Distribution of resting spores of the Lymantria dispar pathogen Entomophaga maimaiga in soil and on bark

Cadavers of late instar Lymantria dispar (gypsy moth) larvae killed by the fungal pathogen Entomophaga maimaiga predominantly contain resting spores (azygospores). These cadavers frequently remain attached to tree trunks for several weeks before they detach and fall to the ground. Density gradient centrifugation was used to quantify resting spores in the soil and on tree bark. Titers of resting spores were extremely high at 0–10 cm from the base of the tree and the number decreased with distance from the trunk of the tree. Titers were also highest in the organic layer of the soil with numbers decreasing precipitously with increasing depth in the soil. While resting spores were obtained from tree bark, densities per unit area were much lower than those found in the organic soil layer at the base of the tree. Field bioassays were conducted with caged L. dispar larvae to compare infection levels with distance from the tree trunk as well as on the trunk. Highest infection levels were found at 50cm from the tree base with lowest infection on the tree trunk at 0.5 m height, although we expected the highest infection levels among larvae caged at the bases of trees, where highest spore titers occurred. Laboratory experiments demonstrated that L. dispar larvae exposed to resting spore- bearing soil at the soil surface became infected while larvae exposed to soil with resting spores buried at least 1 cm below the surface did not become infected.     

Modeling the Influence of Rainfall and Temperature on the Phenology of Infection of Gypsy Moth, Lymantria dispar, Larvae by the Fungus Entomophaga maimaiga

A computer model driven by daily maximum-minimum temperature and rainfall records was developed to investigate the influence of weather on times when gypsy moth larvae, Lymantria dispar, can become infected by the fungal pathogen Entomophaga maimaiga. In the model, gypsy moth eggs are hatched and neonates are exposed primarily to germinating resting spores in the soil during the spring. Risk of infection is related to forest floor moisture. Larval and fungal development follows a degree-day model. When larvae become 4th instars, they can again become infected by resting spores because they hide in the litter during daylight hours. If rain falls when infected caterpillars die, the fungus sporulates, producing conidia. The number of conidia produced is assumed to directly influence the probability of infection of other larvae. The model was run using weather records from 1990 through 1992. Predicted times of infection were compatible with estimated fungal recruitment rates and changes in field disease prevalence rates. Assumptions about infection mechanisms in the model are discussed as they relate to the real world.     

First record of Entomophaga maimaiga (Entomophthorales: Entomophthoraceae) in Slovakia

The entomopathogenic fungus Entomophaga maimaiga was found for the first time in Slovakia in 2013. Late instar larvae of gypsy moth, Lymantria dispar, from two sites with different population densities were dissected to evaluate the presence of pathogens. The presence of conidia and resting spores of E. maimaiga in gypsy moth cadavers was confirmed from both sites.     

Age-dependent cyclic locomotor activity in the cricket, Gryllus bimaculatus, and the effect of adipokinetic hormone on locomotion and excitability

Excitability and locomotor activity of male and female last instar larvae and adults of the two-spotted cricket are measured under crowded conditions, allowing the animals to interact with conspecifics during observations. Male and female last instar larvae display age-dependent cyclic patterns of activity with maxima during early to mid scotophase and minima during early photophase. A period of low locomotor activity without time of day-dependent cyclic changes starts 1 day before the final moult and lasts until 1 day after the moult. Then, both excitability and locomotor activity increase and become cyclic again within 2 or 3 days. The cyclic changes gradually dampen in adult females older than 6 days and finally cease. When injected into photophase larvae and adults, adipokinetic hormone (AKH) increases excitability and locomotor activity in a dose-dependent manner, whereas it has no such effect when injected into scotophase animals. Other behaviours (jumping, hind wing trembling) that mostly occur in scotophase crickets are also increased by injecting AKH into photophase crickets. We argue that AKH could be responsible for linking the endogenous clock output with the cyclic changes in locomotor activity. Furthermore, AKH may serve to synchronise metabolism and behaviour to optimise larval development and reproduction.     

Diurnal pattern of death and sporulation in Entomophaga maimaiga-infected Lymantria dispar

Entomophaga maimaiga Humber, Shimazu, et Soper (Zygomycotina: Entomophthoraceae) is a naturally occurring obligate fungal pathogen specific to gypsy moth, Lymantria dispar (L.) (Lepidoptera: Lymantriidae) larvae. This fungus is considered the most important natural enemy of this pest insect in North America and Asia. A critically important step for the development of E. maimaiga epizootics is the transmission of propagules to healthy larvae, a process known to require high humidity. Some pathogens are known to manipulate the time of day that hosts die so that propagules are produced to maximize chances of survival and thus enhance transmission. The objective of this study was to assess whether E. maimaiga manipulates L. dispar to die at a certain time of day. Laboratory bioassays were conducted at 15 and 20 °C to record the 24‐h activity pattern of death and sporulation exhibited under an L14:D10 photoperiod and 100% r.h. by four isolates of E. maimaiga in its host L. dispar. Events were recorded every 4 h. Our results clearly demonstrate that E. maimaiga‐infected L. dispar larvae die mainly in the afternoon and that the fungus sporulates during the night. The rhythm was independent of the fungal isolate tested and type of spores produced after larval death. By raising the temperature from 15 to 20 °C, the peak death time narrowed and sporulation was initiated earlier at night.     

Pheromone-binding protein and general odorant-binding protein of Sesamia nonagrioides: sex- and diel-dependent expression

The expression of a pheromone‐binding protein (PBP) and a general odorant‐binding protein (GOBP) in Sesamia nonagrioides (Lef.) (Lepidoptera: Noctuidae) was studied. Lymantria dispar's PBP1 antibody yielded an immunoreactive band with an apparent MW of approximately 14.8 kDa, present specifically in the antennae of both sexes, with males having approximately three‐fold the quantity found in females. Furthermore, Manduca sexta's odorant‐binding protein‐2 (GOBP2) antibody recognized a band at approximately 14.5 kDa in the antennae of both sexes. Levels of both proteins were compared between scotophase and photophase periods in insects that were raised under L16:D8 or under constant light. The level of GOPB2 was significantly lower in both sexes during photophase and continuous light; whereas the level of the PBP was significantly lower in females’ antennae, in males’ antennae it remained at the same level as that found during the scotophase.     

Leaf age effects of elevated CO2-grown white oak leaves on spring-feeding lepidopterans

Folivorous insect responses to elevated CO₂-grown tree species may be complicated by phytochemical changes as leaves age. For example, young expanding leaves in tree species may be less affected by enriched CO₂-alterations in leaf phytochemistry than older mature leaves due to shorter exposure times to elevated CO₂ atmospheres. This, in turn, could result in different effects on early vs. late instar larvae of herbivorous insects. To address this, seedlings of white oak (Quercus alba L.), grown in open-top chambers under ambient and elevated CO₂, were fed to two important early spring feeding herbivores; gypsy moth (Lymantria dispar L.), and forest tent caterpillar (Malacosoma disstria Hübner). Young, expanding leaves were presented to early instar larvae, and older fully expanded or mature leaves to late instar larvae. Young leaves had significantly lower leaf nitrogen content and significantly higher total nonstructural carbohydrate:nitrogen ratio as plant CO₂ concentration rose, while nonstructural carbohydrates and total carbon-based phenolics were unaffected by plant CO₂ treatment. These phytochemical changes contributed to a significant reduction in the growth rate of early instar gypsy moth larvae, while growth rates of forest tent caterpillar were unaffected. The differences in insect responses were attributed to an increase in the nitrogen utilization efficiency (NUE) of early instar forest tent caterpillar larvae feeding on elevated CO₂-grown leaves, while early instar gypsy moth larval NUE remained unchanged among the treatments. Later instar larvae of both insect species experienced larger reductions in foliage quality on elevated CO₂-grown leaves than earlier instars, as the carbohydrate:nitrogen ratio of leaves substantially increased. Despite this, neither insect species exhibited changes in growth or consumption rates between CO₂ treatments in the later instar. An increase in NUE was apparently responsible for offsetting reduced foliar nitrogen for the late instar larvae of both species.     

Baculovirus‐induced tree‐top disease: how extended is the role of egt as a gene for the extended phenotype?

Many parasites alter host behaviour to enhance their chance of transmission. Recently, the ecdysteroid UDP‐glucosyl transferase (egt) gene from the baculovirus Lymantria dispar multiple nucleopolyhedrovirus (LdMNPV) was identified to induce tree‐top disease in L. dispar larvae. Infected gypsy moth larvae died at elevated positions (hence the term tree‐top disease), which is thought to promote dissemination of the virus to lower foliage. It is, however, unknown whether egt has a conserved role among baculoviruses in inducing tree‐top disease. Here, we studied tree‐top disease induced by the baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV) in two different host insects, Trichoplusia ni and Spodoptera exigua, and we investigated the role of the viral egt gene therein. AcMNPV induced tree‐top disease in both T. ni and S. exigua larvae, although in S. exigua a moulting‐dependent effect was seen. Those S. exigua larvae undergoing a larval moult during the infection process died at elevated positions, while larvae that did not moult after infection died at low positions. For both T. ni and S. exigua, infection with a mutant AcMNPV lacking egt did not change the position where the larvae died. We conclude that egt has no highly conserved role in inducing tree‐top disease in lepidopteran larvae. The conclusion that egt is a ‘gene for an extended phenotype’ is therefore not generally applicable for all baculovirus–host interactions. We hypothesize that in some baculovirus–host systems (including LdMNPV in L. dispar), an effect of egt on tree‐top disease can be observed through indirect effects of egt on moulting‐related climbing behaviour.     

Diel rhythms of adipokinetic hormone, fat body response, and haemolymph lipid and sugar levels in the house cricket

Abstract. The adipokinetic hormone (Grb-AKH) content in the corpora car-diaca of the house cricket, Acheta domesticus , varied during a day with two peaks in the scotophase and one peak in the photophase. There were two distinct peaks of total lipid concentration in the blood, one early in the photophase and the other early in the scotophase. Fat body sensitivity to adipokinetic hormone also varied in close synchrony with the lipid rhythm. It was not possible to attribute the rhythm of blood lipid titre unequivocally to either the rhythm of adipokinetic hormone content in the corpora cardiaca or to the rhythm of sensitivity of the fat body to the hormone. In adult crickets the blood carbohydrate titre had two peaks in adult females, one towards the end of scotophase and another in the late photophase (c. 12h apart), but a single peak at the end of scotophase was apparent in last instar larvae. The blood carbohydrate rhythm persists in DD and is therefore endogenous. Carbohydrates were not mobilized by the Grb-AKH, therefore could not be involved in the blood carbohydrate rhythm. Exposure to various day lengths caused shifts in the patterns of the carbohydrate rhythms, but imposition of a cyclic temperature regime had no effect on the rhythm.     

Pheromone‐trapping the nun moth,Lymantria monacha (Lepidoptera: Lymantriidae) in Inner Mongolia,China

The nun moth, Lymantria monacha L., is one of the most important defoliators of Eurasian coniferous forests. Outbreaks during 2011–2015 in the natural/planted larch, and larch‐birch mixed forests of the Greater Khingan Range in Inner Mongolia, China, caused tremendous timber losses from severe defoliation and tree mortality. A series of trapping experiments were conducted in these outbreak areas to evaluate the efficacy of a synthetic species‐specific pheromone lure based on the female pheromone blend of European nun moth populations. Our results clearly show that the nun moth in Inner Mongolia is highly and specifically attracted to this synthetic pheromone, with few gypsy moths (Lymantria dispar) captured. Flight activity monitoring of L. monacha male moths using pheromone‐baited Unitraps at 2 locations during the summer of 2015 indicated that the flight period started in mid‐July, peaking in early August at both locations. Based on male moth captures, there was a strong diurnal rhythm of flight activity throughout the entire scotophase, peaking between 22:00 and 24:00. Unitraps and wing traps had significantly and surprisingly higher catches than the gypsy moth traps. Unitraps fastened to tree trunks 2 m above ground caught significantly more male moths than those at the ground level or at 5 m height. Male L. monacha moths can be attracted to pheromone‐baited traps in open areas 150–200 m distant from the infested forest edge. Our data should allow improvement on the performance of pheromone‐baited traps for monitoring or mass‐trapping to combat outbreaks of this pest in northeastern China.     

Diel variations in plasma glucose concentrations of Malachite Sunbirds Nectarinia famosa

Plasma glucose concentrations in birds are much higher than those in mammals of similar body mass, and they are thought to be kept more or less constant. We investigated plasma glucose concentrations of Malachite Sunbirds (Nectarinia famosa) to determine whether there was a circadian rhythm in plasma glucose and whether plasma glucose concentrations rose at lower temperatures. We explored the possibility that glycogenolysis and gluconeogenesis might be used by birds before daylight to generate body heat during arousal from torpor or from overnight adaptive heterothermy. Plasma glucose concentration of Malachite Sunbirds were relatively high, between 13.6 and 21.4 mmol/L. Plasma glucose concentrations were higher at 5°C than at 25°C, and generally lower during the scotophase, particularly in the early hours of the morning. Therefore, it appears that Malachite Sunbirds are not increasing plasma glucose concentrations during arousal from torpor or to increase scotophase body temperature levels to photophase levels. It is apparent that these nectarivorous birds have a circadian variation in plasma glucose concentrations that is affected by ambient temperature and feeding patterns.     

Foraging in a pathogen reservoir can lead to local host population extinction: a case study of a Lepidoptera-virus interaction

In 1990, natural infestations of the polyphagous vapourer moth, Orgyia antiqua (Lepidoptera: Lymantriidae) in lodgepole pine plantations in northern Scotland, were studied to ascertain the role of host foraging behaviour on the prevalence of nucleopolyhedrovirus (NPV; Baculoviridae) infection in the population. Aerial dispersal of early instar larvae (L1–L3) from the tree canopy onto heather foliage at the forest understorey, with subsequent relocation back onto the tree as late-instar larvae (L4–L6) appeared to play a significant role in the development of a widespread virus epizootic in which approximately 80% of L4–L6 individuals succumbed to disease. Bioassays of foliage 1 year later showed that the distribution of NPV followed a pronounced vertical gradient through the forest canopy culminating in high concentrations of virus in the forest understorey. Experimental systems comprising potted pine trees positioned above heather bases showed that NPV infections could be acquired by early stage larvae following dispersal from the tree and feeding on the undercanopy vegetation, then translocated to the tree component for secondary transmission to susceptible tree-feeding individuals. Behavioural studies indicated that the tendency for first-, second- and third-instar larvae to disperse to the understorey was probably not influenced by larval density on the tree but was strongly dependent on larval instar. In contrast, the tendency for larvae to relocate from the understorey heather to the tree was affected by both larval density and larval instar, suggesting that both these factors may significantly affect virus acquisition, translocation and transmission in the host population. In the present study, the heather understorey appeared to act as a pathogen reservoir in which virus could persist between host generations. Spatial heterogeneity in virus distribution combined with host foraging behaviour (dispersal and feeding) resulted in the pathogen playing a major role in host population dynamics over an extended time period (3 years). The reservoir theory is supported by the observation that similar dynamics were not observed in O. antiqua populations at neighbouring sites which lacked understorey food plants. Received: 8 June 1998 / Accepted: 5 October 1998     

Thermal orientation of Anthonomus pomorum (Coleoptera: Curculionidae) in early spring

Abstract. The selection of habitats with favourable temperature by the apple blossom weevil Anthonomus pomorum (L.) is investigated in a temperature gradient arena with a range of approximately 0–15 °C. Single female and male weevils are tested in the arena 2, 4 and 6 days after termination of diapause, during photophase and during scotophase. During photophase, weevils of both sexes choose the warmest part of the temperature gradient arena, irrespective of the time elapsed after diapause. During scotophase, high temperature is favoured by male weevils, as well as by females 2 and 4 days after diapause. However, 6 days after termination of diapause, females show no thermal preference in the temperature gradient arena during scotophase, indicating that thermal choice of female A. pomorum in the scotophase changes with time after the termination of diapause. The results suggest that both sexes benefit from thermoregulation by habitat choice during photophase when the weevils are flight active and colonize apple trees.     

Uscana lariophaga, West-African egg parasitoid of the cowpea bruchid beetleCallosobruchus maculatus: photoperiod, parasitization and eclosion interactions

The diurnal pattern of parasitization and eclosion of the trichogrammatidUscana lariophaga Steffan, egg parasitoid of the cowpea bruchidCallosobruchus maculatus (Fabricius) was studied for L12:D12 photoperiods. The wasp parasitized eggs throughout each 24-h period with about 70% of the parasitization taking place during the first 12 h regardless whether it was the photophase or the scotophase. More female progeny was produced when the first 12-h period was photophase instead of scotophase. Eclosion of wasps took place over a period of 4 days and occurred during the second half of the scotophase and the first half of the photophase. The number of wasps eclosed during the photophase was similar to that eclosed during the scotophase. Average development time was 8.9 days at 30°C. Male development was completed 6 to 8 h before the females, hence a higher percentage of females emerged in the later eclosion peaks. During scotophases more females eclosed than during photophases. Although the results indicate that the wasp is able to perform under dark storage conditions the effect of permanent low light intensity remains to be studied.     

Spruce budworm growth,development and food utilization on young and old balsam fir trees

Laboratory rearing of spruce budworm, Choristoneura fumiferana, in conjunction with field rearing, gravimetric analyses, a transfer experiment, and foliage chemical analyses at six dates during the period of budworm feeding activity indicated that the age of balsam fir, Abies balsamea, trees (70-year-old mature trees or 30-year-old juvenile trees) affected tree suitability for the spruce budworm via the chemical profile of the foliage. Insects reared on old trees had greater survival and pupal weight, shorter development times, and caused more defoliation than those reared on young trees. Young trees were more suitable for the development of young larvae (instars 2–5), while old trees were more suitable for the development of older, sixth-instar larvae. These results were confirmed by the laboratory transfer experiment. Young larvae fed foliage from young trees had higher relative growth rates (RGR), digestibility (AD), and efficiency of conversion of ingested foliage (ECI) than those fed foliage from old trees. These differences appeared to be related to the high N:tannins ratio, and the high contents of P present in young trees during the development of the young larvae. Old larvae fed foliage from old trees had higher relative growth rates, relative consumption rates (RCR), and digestibility of the foliage than those fed foliage from young trees. The high digestibility of the foliage of old trees was compensated for by a lower efficiency of conversion of digested food (ECD), which in turn resulted in no significant effect of tree age on the efficiency of conversion of ingested foliage by old larvae. The low relative consumption rate of old larvae fed foliage from young trees appeared to be related to the low N:tannins ratio, and the high contents of bornyl acetate, terpinolene, and °-3-carene present in young trees during the budworm sixth instar. Variations in these compounds in relation to tree age may serve as mechanisms of balsam fir resistance to spruce budworm by reducing the feeding rate of sixth instar larvae.     

Effects of heavy metal stress on free amino acids in the haemolymph and proteins in haemolymph and total body tissue of Lymantria dispar larvae parasitized by Glyptapanteles liparidis

Heavy metal contamination of the forest pest insect Lymantria dispar (L.) (Lepidoptera; Lymantriidae), the gypsy moth, can alter its haemolymph composition, as has already been shown for carbohydrates and lipids in recent studies. L. dispar larvae are frequently parasitized by Glyptapanteles liparidis (Bouché) (Hymenoptera; Braconidae) larvae, which can—to some extent—regulate the population size of the pest insect. The parasitoids feed on the haemolymph of L. dispar larvae; hence, a different haemolymph composition of the host alters the trophic situation of the parasitoids. The aim of the present study was to investigate whether metal contamination also affects the concentrations of free amino acids in L. dispar haemolymph, and protein concentrations in their haemolymph and tissue. L. dispar larvae were parasitized on the first day of the second instar and then reared on diets contaminated with Cd, Pb, Cu or Zn at two concentrations each. Haemolymph and total body tissue of the larvae (fourth instar/third day) were analyzed. The concentrations of the free amino acids were elevated in five out of the eight contamination groups (Cd6, Pb4, Cu6, Cu10, Zn60), whereas haemolymph protein concentrations were significantly reduced in all contaminated individuals. The haemolymph protein concentration was 18 mg/ml in the control group and decreased to less than 10 mg/ml due to cadmium and zinc contamination at both concentrations and in the low copper contamination group. In contrast, total body proteins (136 g/mg dry weight in the control group) were elevated due to heavy metal stress. Analyses of haemolymph protein concentrations during the fourth instar demonstrated an increase of the proteins from day one to day four (followed by a decrease on the fifth day) in the control group and the cadmium contamination group. A steady increase of proteins from the first to the fifth day in the copper and zinc contaminated larvae indicated a retarded development in these groups. Thus, the present study along with other recent studies demonstrated, that heavy metal stress changes the concentrations of all main haemolymph compounds of L. dispar larvae.     

Photoperiodismus bei Aphiden. Gegensätzliche Reaktionsmechanismen bei Pterocomma jacksoni Theob. und Megoura viciae Bckt. (Homoptera,Rhynchota)

Summary Viviparous females of Pterocomma and Megoura were chilled daily for 4 hours at +1° C throughout larval stages. Chilling Pterocomma during photophase turns long day into short day, whereas chilling during scotophase has no effect. On the other hand chilling Megoura during photophase has no effect, whereas chilling during scotophase turns short day into long day. When the neurosecretory cells of the pars intercerebralis were cauterized, females of Pterocomma became permanent oviparaproducer, females of Megoura permanent viviparaproducer.     

Competition between the gypsy moth, Lymantria dispar, and the northern tiger swallowtail, Papilio canadensis: interactions mediated by host plant chemistry, pathogens, and parasitoids

The gypsy moth, Lymantria dispar, and the northern tiger swallowtail, Papilio canadensis, overlap geographically as well as in their host ranges. Adult female swallowtails are incapable of distinguishing between damaged and undamaged leaves, and the opportunities for competition between these two species are numerous. We designed field and laboratory experiments to look for evidence of indirect competition between P. canadensis and L. dispar larvae. Swallowtail caterpillars were reared in the laboratory on leaves from gypsy-moth-defoliated and undefoliated trees to explore host-plant effects. We tested for pathogen-mediated interactions by rearing swallowtail larvae on both sterilized and unsterilized leaves from defoliated and undefoliated sources. In addition, we measured the effects of known gypsy moth pathogens, as well as gypsy moth body fluids, on the growth and survival of swallowtail larvae. Field experiments were designed to detect the presence of parasitoid-mediated competition, as well: we recorded parasitism of swallowtail caterpillars placed in the field either where there were no gypsy moth larvae present, or where we had artificially created dense gypsy moth populations. We found evidence that swallowtails were negatively affected by gypsy moths in several ways: defoliation by gypsy moths depressed swallowtail growth rate and survival, whether leaves were sterilized or not; sterilization significantly reduced the effect of defoliation, and gypsy moth body fluids proved lethal; and swallowtail caterpillars suffered significantly increased rates of parasitism when they were placed in the field near gypsy moth infestations.