diverse effects on hosts ranging from influencing host fitness to manipulating reproduction. Here we provide evidence that a Rickettsia symbiont causes thelytokous parthenogenesis (in which mothers produce only daughters from unfertilized eggs) in a parasitoid wasp, Pnigalio soemius (Hymenoptera: Eulophidae). Feeding antibiotics to thelytokous female wasps resulted in production of progeny that were almost all males. Cloning and sequencing of a fragment of the 16S rRNA gene amplified with universal primers, diagnostic PCR screening of symbiont lineages associated with manipulation of reproduction, and fluorescence in situ hybridization (FISH) revealed that Rickettsia is always associated with thelytokous P. soemius and that no other bacteria that manipulate reproduction are present. Molecular analyses and FISH showed that Rickettsia is distributed in the reproductive tissues and is transovarially transmitted from mothers to offspring. Comparison of antibiotic-treated females and untreated females showed that infection had no cost. Phylogenetic analyses of 16S rRNA and gltA gene sequences placed the symbiont of P. soemius in the bellii group and indicated that there have been two separate origins of the parthenogenesis-inducing phenotype in the genus Rickettsia. A possible route for evolution of induction of parthenogenesis in the two distantly related Rickettsia lineages is discussed.The genus Rickettsia contains a group of obligate intracellular symbionts of eukaryotic cells and belongs to the family Rickettsiaceae in the order Rickettsiales of the Alphaproteobacteria (58, 90). Many species have medical importance as they are pathogens of humans and other vertebrates; pathogenic Rickettsia species infect their hosts through blood-feeding arthropods, including lice, fleas, ticks, and mites (51, 80). In addition to Rickettsia species that cause infectious diseases in vertebrates, symbiotic species have been found in disparate groups of organisms, including arthropods, annelids, amoebae, hydrozoa, and plants (53). Rickettsia appears to be especially common in arthropods, having been found in a wide range of taxa in the classes Entognatha (springtails), Insecta (booklice, lice, bugs, leafhoppers, aphids, whiteflies, fleas, flies, lacewings, moths, beetles, and wasps), and Acarina (ticks and mites) (86). However, in most cases, the effect of Rickettsia on the invertebrate host has not been established yet. In general, Rickettsia bacteria are facultative symbionts, but in the booklouse Liposcelis bostrychophila the association is strictly obligate and Rickettsia has an essential role in oocyte development (54, 92). Facultative symbiotic Rickettsia strains have been reported to negatively affect some aspects of host fitness, causing reductions in body weight, fecundity, and longevity in the pea aphid (16, 60, 64), reductions in viability in some blood-feeding arthropod vectors (5, 46), and increased susceptibility to insecticides in the sweet potato whitefly (41). There is also evidence that Rickettsia has positive effects on host fitness, such as a larger body size in infected leeches (40) and a possible role in the oogenesis of a bark beetle (93). Finally, facultative symbiotic rickettsiae can be reproductive parasites of insects. Rickettsia strains are the causal agents of male killing (infected male embryos die) in some ladybird (79, 88) and buprestid leaf-mining (42) beetles. They are also the cause of thelytokous parthenogenesis (in which mothers produce only daughters from unfertilized eggs) in a parasitoid wasp (32). Both kinds of reproductive manipulation bias the host sex ratio toward females and favor the spread of the transovarially inherited Rickettsia strains in the infected populations. In general, Rickettsia is transmitted primarily vertically to host progeny, but in pathogenic species there is concomitant horizontal transmission via intermediate vertebrate hosts, which plays an important role in maintaining the infection in populations of blood-feeding arthropods (53, 57). An exception is Rickettsia prowazekii, the epidemic typhus agent, which spreads only via horizontal transmission in louse host populations (5). Only one Rickettsia is known to be a plant pathogen, and leafhoppers transfer this pathogen horizontally between plants (20). The fact that Rickettsia can be transmitted horizontally and then perpetuated vertically in host descendants has probably been one of the most important factors determining the enormous diversity of Rickettsia symbiotic associations. This point has been emphasized by analyses that have revealed considerable incongruence between Rickettsia and host phylogenies, indicating that horizontal transfer has occurred multiple times over evolutionary timescales (53, 54, 86).In addition to Rickettsia, diverse heritable bacteria are known to manipulate host reproduction to enhance their transmission in arthropods (12, 23). Wolbachia (order Rickettsiales, family Anaplasmataceae), a close relative of Rickettsia (90), is able to induce all known forms of manipulation of reproduction, including cytoplasmic incompatibility, feminization of genetic males, male killing, and parthenogenesis (68). Previously, only Cardinium (Sphingobacteria) has been shown to cause a similar range of reproductive phenotypes, except for male killing (35). The emerging diversity of Rickettsia associated with arthropods (53, 86), combined with evidence that it can manipulate host reproduction in more than one way, suggests that this symbiont may also be a master manipulator.In the Hymenoptera, the dominant mode of reproduction is arrhenotoky; that is, diploid females develop from fertilized eggs, and haploid males develop from unfertilized eggs (76). However, thelytokous parthenogenesis is common, and in some lineages, like the superfamilies Chalcidoidea and Cynipoidea, it is strongly associated with Wolbachia or Cardinium infection (33, 35). Parthenogenesis-inducing (PI) bacteria cause restoration of diploidy in unfertilized haploid eggs, which results in female offspring (28, 50, 69). PI Wolbachia and PI Cardinium also occur in other groups of haplodiploid arthropods, such as mites (82), scale insects (56), and thrips (4). Previously, the only example of PI caused by Rickettsia was found in the eulophid parasitoid wasp Neochrysocharis formosa (1, 32). Besides PI bacteria, uniparental (thelytokous) reproduction in haplodiploid arthropods can also be caused by feminizing bacteria that are able to interact with the host sex determination system and force the development of genotypic males toward functional phenotypic females. To date, only Cardinium has been reported to be a causal agent of feminization in haplodiploid arthropods, and only two examples are known: a mite in which Cardinium causes haploid male embryos to develop as haploid females (18, 83) and a parasitoid wasp in which diploid males are converted to females (27).In this paper, thelytokous reproduction in a parasitoid wasp, Pnigalio soemius (Hymenoptera: Eulophidae), was studied. This wasp, which is probably a complex of cryptic species (8), is a solitary ectoparasitoid that attacks larvae of many leafminer insect species in the orders Coleoptera, Diptera, Hymenoptera, and Lepidoptera (48), some of which are pests of agricultural crops (37, 61). Female P. soemius wasps paralyze host larvae by injection of venom and subsequently lay a single egg next to the host inside a leaf mine; then the parasitoid larva eats the killed host (7). Commonly, P. soemius reproduces biparentally, and the occurrence of thelytoky has not been reported previously. The aims of this study were to determine whether symbiotic bacteria are involved in manipulating the reproduction of P. soemius and then to determine the taxonomic affiliation and phenotype of the manipulators of reproduction discovered. By using antibiotic treatments and karyological analysis of the insect studied, molecular and phylogenetic characterization of the symbiotic bacteria, and detection of intracellular symbionts by means of fluorescence in situ hybridization, it was demonstrated that a PI Rickettsia causes thelytokous reproduction in P. soemius.     

Absence of Complementary Sex Determination in the Parasitoid Wasp Genus Asobara (Hymenoptera: Braconidae)

An attractive way to improve our understanding of sex determination evolution is to study the underlying mechanisms in closely related species and in a phylogenetic perspective. Hymenopterans are well suited owing to the diverse sex determination mechanisms, including different types of Complementary Sex Determination (CSD) and maternal control sex determination. We investigated different types of CSD in four species within the braconid wasp genus Asobara that exhibit diverse life-history traits. Nine to thirteen generations of inbreeding were monitored for diploid male production, brood size, offspring sex ratio, and pupal mortality as indicators for CSD. In addition, simulation models were developed to compare these observations to predicted patterns for multilocus CSD with up to ten loci. The inbreeding regime did not result in diploid male production, decreased brood sizes, substantially increased offspring sex ratios nor in increased pupal mortality. The simulations further allowed us to reject CSD with up to ten loci, which is a strong refutation of the multilocus CSD model. We discuss how the absence of CSD can be reconciled with the variation in life-history traits among Asobara species, and the ramifications for the phylogenetic distribution of sex determination mechanisms in the Hymenoptera.     

Host discrimination modulates brood guarding behaviour and the adaptive superparasitism in the parasitoid wasp Trissolcus semistriatus (Hymenoptera: Scelionidae)

Because hosts utilized by parasitoids are vulnerable to further oviposition by conspecifics, host guarding benefits female wasps. The present study aims to test whether female adults regulate brood guarding behaviour by host discrimination in a solitary parasitoid Trissolcus semistriatus by presenting an intact or parasitized host egg mass to a female adult. Virgin females without oviposition experience have host discrimination ability, which enables them to adjust the number of eggs laid in the hosts. Mating experience increases superparasitism by female adults, whereas mated females achieve a higher discrimination ability as a result of oviposition experience and show a lower superparasitism rate. As expected, females exhibit brood guard after parasitizing an intact host egg mass, whereas those females visiting a previously parasitized host egg mass, do not. Because the survival of eggs in superparasitized hosts is relatively low, regulating brood guarding behaviour by host discrimination is adaptive for female wasps.     

Lethal and sublethal effects of fenitrothion and deltamethrin residues on the egg parasitoid Trissolcus grandis (Hymenoptera: Scelionidae)

Effects of fenitrothion and deltamethrin, the most commonly used insecticides in Iran for controlling Eurygaster integriceps Puton (Heteroptera: Scutelleridae), in wheat and barley were assessed on adults and preimaginal stages of egg parasitoid Trissolcus grandis Thompson (Hymenoptera: Scelionidae). Adult parasitoids exposed to field recommended concentrations of the insecticides suffered 100% mortality within 24 h. LC50 values of fenitrothion and deltamethrin for T. grandis were 8.1 and 3.9 microg (AI) /ml, respectively. Both insecticides and the preimaginal stage of exposure had a significant influence on the level of adult emergence from host eggs treated with field recommended rates. Fenitrothion and deltamethrin reduced the emergence rates by 18 and 34.4%, respectively, compared with the control. However, neither insecticide significantly affected the longevity or reproductive capacity of emerged females, or the sex ratio of their progeny. This study revealed that application of these insecticides should be avoided in early season to conserve natural or released populations of T. grandis. Both insecticides seemed to be detrimental to the parasitoid and need to be applied cautiously through season.