Harlequin Ladybird Harmonia Axyridis Research Articles

Responses of avian predators to the polymorphic harlequin ladybird (Harmonia axyridis)

Abstract The harlequin ladybird (Harmonia axyridis) shows a high level of colour polymorphism. Particular forms differ in their colour combination, pattern and abundance. Two species of wild‐caught passerines native to Central Europe were offered various forms of ladybirds, differing in their colour pattern and abundance in nature. We predicted that those forms that are more abundant in the wild are better protected, as they are more familiar to predators. Forms novemdecimsignata and spectabilis, which represent 97% of individuals in the wild population, significantly differ in their visual appearance (mostly orange vs. mostly black). The form axyridis is very rare in the wild, and suturalis and aninkae do not occur in the wild and were derived from laboratory breedings As predators, we used great tits (Parus major), which are very aversive towards ladybirds, and tree sparrows (Passer montanus), which are quite willing to attack and even eat ladybirds. We compared bird responses to particular ladybird colour forms and included a brown‐painted control to test the effect of conspicuous colours. We showed that both species of birds attacked all conspicuous forms of the ladybird equally and usually very rarely. The brown‐painted novemdecimsignata form was attacked more frequently compared with the conspicuous forms, showing that the visual appearance prevents birds from attacking any conspicuous colour combination. Sparrows tended to eat the attacked ladybirds of forms novemdecimsignata, spectabilis and brown painted. The rare axyridis form and laboratory forms were very well protected from the attack, very likely due to neophobia. We may conclude that despite the outstanding polymorphism of H. axyridis, its protection against avian predators is very effective as long as the red‐and‐black pattern is preserved.

How do invasion syndromes evolve? An experimental evolution approach using the ladybird Harmonia axyridis

Experiments comparing native to introduced populations or distinct introduced populations to each other show that phenotypic evolution is common and often involves a suit of interacting phenotypic traits. We define such sets of traits that evolve in concert and contribute to the success of invasive populations as an ‘invasion syndrome’. The invasive Harlequin ladybird Harmonia axyridis displays such an invasion syndrome with, for instance, females from invasive populations being larger and heavier than individuals from native populations, allocating more resources to reproduction, and spreading reproduction over a longer lifespan. Invasion syndromes could emerge due to selection acting jointly and directly on a multitude of traits, or due to selection on one or a few key traits that drive correlated indirect responses in other traits. Here, we investigated the degree to which the H. axyridis invasion syndrome would emerge in response to artificial selection on either female body mass or on age at first reproduction, two traits involved in their invasion syndrome. To further explore the interaction between environmental context and evolutionary change in molding the phenotypic response, we phenotyped the individuals from the selection experiments in two environments, one with abundant food resources and one with limited resources. The two artificial selection experiments show that the number of traits showing a correlated response depends upon the trait undergoing direct selection. Artificial selection on female body mass resulted in few correlated responses and hence poorly reproduced the invasion syndrome. In contrast, artificial selection on age at first reproduction resulted in more widespread phenotypic changes, which nevertheless corresponded only partly to the invasion syndrome. The artificial selection experiments also revealed a large impact of diet on the traits, with effects dependent on the trait considered and the selection regime. Overall, our results indicate that direct selection on multiple traits was likely necessary in the evolution of the H. axyridis invasion syndrome. Furthermore, they show the strength of using artificial selection to identify the traits that are correlated in different selective contexts, which represents a crucial first step in understanding the evolution of complex phenotypic patterns, including invasion syndromes.

Exotic predators are not better biocontrol agents: the harlequin ladybird is not the most voracious in Mexico.

The use of exotic species for pest biocontrol has been a common pathway for introduction and dispersal of invasive species that may have undesired outcomes. Biocontrol agents are believed to be a less damaging alternative than pesticides, but some species may also prey on or parasitize native species or outcompete them for resources. The harlequin ladybird (Harmonia axyridis) is a well-known biocontrol agent originally from Asia that has established invasive populations in 59 countries around the globe. Harlequin ladybirds are generalist predators that in addition to pests prey on an array of different species including other coccinelids’ eggs and larvae. In Mexico, native ladybirds that share ecological requirements with harlequin ladybirds are at risk of being outcompeted and predated upon. The aim of our study was to compare the foraging efficiency of harlequin ladybirds against three species of native coccinelids when preying on aphids. We investigated the foraging behaviour of ladybirds alone and in pairs with a conspecific, a native heterospecific or an exotic heterospecific. We found that the native Cycloneda sanguinea was the species that consumed the most aphids, while Hippodamia convergens was the fastest to find and consume each aphid. Harlequin ladybirds and H. convergens consumed the same number of aphids while P. vittigera consumed less. Conspecific competition was stronger than heterospecific competition. We discuss the suitability of using the exotic invasive harlequin ladybird for aphid biocontrol in comparison with native coccinelids.

Warm Mesoclimate Advances the Seasonal Dynamics of Harmonia axyridis in Urban Habitats

In 2016–2019, seasonal changes in the abundance of the harlequin ladybird Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae) were established in the city center of Prague, Central Europe, and in its outskirts. Adults were sampled from lime trees (Tilia spp.) at regular intervals throughout the growing season. The abundance of H. axyridis paralleled the course of abundance of its prey, the aphid Eucallipterus tiliae L., which peaks either early or late in the season. As a result, the seasonal dynamics of H. axyridis were unimodal, with a peak in the early (late June—early July of 2017 and 2019) or late (late July—mid-September of 2016 and 2018) period of the season. In the early period, there was a small (1–4 days) difference in the timing of the peak of H. axyridis between the city center and the outskirts. In the late period, the peak occurred significantly earlier (by 13–21 days) in the city center due to the warmer climate there than in the outskirts. The difference in the timing of the population peak between both locations disappeared after recalculating the calendar to thermal time (number of day degrees above 10.6°C thresholds elapsed from the end of H. axyridis hibernation). The warm mesoclimate of the city center advances the seasonal dynamics of H. axyridis, contributing to the success of this invasive species in urban habitats.

PREVALENCE AND DISTRIBUTION OF DIFFERENT NATURAL ENEMIES IN THE HARLEQUIN LADYBIRD HARMONIA AXYRIDIS (COLEOPTERA COCCINELLIDAE)

The harlequin ladybird Harmonia axyridis (Coleoptera: Coccinellidae) is one of the most common invasive alien species in the world, which has a global impact on biodiversity and agriculture, and causes economic loss and harm to humans. This insect can be infected by different natural enemies, but their interactions remain understudied. In the present study, 581 adult Ha. axyridis specimens were collected from 21 localities in Hungary with the help of citizen scientists and examined for natural enemies. The parasitic nematode Parasitylenchus bifurcatus (Nematoda: Allantonematidae) is reported in Hungary on the harlequin ladybird for the first time. The ladybirds were also infected by the ectoparasitic fungus Hesperomyces virescens (Ascomycota: Laboulbeniales) and parasitoid Dinocampus coccinellae (Hymenoptera: Braconidae). The nematode parasite was present in all localities where Hesperomyces-infected ladybirds appeared. The prevalence of recorded natural enemies varied by locality: for P. bifurcatus it ranged from 1% to 8%; for He. virescens from 2% to 36%; and for the parasitoid it was 4%. Besides, a statistically significant positive correlation was shown between the nematode and the fungus co-infection using Spearman’s correlation. A study on the multiparasitism of harlequin ladybirds was not yet performed in Hungary. The study area is located between two distant areas in Europe where co-infections had already been observed, indicating that this is probably not a sporadic phenomenon, but that parasitism of the harlequin ladybird by multiple natural enemies might be prevalent throughout the area.

Antennal Transcriptome Analysis and Identification of Candidate Chemosensory Genes of the Harlequin Ladybird Beetle, Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae).

Simple SummaryThe predatory harlequin ladybird Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae) has been widely released for classical and augmentative biological control programs of insect herbivores and is now distributed worldwide. Because of its invasive behavior and the threat it can pose to local biodiversity, this ladybird has been adopted as a model species for invasive biocontrol predators. A huge amount of existing literature is available on this species. However, little is known about the mechanisms underlying H. axyridis smell and taste, even though these senses are important in this ladybird for courtship, mating, and for locating suitable habitats for feeding and oviposition. Here we describe the first chemosensory gene repertoire that is expressed in the antennae of male and female H. axyridis. Our findings would likely represent the basis for future functional studies aiming at increasing the efficacy of H. axyridis in biological control or at reducing its populations in those areas where the ladybird has become a matter of concern due to its invasiveness.In predatory ladybirds (Coleoptera: Coccinellidae), antennae are important for chemosensory reception used during food and mate location, and for finding a suitable oviposition habitat. Based on NextSeq 550 Illumina sequencing, we assembled the antennal transcriptome of mated Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae) males and females and described the first chemosensory gene repertoire expressed in this species. We annotated candidate chemosensory sequences encoding 26 odorant receptors (including the coreceptor, Orco), 17 gustatory receptors, 27 ionotropic receptors, 31 odorant-binding proteins, 12 chemosensory proteins, and 4 sensory neuron membrane proteins. Maximum-likelihood phylogenetic analyses allowed to assign candidate H. axyridis chemosensory genes to previously described groups in each of these families. Differential expression analysis between males and females revealed low variability between sexes, possibly reflecting the known absence of relevant sexual dimorphism in the structure of the antennae and in the distribution and abundance of the sensilla. However, we revealed significant differences in expression of three chemosensory genes, namely two male-biased odorant-binding proteins and one male-biased odorant receptor, suggesting their possible involvement in pheromone detection. Our data pave the way for improving the understanding of the molecular basis of chemosensory reception in Coccinellidae.

Invasion history of Harmonia axyridis (Pallas, 1773) (Coleoptera: Coccinellidae) in Ecuador.

Harmonia axyridis is a ladybird extensively used around the world for biological control of agricultural pests. However, it has become invasive in several countries, producing negative ecological and socio-economic impacts. Herein, we review the invasion history of the Harlequin Ladybird Harmonia axyridis (Pallas, 1773) in Ecuador. Although first reported in Ecuador in 2012, museum specimens date back to 2004 and it is currently established across the country, especially along the Andean region. Due to its invasive nature, further studies are urgently needed to evaluate possible impacts of H. axyridis on the Ecuadorian biodiversity and agroindustry.

Effect of gamma irradiation and subsequent cold storage on the development and predatory potential of seven spotted ladybird beetle Coccinella septempunctata Linnaeus (Coleoptera; Coccinellidae) larvae

Seven spot ladybird beetle, (Coccinella septempunctata) is a widely distributed natural enemy of soft-bodied insect pests especially aphids worldwide. Both the adult and larvae of this coccinellid beetle are voracious feeders and serve as a commercially available biological control agent around the globe. Different techniques are adopted to enhance the mass rearing and storage of this natural enemy by taking advantage of its natural ability to withstand under extremely low temperatures and entering diapause under unfavorable low temperature conditions. The key objective of this study was to develop a cost effective technique for enhancing the storage life and predatory potential of the larvae of C. septempunctata through cold storage in conjunction with the use of nuclear techniques, gamma radiations. Results showed that the host eating potential of larvae was enhanced as the cold storage duration was increased. Gamma irradiation further enhanced the feeding potential of larvae that were kept under cold storage. Different irradiation doses also affected the development time of C. septempuntata larvae significantly. Without cold storage, the lower radiation doses (10 and 25 GY) prolonged the developmental time as compared to un-irradiated larvae. Furthermore, the higher dose of radiation (50GY) increased the developmental time after removal from cold storage. This study first time paves the way to use radiation in conjunction with cold storage as an effective technique in implementation of different biological control approaches as a part of any IPM programs

Citizen science reveals the distribution of the invasive harlequin ladybird (Harmonia axyridis Pallas) in Argentina

The harlequin ladybird (Harmonia axyridis) was introduced in numerous countries around the world as a biological control agent but nowadays it is considered as a pest itself. It is a highly invasive species that has negative impacts on native biodiversity, may affect some crops, and causes a nuisance to humans. Despite its relevance worldwide, the invasion by H. axyridis in South America has received little attention. Here we used a citizen science approach to study the distribution of H. axyridis in Argentina where the species was intentionally introduced in 1986. We also informally tested and evaluated the preferences for different methods for reporting records: iNaturalist website, Google form, email, WhatsApp, and Facebook. Most records were reported through WhatsApp and iNaturalist. We found that H. axyridis is widely distributed in Argentina but there are large gaps, particularly in arid regions. The invasion seems to be recent in the south of the country. Citizen science is a valuable tool for mapping invasive alien species in a large country like Argentina, and will help to monitor the continuous expansion of H. axyridis.

Across-stage consequences of thermal stress have trait-specific effects and limited fitness costs in the harlequin ladybird, Harmonia axyridis

The fitness consequences of thermal plasticity of functional traits of insects, and the life stages at which these responses manifest, remain a subject of much debate. Here, we examine whether a short-term exposure to cold, medium or warm fluctuating temperature regimes during development (larval or adult) affects adult performance, thermal tolerance and fitness of the globally invasive harlequin ladybird Harmonia axyridis. We found plastic responses of a metric of heat stress resistance—critical thermal maximum—and of preferred body temperature after adult temperature exposure, but not in other traits measured. By contrast, exposure of larval stages resulted in plasticity of adult walking performance (height and breadth of the curve) but not tolerance or preference. We found distinct fitness responses between larval and adult treatments, but a composite fitness index revealed negligible effects on reproductive output. These results suggest that the drivers underlying the plasticity of temperature tolerance and temperature selection are different to those shaping the plasticity of walking speed. By testing specific predictions based on current theory of developmental and reversible plasticity, this study contributes novel data to plastic responses of behaviour, stress resistance and fitness to temperature exposure across life stages and thus, provides insights to the broader evolutionary and ecological significance of these responses.

Factors determining variation in colour morph frequencies in invasive Harmonia axyridis populations

The Harlequin ladybird Harmonia axyridis Pallas, native to eastern Asia, is an invasive, non-native species that has recently achieved an almost worldwide distribution. A conspicuous feature of this species is colour polymorphism of the elytra. In its native area, the populations consist of a recessive non-melanic morph, several dominant melanic morphs and small numbers of other (rare) morphs. The morph proportions in native populations have been intensively studied and vary with geographic area, climate and time. In contrast, colour polymorphism in invaded regions has been little studied. We examine and try to account for the morph frequencies observed across the different invaded regions. In America, monomorphic populations consist of the non-melanic morphs while European populations contain also melanic morphs. In particular geographic areas of Europe, the average percentage of the non-melanic morphs varied between 78 and 99%. It was highest in the lowlands of northern Italy and central and northern Europe and decreased in the Alps and western (Spain, UK) and eastern (southeast Russia) margins of the recently invaded area. In central Europe the frequency of the non-melanic morphs decreased over the course of the year but increased over the years from 2010 to 2018. The local differences might thus arise through gradual change of the morph composition of the founder invasive, non-native population. However, the variation in non-melanic morph frequency was not correlated with climatic characteristics that might affect coccinellid polymorphism. The observed rate of change in morph proportions in our data was too small to explain the diversification of what was supposedly a uniform invasive, non-native population at the point of introduction.

Actions on mammalian and insect nicotinic acetylcholine receptors of harmonine-containing alkaloid extracts from the harlequin ladybird Harmonia axyridis

The harlequin ladybird, Harmonia axyridis (H. axyridis), possesses a strong chemical defence that has contributed to its invasive success. Ladybird beetle defensive chemicals, secreted in response to stress and also found on the coating of laid eggs, are rich in alkaloids that are thought to be responsible for this beetle's toxicity to other species. Recent studies have shown that alkaloids from several species of ladybird beetle can target nicotinic acetylcholine receptors (nAChRs) acting as receptor antagonists, hence we have explored the actions of alkaloids of the ladybird H. axyridis on both mammalian and insect nAChRs. Electrophysiological studies on native and functionally expressed recombinant nAChRs were used to establish whether an alkaloid extract from H. axyridis (HAE) targeted nAChRs and whether any selectivity exists for insect over mammalian receptors of this type. HAE was found to be an inhibitor of all nAChRs tested with the voltage-dependence of inhibition and the effect on ACh EC50 differing between nAChR subtypes. Our finding that an HAE fraction consisting almost entirely of harmonine had a strong inhibitory effect points to this alkaloid as a key component of nAChR inhibitory actions. Comparison of HAE inhibition between the mammalian and insect nAChRs investigated indicates some preference for the insect nAChR supporting the view that investigation of ladybird alkaloids shows promise as a method for identifying natural product leads for future insecticide development.

Fifty Shades of the Harlequin Ladybird and a Sexually Transmitted Fungus.

The ectoparasitic fungus Hesperomyces virescens was studied on its invasive host, the harlequin ladybird Harmonia axyridis, in the Czech Republic. A primary aim was to examine the relationship between fungal infection and elytral coloration of the ladybird. Furthermore, the role of host sex and mating status of females were analyzed. Beetles (n = 1,102) were sampled during autumn migration, and then sexed, weighed, and screened for infection. Females were dissected for detection of sperm in their spermathecae. Ladybirds were sorted according to color form and absorbance spectrophotometry was used to quantify carotenoid contents in their elytra. In individuals of the nonmelanic succinea form, the degree of melanization was measured using digital photographs and putative age groups were estimated based on background color of elytra. Sexual differences in infection patterns indicated transmission during copulation: males were infected mostly on elytra and venter, and females had infection almost exclusively on elytra. Mated females had higher infection rate than virgins. There was no influence of genetic color form on the fungal infection. Putative age groups (visual sorting to yellow, orange, and red) correlated with fungal infection. Infected individuals had elevated elytral carotenoid levels in comparison to uninfected individuals, which could be explained by host age. Infection-free succinea beetles were extensively melanized because they emerged later in the season at lower temperatures which induced melanization. Overall, we highlight that H. axyridis is a multivoltine species whose age, if not taken into account in ecophysiological studies, might present a considerable confounding factor.

Silencing of the DNA methyltransferase 1 associated protein 1 (DMAP1) gene in the invasive ladybird Harmonia axyridis implies a role of the DNA methyltransferase 1-DMAP1 complex in female fecundity.

The invasive harlequin ladybird Harmonia axyridis is a textbook example of polymorphism and polyphenism as the temperature during egg development determines the frequency of melanic morphs and the number and size of black spots in nonmelanic morphs. Recent concepts in evolutionary biology suggest that epigenetic mechanisms can translate environmental stimuli into heritable phenotypic changes. To investigate whether epigenetic mechanisms influence the penetrance and expressivity of colour morphs in H. axyridis, we used RNA interference to silence key enzymes required for DNA methylation and histone modification. We found that neither of these epigenetic mechanisms affected the frequency of different morphs, but there was a significant impact on life-history traits such as longevity and fecundity. Strikingly, we found that silencing the gene encoding for DNA methyltransferase 1 associated protein 1 (DMAP1) severely reduced female fecundity, which correlated with an abundance of degenerated ovaries in DMAP1-knockdown female beetles. Finally, we observed significant differences in DMAP1 expression when we compared native and invasive H. axyridis populations with a biocontrol strain differing in egg-laying capacity, suggesting that the DNA methyltransferase 1-DMAP1 complex may influence the invasive performance of this ladybird.

Invasion success of a widespread invasive predator may be explained by a high predatory efficacy but may be influenced by pathogen infection

Invasive alien species (IAS) can drive community change through ecological interactions. Parasites and pathogens can play an important role in community function including mitigating or enhancing IAS impacts. Despite this, the degree to which pathogen pressure influences IAS impacts remains poorly understood. We quantified the predatory behaviour of the highly invasive alien harlequin ladybird (Harmonia axyridis) and two UK native species, the 7-spot (Coccinella septempunctata) and 2-spot (Adalia bipunctata) ladybirds, using comparative functional response experiments. We investigated the impacts of pathogen infection on the predatory ability of the ladybirds by exposing individuals to Beauveria bassiana, a widespread entomopathogen. Invasive H. axyridis was a more efficient predator than both the native A. bipunctata and C. septempunctata, often having higher attack and/or lower prey handling time coefficients, whereas native A. bipunctata were the least efficient predators. These differences were found in both adult and larval life-stages. Beauveria bassiana infection significantly altered the predatory efficiency of adult and larval ladybird predators. The effects of pathogenic infection differed between species and life-stage but in many cases infection resulted in a reduced predatory ability. We suggest that the interactions between IAS and pathogens are integral to determining invasion success and impact.

Microsatellite Polymorphism Suggests High Genetic Diversity But Disrupted Gene Flow in the Two-Spot Ladybird Adalia bipunctata (Linnaeus, 1758) (Coleoptera: Coccinellidae) Populations from Diverse Environments

The two-spot ladybird Adalia bipunctata (Linnaeus, 1758) (Coleoptera: Coccinellidae) is a widely distributed generalist, feeding preferably on aphids. The species inhabits various environments, including anthropogenic landscape, where pollution and microclimate enables to attain very high numbers of favorite prey. Recently, the abundance of the two-spot ladybird has declined, mainly as an after-effect of invasion of the competing species — the harlequin ladybird Harmonia axyridis (Pallas, 1773). High phenotypic polymorphisms made the two-spot ladybird classic model for population genetic studies of Coccinellidea. However, studies investigating diversity of the species at DNA level have Coccinellidae been scarce. In our investigation we used microsatellite markers to characterize genetic diversity in the two-spot ladybird populations from different environments. We made an attempt to amplify 13 loci, identified previously in the genome of the species, however due to lack of amplification, monomorphisms and presence of the null alleles, the final set of six markers useful for analysis on a population level was obtained. Analyzing genotypes of 124 individuals, we confirmed high genetic diversity of the species, suggested previously by studies, applying allozymes and mitochondrial DNA. Also, microsatellites revealed weakly pronounced genetic structure in a large scale (localities separated by a distance of 500 kilometers). However, we found some indications (signs of the Wahlund effect) that gene flow could be disturbed among sites in diverse environments (rural, suburban and urban). Our data suggested that invasion of the harlequin ladybird has not yet affected populations of the two-spot in such extent to alter genetic diversity or genetic structure.

Effects of so-called "environmentally friendly" agrochemicals on the harlequin ladybird Harmonia axyridis (Coleoptera: Coccinelidae)

A variety of plant protection products and other agrochemicals are used in agro-ecosystems. Products approved for integrated pest management (IPM) or organic farming should have minimal negative side effects on beneficial insects. The Asian harlequin ladybird Harmonia axyridis (Pallas, 1773) (Coleoptera: Coccinellidae) has become a widespread and important generalist predator of certain agricultural pests, mainly aphids, throughout Europe. We studied the effects of two agrochemicals, Boundary SW® (auxiliary plant protection product) and Prev B2® (foliar boron fertilizer), usually regarded as environmentally friendly and known to have insecticidal side effects against some fruit and vegetable pests (e.g., aphids, spider mites, weevils), on the last larval instar and adults of Harmonia axyridis. The conventional organophosphate insecticide Reldan 22® was used as a chemical standard for evaluating the lethal effect, because this product is usually effective against a broad spectrum of insects, and indeed was immediately lethal for both the adults and larvae of this species. However, whereas Prev B2® had no effect, adult ladybirds sprayed with Boundary SW® survived only for up to 25 h and also none of the larvae completed their development. Thus, although our experiments were not made under natural conditions, the use of Boundary SW® cannot be recommended for IPM and organic farming in terms of safeguarding insect predators such as Harmonia axyridis until further more detailed testing.

Response of passerine birds and chicks to larvae and pupae of ladybirds

1. Few, if any, experimental tests have demonstrated the anti‐predator protection of the developmental stages of ladybirds (Coleoptera: Coccinellidae) against vertebrates, despite the fact that both the visual appearance of ladybirds and the content of defensive compounds fulfil the definition of an aposematic prey.2. In this study, avian predators of three species were confronted with fourth‐instar larvae and pupae of the harlequin ladybird (Harmonia axyridis) – a large, conspicuous, and toxic ladybird species.3. The selected predators differed in their individual experience and attitude to ladybirds. Wild‐caught great tits (Parus major) strongly avoided attacking ladybirds in general, whereas wild‐caught tree sparrows (Passer montanus) were willing to include ladybirds in their diet. Domestic chicks (Gallus gallus domesticus) have never been confronted with ladybirds but usually show avoidance of aposematic signals. In this study, great tits and domestic chicks avoided both developmental stages, but in the case of chicks the avoidance had to be learned over the course of repeated encounters. Sparrow avoidance was significantly lower, with more than one‐third of the prey being attacked and eaten.4. The protection of both developmental stages of ladybirds is similar to adults, despite substantially different visual appearance.

Impact of the neozoic ladybird Harmonia axyridis on the coccinellid community in Bohemia

Since 2007, when Asian harlequin ladybird Harmonia axyridis (Pallas) has become a part of coccinellid entomofauna in Bohemian hop gardens, the spectrum of aphidophagous coccinelids has changed considerably. Whereas, in the period before its invasion native species Coccinella septempunctata L., Adalia bipunctata L. and Propylea quatuordecimpunctata L. used to be the dominant predators of damson hop aphid (Phorodon humuli Schrank), Harmonia axyridis has been the most frequent ladybird species in Zatec (Saaz) hop growing region nowadays. Population densities of adults of aphidophagous ladybirds in the middle of the eighties (1985), shortly before the invasion of the above-mentioned Asian species (2004) and in the recent years (2011 and 2015) are compared. Whereas four years ago harlequin ladybird represented just 10% of the total Coccinellidae, in 2015 it forms nearly half of coccinellid adults observed in hop gardens in NW Bohemia. It is followed by tiny species P. quatuordecimpunctata and A. bipunctata, which consist together more than one third, whereas in the middle of the eighties they generally formed two thirds of the coccinellid populations. On the other hand adults of C. septempunctata have become less abundant. It may be caused by aggressiveness of H. axyridis, whose larvae were observed to feed on eggs and young larvae of C. septempunctata. To prevent attacks from larvae of H. axyridis, females of C. septempunctata appear on hop plants and begin to lay eggs earlier as illustrated by the presence of the third and the fourth instars larvae in the time when larvae of H. axyridis reach much smaller size and are not competitive enough. Decline in population densities of native species of ladybirds on response to arrival of the Asian invasive species, their responses and potential efficiency of aphidophagous coccinelids, including H. axyridis, on damson hop aphid will need to be observed in the future so as us to know more about this issue.

Insects in anthelminthics research: Lady beetle-derived harmonine affects survival, reproduction and stem cell proliferation of Schistosoma mansoni.

Natural products have moved into the spotlight as possible sources for new drugs in the treatment of helminth infections including schistosomiasis. Surprisingly, insect-derived compounds have largely been neglected so far in the search for novel anthelminthics, despite the generally recognized high potential of insect biotechnology for drug discovery. This motivated us to assess the antischistosomal capacity of harmonine, an antimicrobial alkaloid from the harlequin ladybird Harmonia axyridis that raised high interest in insect biotechnology in recent years. We observed remarkably pleiotropic effects of harmonine on physiological, cellular, and molecular processes in adult male and female Schistosoma mansoni at concentrations as low as 5 μM in vitro. This included tegumental damage, gut dilatation, dysplasia of gonads, a complete stop of egg production at 10 μM, and increased production of abnormally shaped eggs at 5 μM. Motility was reduced with an EC50 of 8.8 μM and lethal effects occurred at 10–20 μM within 3 days of culture. Enzyme inhibition assays revealed acetylcholinesterase (AChE) as one potential target of harmonine. To assess possible effects on stem cells, which represent attractive anthelminthic targets, we developed a novel in silico 3D reconstruction of gonads based on confocal laser scanning microscopy of worms after EdU incorporation to allow for quantification of proliferating stem cells per organ. Harmonine significantly reduced the number of proliferating stem cells in testes, ovaries, and also the number of proliferating parenchymal neoblasts. This was further supported by a downregulated expression of the stem cell markers nanos-1 and nanos-2 in harmonine-treated worms revealed by quantitative real-time PCR. Our data demonstrate a multifaceted antischistosomal activity of the lady beetle-derived compound harmonine, and suggest AChE and stem cell genes as possible targets. Harmonine is the first animal-derived alkaloid detected to have antischistosomal capacity. This study highlights the potential of exploiting insects as a source for the discovery of anthelminthics.