Serratia marcescens as a rapid indicator of Microctonus hyperodae oviposition activity in Listronotus maculicollis and potential application of the technique to host-specificity testing

Listronotus maculicollis (Dietz) (Coleoptera: Curculionidae) is a potential novel host of the braconid parasitoid Microctonus hyperodae Loan, but initial studies have shown that levels of parasitism are lower than in the natural host L. bonariensis (Kuschel). A novel bacterial indicator test was used to determine whether the lower level of parasitism was due to behavioural factors, lack of oviposition, or host resistance. The incidence of ovipositor penetration by the parasitoid M. hyperodae into adult L. maculicollis was measured by immersing the ovipositor of the parasitoid in the facultative pathogen, Serratia marcescens Bizio. Adult weevils were then exposed to parasitoids for up to 72 h and rapid mortality used as an indicator of oviposition penetration. Survival was assessed after six days and surviving weevils were dissected and examined for parasitoid larvae. Mortality among L. maculicolis exposed to parasitoids treated with S. marcescens was significantly higher (P<0.001) than the controls but significantly lower (P<0.001) than in the natural host, L. bonariensis. Dissection of weevils exposed to uncontaminated parasitoids revealed that parasitism in L. maculicolis was significantly (P<0.001) less than parasitism in L. bonariensis. Serratia marcescens-induced mortality plus parasitism of surviving weevils in the parasitoid plus bacteria treatments produced a similar overall effect. Application of bacteria to the parasitoid ovipositor provided a rapid, simple test for ovipositor penetration, which shows potential for separation of behavioural and physiological defence mechanisms in parasitoid/host range studies.     

Influence of host diet on parasitoid fitness: unravelling the complexity of a temperate pastoral agroecosystem

Parasitoid fitness is closely associated with the condition of the host insect, and the condition of the host is partly dependent on the quality and quantity of plant food available. The relationship between host diet and parasitoid fitness therefore has the potential to indirectly mediate complex multitrophic interactions. This relationship may be influenced both by the presence of mycotoxins from endophytic fungi, which share symbiotic relationships with plants and cause reductions in host-plant quality for herbivores, and by the availability of beneficial nutrient-rich plant foods such as pollen. This study used a multitrophic system involving ryegrass, a fungal endophyte, pollen, and an insect herbivore and its parasitoid to study the effect of host diet on parasitoid fitness. Previous studies showed that the larval stage of the parasitoid was negatively influenced by the presence of fungal-derived alkaloids in the diet of its host, but no assessment of effects on the adult stage of the parasitoid had been made. Similarly, the effect of pollen in the diet of the host on the fitness of the parasitoid had not previously been examined. In this study, the fitness of parasitoids reared from hosts fed endophyte-free ryegrass, endophyte-free ryegrass plus pollen, or endophyte-infected ryegrass was assessed in two laboratory experiments. Host insects exhibited significant responses to treatment, but remarkably, there were no significant treatment effects on parasitoid development times, the number of successful parasitoid emergences, tibia length, or preoviposition egg complement. The absence of a strong relationship between host diet and parasitoid fitness in this study differs markedly from studies of other parasitoid–host systems in that the adult stage of this parasitoid appears remarkably unresponsive to variation in host condition. This work contributes to a better understanding of multitrophic interactions and to the refinement of integrated pest management tactics in a temperate pastoral agroecosystem.     

Behavioural Acceptability of Sitona lepidus (Coleoptera: Curculionidae) to the Parasitoid Microctonus aethiopoides (Hymenoptera: Braconidae) Using the Pathogenic Bacterium Serratia marcescens Bizio

Microctonous aethiopoides Loan has been introduced into New Zealand to control the lucerne pest Sitona discoideus Gyllenhal (Coleoptera: Curculionidae). Sitona lepidus Gyllenhal (Coleoptera: Curculionidae) a pest of clover ( Trifolium spp.), has recently established in New Zealand. Laboratory experiments to test the potential of M. aethiopoides to parasitize S. lepidus has resulted in very low levels of parasitism. To investigate whether there were behavioural or physiological barriers to successful parasitism, two experiments were conducted using the insect pathogenic bacterium. Serratia marcescens Bizio as a marker for parasitoid ovipositor penetration. Firstly, M. aethiopoides 'treated' with S. marcescens were exposed to weevils and rapid weevil mortality was used to indicate ovipositor penetration. Up to 50% mortality of S. lepidus occurred, which was comparable with mortality observed in the permissive host Listronotus bonariensis . Dissection of S. lepidus exposed to parasitoids treated with distilled water showed that ca. 21% contained parasitoid eggs of which 98% were nonviable. In the second experiment, exposure periods of 24, 48 and 72 h to S. marcescens -treated parasitoids produced an increase in S. lepidus mortality of 14, 28 and 38%, respectively. There was 3% successful parasitoid development in weevils exposed for 72 h to parasitoids treated with distilled water. M. aethiopoides has been shown to develop successfully in a wide range of non-target weevil species both in the laboratory and field. Possible reasons for poor survival of M. aethiopoides immature stages in S. lepidus are discussed.     

Virus-like particles in the ovaries of Microctonus aethiopoides Loan (Hymenoptera: Braconidae): comparison of biotypes from Morocco and Europe

Virus-like particles (MaVLP) have been discovered in the ovarial epithelial cells of the solitary, koinobiont, endoparasitoid, Microctonus aethiopoides Loan (Hymenoptera: Braconidae) introduced to New Zealand originally from Morocco to control the lucerne pest Sitona discoideus Gyllenhal (Coleoptera: Curculionidae). MaVLP have been found in all females examined. It has been suggested, although not demonstrated, that like many other such VLP found in parasitoids, MaVLP might play a role in host immunosuppression. Since another biotype of M. aethiopoides from Ireland has been proposed for introduction to control the white clover pest, Sitona lepidus Gyllenhal, in New Zealand, it was considered that females from this biotype warranted transmission electron microscope examination for VLP. No VLP were observed in ovarian tissues of specimens collected from three different locations in Ireland. Similarly, none were found in M. aethiopoides sourced from France, Wales, and Norway. These observations are discussed in relation to quarantine host specificity tests with the Irish biotype, which found that the host range of the Irish biotype is likely to be less extensive than that of the Moroccan biotype already in New Zealand.     

Phenology of native weevils (Coleoptera: Curculionidae) in New Zealand pastures and parasitism by the introduced braconid,Microctonus aethiopoides Loan (Hymenoptera: Braconidae)

Abstract

The phenology of native brachycerine weevil species at seven pasture sites in Otago, Canterbury and Waikato was studied by regular quantitative sampling of adults. Weevils were identified to species, and dissected to record reproductive status and parasitism by introduced braconid parasitoids in the genus Microctonus. Climatic data assisted in the interpretation of some population density patterns. Weevil population density was estimated for periods of two to five years at the selected sites. Species in the Entimini (species of Irenimus and Nicaeana) were generally univoltine, with adults emerging in winter‐spring. The main period of reproductive activity was spring, and parasitism by Microctonus aethiopoides reached its highest incidence in January. Low level parasitism of native weevil species by M. aethiopoides was detected at all sites, and by M. hyperodae at two sites. At one site in Otago, parasitism by M. aethiopoides was higher and could have affected the population density of Irenimus aemulator (Broun) and Nicaeana sp. Most parasitism occurred after the main reproductive period of weevils in spring, but a putative second generation in some species might be more affected by parasitoid attack. A native rhytirhinine species, Steriphus variabilis, differed from the entimines because adults emerged in autumn and spring, and may be bivoltine. Mechanisms of M. aethiopoides parasitism of non‐target species in the field are discussed.     

Influence of Clavicipitaceous Endophyte Infection in Ryegrass on Development of the ParasitoidMicroctonus hyperodaeLoan (Hymenoptera: Braconidae) inListronotus bonariensis(Kuschel) (Coleoptera: Curculionidae)

Laboratory experiments were conducted to examine the effect of ryegrass infection by the endophytic fungusAcremonium loliiLatch, Christensen and Samuels onMicroctonus hyperodaeLoan, a parasitoid ofListronotus bonariensis(Kuschel). Progression of parasitoids through the larval instar stages was shown to depend on adequate nutrition of the weevil host. Compared to confinement on endophyte-free ryegrass, parasitized weevils held on nonpreferred diets comprising leaf segments from endophyte-infected ryegrass and switchgrass contained parasitoid larvae with retarded development. Similarly, development of parasitoid larvae was retarded in hosts feeding on artificial diet containing diterpenes and alkaloids ofA. loliiorigin. Several diterpenes incorporated into the diet reduced survival of the parasitoid larvae. Attack rate of parasitoids was reduced when the quality of potential host weevils was compromised by confinement on nonpreferredA. lolii-infected ryegrass or without food for 14 days.     

South American Origins of Microctonus hyperodae Loan (Hymenoptera: Braconidae) Established in New Zealand as Defined by Morphometric Analysis

Eight South American geographic populations of the thelytokous parasitoid Microctonus hyperodae Loan (Hymenoptera: Braconidae) were released in New Zealand in 1991 to assist in the suppression of the pasture pest Listronotus bonariensis (Kuschel) (Coleoptera: Curculionidae). With one exception, parasitoids from each South American geographic population were released in equal numbers at each New Zealand release site. It was postulated that the South American geographic population(s) best suited to the conditions encountered at each New Zealand release locality would eventually become prevalent there. A morphometric analysis of adult parasitoids of known South American origins, reported previously, allowed M. hyperodae derived from west of the Andes (i.e. two collection sites in Chile) to be distinguished from parasitoids derived from east of the Andes (i.e. three collection sites in Argentina and one each in Brazil and Uruguay). Parasitoids derived from a fourth site in Argentina (S. C. de Bariloche) could not be clearly discriminated from either the 'east of the Andes' or 'west of the Andes' categories. A morphometric analysis of M. hyperodae adults collected from five of the New Zealand release sites from 1992-1994 is presented in this contribution. The analysis indicated that parasitoids derived from east of the Andes were significantly more prevalent than expected. The possible reasons for the initial success in New Zealand of one or more east of the Andes populations include the greater fecundity of M. hyperodae collected in Uruguay and the likelihood that M. hyperodae from east of the Andes co-evolved more recently with the stock from which New Zealand's L. bonariensis was founded.     

Evidence of Host Discrimination by Microctonus hyperodae Loan (Hymenoptera: Braconidae, Euphorinae), a Parasitoid of the Argentine Stem Weevil Listronotus bonariensis (Kuschel) (Coleoptera: Curculionidae)

Microctonus hyperodae is a solitary endoparasitoid of the Argentine stem weevil, Listronotus bonariensis. Early investigation into the biology of the parasitoid indicated that there was no discrimination between parasitized and unparasitized hosts. However, dissection data from two experiments were analyzed according to three mathematical models. Model I was based on the Poisson distribution and assumed random selection of hosts. The host discrimination model (model II) assumed that parasitized hosts had reduced attractiveness to searching parasitoids. A competition model (model III) made the assumption that competition between the early immature parasitoid stages led to premature mortality, which was not accounted for in the dissection results. The dissection data indicated statistically significant departure from the Poisson model. Results from the laboratory experiment indicated that host discrimination explained the results more accurately than parasitoid larval competition. Models II and III both provided adequate fits to the field data, although the departure from the observed data was greatest for the competition model. Both intra- and inter-ecotypic host recognition were evident, and there was some indication that clonal recognition took place. This is the first evidence of host discrimination by a member of the genus Microctonus. It has provided further indication of the success of M. hyperodae as a biological control agent.     

Response of parasitoid egg load to host dynamics and implications for egg load evolution

A theoretical debate about whether parasitoids should be time or egg limited now recognizes both as feasible, and interest has turned to determining the circumstances under which each might arise in the field, and their implications for parasitoid behaviour and evolution. Egg loads of parasitoids sampled from the field are predicted to show a negative response to host availability, but empirical support for this relationship is scarce. We measured how a parasitoid's egg load responded to seasonal fluctuations in host population density and recorded the predicted correlation. In early summer, parasitoids were at high risk of time limitation due to low host availability, and in late summer, their offspring were at greater risk of egg limitation due to high host availability. Despite clear seasonal changes in selection pressures on egg load and lifespan, the parasitoid showed no evidence of seasonal variation in its reproductive strategy. We made minor modifications to a previously published model to explore the effects of seasonal variation in host availability on optimal investments in eggs and lifespan and obtained several new results. In particular, under circumstances analogous to some of those observed in our field study, temporal stochasticity in reproductive opportunities can cause investments in eggs to increase, rather than decrease as previously predicted. Our model results helped to explain the parasitoid's lack of a seasonally varying reproductive strategy. Understanding the evolution of parasitoid egg load would benefit from a shift of research emphasis from purely stochastic variation in parasitoid reproductive opportunities to greater consideration of host dynamics.     

Geographic Variation in Egg Load of Microctonus hyperodae Loan (Hymenoptera: Braconidae) and its Implications for Biological Control Success

Eight South American geographic populations of the parthenogenic, proovigenic, koinobiont parasitoid Microctonus hyperodae Loan (Hymenoptera: Braconidae: Euphorinae) were introduced to New Zealand to assist management of the pasture pest Listronotus bonariensis (Kuschel) (Coleoptera: Curculionidae). Geographic variation in fecundity has been suggested as a reason for the populations' differential successes in establishing in New Zealand. This study investigated whether geographic variation in fecundity was due to corresponding variation in pre-oviposition egg load (other possible sources of fecundity variation include searching efficiency, egg survival and female longevity). Variation in egg load accounted for that in fecundity, but also showed that the variation in fecundity was not as great as it had first appeared. Geographic variation in egg load did not explain the pattern of population establishment observed in New Zealand. Egg load was proportional to parasitoid size and this relationship was stronger in populations originating from west, rather than east, of the Andes. A new method for making parasitoid eggs visible is described.