Synthesis of fluorescent Molecularly Imprinted Polymer Nanoparticles Sensing Small Neurotransmitters with High Selectivity Using Immobilized Templates with Regulated Surface Density.

To develop nanosensors to probe neurotransmitters, we synthesized fluorescent-functionalized molecularly imprinted polymeric nanoparticles (fMIP-NPs) using monoamine neurotransmitters (serotonin and dopamine) immobilized on glass beads as templates. The size and fluorescence intensity of the fMIP-NPs synthesized with blended silane couplers increased with the presence of the target but were insensitive to the target analogs (L-tryptophan and L-dopa, respectively). However, when the template is anchored by a pure silane agent, both the fluorescence intensity and particle size of the fMIP-NPs were sensitive to the structural analog of the template. Another fMIP-NP was synthesized in the presence of poly([2-(methacryloyloxy)ethyl] trimethylammonium chloride (METMAC)-co-methacrylamide) grafted onto glass beads as a dummy template for acetylcholine. Acetylcholine increased the diameter and fluorescence intensity of the fMIP-NP, but choline had no effect. When the homopolymer of METMAC was used as a template, the fluorescence intensity and size of the resulting nanoparticles were not responsive to either acetylcholine or choline. The principle of increased fluorescence intensity due to specific interaction with the target substance is probably due to the increased distance between the fluorescent functional groups and decreased self-quenching due to the swelling caused by the specific interaction with the template. The results also indicate that MIP nanoparticles prepared by solid-phase synthesis can be used for targeting small molecules, such as the neurotransmitters addressed in this study, by adjusting the surface density of the template.

High Variability in Pre-Oviposition Time Independent of Diet Available at Eclosion: A key Reproductive Trait in the Ladybird Beetle Harmonia axyridis (Coleoptera: Coccinellidae) in Its Native Range.

While insect predators need high-quality food to reach sexual maturity and reproduce, starvation following adult eclosion may occur when prey are missing in agroecosystems. This study explores whether the type of diet available at eclosion determines the future fecundity of newly emerged adult predators. In a laboratory experiment, three different diets (i.e., flowers of Perilla frutescens (L.) Britton, eggs of Ephestia kuehniella Zeller as prey, or no food) were offered to adult females of the ladybird beetle Harmonia axyridis Pallas during their first three days after adult eclosion. On the fourth day, each female was paired with a prey-fed male and the pair was subsequently fed with prey. Diet at eclosion did not affect pre-oviposition time, the number of eggs oviposited daily, or the viability of egg batches. High variability in pre-oviposition time among females was observed for all diets. Significant negative linear relationships were found between pre-oviposition time and both the number of eggs oviposited daily and the viability of egg batches. This study clarifies that the food readily available at adult eclosion does not affect the capacity of H. axyridis to reproduce, provided that adults find prey within a few days. More generally, it shows that the reproductive traits of H. axyridis allow this generalist predator to be highly adapted to heterogenous environments in its native range. It is concluded that the variety of habitats offered by diversified agroecosystems may highly benefit the ladybird beetle H. axyridis, and potentially enhance its ability to biologically regulate crop pests.

Direct and indirect effect of cannibalism and intraguild predation in the two sibling Harmonia ladybird beetles.

In this study, we focused on the direct (i.e., predation) and indirect (i.e., potential threat from coexisting with a larger individual) effects of cannibalism and intraguild predation (IGP) during larval stages of two sibling ladybird beetles. These effects play an important role in the coexistence of the generalist-common Harmonia axyridis and specialist-rare H. yedoensis (Coleoptera: Coccinellidae). Direct predation effect of cannibalism and IGP was asymmetric in the two sibling ladybird beetles; the fourth instar larvae of H. axyridis were better intraguild predators than cannibals, while the reverse was true in the larvae of H. yedoensis. Neither cannibalism nor IGP significantly affected female body weight in either species. Larval H. axyridis surviving exposure to cannibalism or IGP had a reduced number of ovarioles as adults, whereas adult H. yedoensis ovarioles were not affected. For the indirect effects, longer developmental times in males and females and a lower total number of ovarioles in females were detected in H. axyridis. In H. yedoensis, shorter developmental time of males, lighter adult weight and fewer total ovarioles in females were observed. Olfactometer choice experiments clarified that the fourth instar larvae of H. axyridis avoided the first instar conspecific larvae, while those of H. yedoensis were attracted to the odors from H. axyridis and conspecifics. Thus, H. axyridis has an avoidance mechanism only for cannibalism but not for IGP, whereas H. yedoensis does not have any avoidance mechanism. These different behaviors in the direct and indirect effects of cannibalism and IGP observed in the laboratory may play important roles in the coexistence of generalist-common H. axyridis and specialist-rare H. yedoensis in natural conditions, compensating for the large handicap of H. yedoensis at reproductive interference from H. axyridis.

Fitness costs of reflex bleeding in the ladybird beetle Harmonia axyridis: the role of parental effects.

Reflex bleeding is an important antipredator defense mechanism in Coccinellidae. We examined the costs of reflex bleeding in larval and adult stages of the ladybird beetle Harmonia axyridis on offspring fitness and reproductive performance through the comparisons between bled and control larvae, reciprocal crosses of bled/control beetles, and early and late clutch phenotypes. Beetles bled during their larval stage spent a longer time in development and weighed less than controls. Egg fertility was reduced for crosses where either one or both parents had been bled during the larval or adult stage. Offspring crosses that included a parent bled during the larval stage suffered fitness costs in development and female body mass, while those bled during the adult stage suffered no transgenerational costs. Males that suffered bleeding during their larval stage accelerated progeny development of nonbled females in early clutches, suggesting a positive transgenerational effect of larval bleeding, while males that did not suffer bleeding accelerated progeny development of bled females in later clutches. As the underlying effects of bleeding on females' offspring in the early clutches were diminished in the late ones, suggesting another transgenerational effect. The strengths of maternal and paternal effects on progeny development of parents bled at the larval stage were higher in earlier clutches. This study suggests that H. axyridis adults are less affected than larvae by the frequent use of the defensive secretions in their stressful habitats.

Increase in male reproductive success and female reproductive investment in invasive populations of the harlequin ladybird Harmonia axyridis.

Reproductive strategy affects population dynamics and genetic parameters that can, in turn, affect evolutionary processes during the course of biological invasion. Life-history traits associated with reproductive strategy are therefore potentially good candidates for rapid evolutionary shifts during invasions. In a series of mating trials, we examined mixed groups of four males from invasive and native populations of the harlequin ladybird Harmonia axyridis mating freely during 48 hours with one female of either type. We recorded the identity of the first male to copulate and after the 48 h-period, we examined female fecundity and share of paternity, using molecular markers. We found that invasive populations have a different profile of male and female reproductive output. Males from invasive populations are more likely to mate first and gain a higher proportion of offspring with both invasive and native females. Females from invasive populations reproduce sooner, lay more eggs, and have offspring sired by a larger number of fathers than females from native populations. We found no evidence of direct inbreeding avoidance behaviour in both invasive and native females. This study highlights the importance of investigating evolutionary changes in reproductive strategy and associated traits during biological invasions.

Asymmetric reproductive interference between specialist and generalist predatory ladybirds.

1. Closely related species often differ greatly in the quality and breadth of resources exploited, but the actual mechanisms causing these differences are poorly understood. Because in the laboratory specialized species often survive and perform as well or better on host species that are never utilized in nature, negative ecological interactions restricting host range must exist. Here, we focused on reproductive interference, which has been theoretically predicted to drive niche separation between closely related species with overlapping mating signals. 2. We examined the interspecific sexual interactions in relation to ecological specialization and generalization in two sibling ladybird species, Harmonia yedoensis and Harmonia axyridis. Harmonia yedoensis is a specialist predator that preys only on pine aphids, which are highly elusive prey for ladybird hatchlings, whereas H. axyridis is a generalist predator with a broad prey and habitat range. 3. We experimentally showed that conspecific sperm fertilized the vast majority of eggs regardless of mating order (i.e. conspecific sperm precedence) when a female of H. yedoensis or H. axyridis mated with both a conspecific and a heterospecific male. Moreover, we demonstrated that mating opportunities of H. yedoensis females strongly decreased as heterospecific density increased relative to conspecific density. In contrast, in H. axyridis, female mating success was high regardless of conspecific or heterospecific density. 4. Our results suggest that the generalist H. axyridis should be dominant to the specialist H. yedoensis in terms of reproductive interference. Our results support the hypothesis that asymmetric reproductive interference from the dominant species may force the non-dominant species to become a specialist predator that exclusively utilizes less preferred prey in nature.

Tolerance of Drosophila flies to ibotenic acid poisons in mushrooms.

The mushroom genus Amanita has a spectrum of chemical compounds affecting survival and performance of animals. Ibotenic acid is one of such compounds found in some Amanita mushrooms. We studied the effects of ibotenic acid and its derivative, muscimol, on egg-to-pupa survival, pupation time, and pupal size in five Drosophila species (Diptera: Drosophilidae), Drosophila bizonata, Drosophila angularis, Drosophila brachynephros, Drosophila immigrans, and Drosophila melanogaster. The first three species are mycophagous and use a wide range of mushrooms for breeding, whereas D. immigrans and D. melanogaster are frugivorous. We reared fly larvae on artificial medium with 500, 250, 125, and 62.5 microg/ml of ibotenic acid and/or musimol. The three mycophagous species were not susceptible to ibotenic acid, whereas the two frugivorous species were affected. In experiments with D. melanogaster, muscimol was less toxic than ibotenic acid.

Shoot development and extension of Quercus serrata saplings in response to insect damage and nutrient conditions.

BACKGROUND AND AIMS: Plants have the ability to compensate for damage caused by herbivores. This is important to plant growth, because a plant cannot always avoid damage, even if it has developed defence mechanisms against herbivores. In previous work, we elucidated the herbivory-induced compensatory response of Quercus (at both the individual shoot and whole sapling levels) in both low- and high-nutrient conditions throughout one growing season. In this study, we determine how the compensatory growth of Quercus serrata saplings is achieved at different nutrient levels. METHODS: Quercus serrata saplings were grown under controlled conditions. Length, number of leaves and percentage of leaf area lost on all extension units (EUs) were measured. KEY RESULTS: Both the probability of flushing and the length of subsequent EUs significantly increased with an increase in the length of the parent EU. The probability of flushing increased with an increase in leaf damage of the parent EU, but the length of subsequent EUs decreased. This indicates that EU growth is fundamentally regulated at the individual EU level. The probabilities of a second and third flush were significantly higher in plants in high-nutrient soil than those in low-nutrient soil. The subsequent EUs of damaged saplings were also significantly longer at high-nutrient conditions. CONCLUSIONS: An increase in the probability of flushes in response to herbivore damage is important for damaged saplings to produce new EUs; further, shortening the length of EUs helps to effectively reproduce foliage lost by herbivory. The probability of flushing also varied according to soil nutrient levels, suggesting that the compensatory growth of individual EUs in response to local damage levels is affected by the nutrients available to the whole sapling.