Substrate Marking by an Invasive Ladybeetle: Seasonal Changes in Hydrocarbon Composition and Behavioral Responses

The multicolored Asian ladybeetle, Harmonia axyridis (Pallas), aggregates inside dwellings during the winter to survive the cold. Recent published reports have highlighted that overwintering individuals use hydrocarbon markings deposited on surfaces by conspecifics to orient toward aggregation sites. In the current study, monthly GC-MS analyses revealed seasonal modifications in the chemical profile of substrate markings deposited by moving individuals. The markings of overwintering ladybeetles contained larger proportions of heptacosadiene, nonacosadiene, hentriacontadienes, and methyl-nonacosanes, along with a lower proportion of heptacosene and nonacosene. This finding suggests the importance of the unsaturated and/or branched hydrocarbons in the H. axyridis aggregation process. Subsequently, we conducted behavioral assays to test whether (1) there is seasonal variation in the behavioral response of H. axyridis individuals toward substrate markings deposited by conspecifics in the same physiological state and (2) the observed behavioral modification is due to a change in ladybeetle sensitivity and/or a change in the chemical composition of the substrate marking. The results indicate that overwintering individuals exhibit a stronger “following” response toward conspecific substrate markings. This behavior is linked to both the physiological state of ladybeetles and the specific chemical profile of the marking biomolecules deposited under overwintering conditions.     

Role of long-chain hydrocarbons in the aggregation behaviour of Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae)

The multicoloured Asian ladybirds, Harmonia axyridis (Pallas), form large aggregations inside dwellings to survive cold winters, causing annoyance to householders due to their abundance, allergenic properties and problems consequent on reflex bleeding. Flight to overwintering sites and macrosite choice of this species is well documented. H. axyridis shows a hypsotactic behaviour and a clear preference for contrasting visual elements. However, how the microsite is selected remains undocumented, although a better understanding of the factors implicated could lead to the development of new control methods for this pest. In this work, we hypothesised that non-volatile compounds are involved in the microsite choice and aggregation of this beetle. Long chain hydrocarbons were identified at aggregation sites, comprising saturated and unsaturated homologues. An aggregation bioassay was then conducted on overwintering individuals, highlighting the retention capacity of the identified compounds on the tested ladybirds. Additional investigations showed that H. axyridis males and females, originating from overwintering sites, deposit a similar blend of molecules while walking. A Y-shaped tube assay revealed that this blend is used by male and female congeners as cue allowing individuals to orientate towards the treated side of the olfactometer. These results suggest the use of two different blends of long chain hydrocarbons by H. axyridis during its aggregative period, the first one to lead conspecifics towards aggregation sites (microsites) and the second to ensure the cohesion of the aggregation. These findings support the potential use of these blends, in association with volatiles, in the design of traps in order to control infestations of this species in dwellings.     

Is Contact Between Conspecifics Involved in the Cohesion of Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae) Aggregations?

The multicolored Asian ladybeetle, Harmonia axyridis (Pallas), exhibits a gregarious behavior during unfavorable winter conditions. Although this behavior is currently described as a phenomenon occurring only during winter, H. axyridis aggregations can also be observed outside overwintering conditions. However, the substrate markings previously highlighted as being involved in the wintry aggregation of this exotic species do not seem to be used by non-overwintering individuals to aggregate. This fact suggests then that other cues are responsible for the induction of this behavior. In this work, we have tested the hypothesis that direct contact between non-overwintering individuals stimulates the establishment of clusters. Binary choice experiments highlighted the involvement of elytral cuticular compounds in this phenomenon. Chromatographic analyses showed that the active extracts contained mainly hydrocarbons, including saturated, mono-unsaturated, and di-unsaturated homologues. Physical contact also seems to be involved in the non-overwintering aggregative behavior of H. axyridis, but to a lesser extent than these natural compounds. These findings could eventually be used to develop new control methods of these pest populations and so, reduce the adverse impacts it causes on biodiversity.     

Mode of overwintering of invasive <Emphasis Type="Italic">Harmonia axyridis</Emphasis> in the Netherlands

After establishment of Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae) in Europe, population densities of native ladybird species have decreased. The post-hibernation onset of female reproduction, a key characteristic influencing population dynamics and competition with related species, was studied. Hibernating individuals were collected and transferred to outdoor cages to continue overwintering. Every two weeks a sample of individuals was transferred to long-day, warm conditions. Intensity of dormancy was studied by determining the pre-oviposition period and ovarian development. Pre-oviposition periods were short throughout our observations, indicating that Harmonia axyridis was not in diapause but in a quiescent state. H. axyridis becomes active rapidly when temperature rises in spring but is not active earlier in the year than native species. Neither the mode of overwintering, nor the onset of spring activity can explain the invasion success of H. axyridis.     

The chemical ecology of <Emphasis Type="Italic">Harmonia axyridis</Emphasis>

We review the chemical ecology of the ladybird beetle Harmonia axyridis from the perspective of its invasiveness and the deleterious effects it exerts in the regions it has colonised. We outline the nature and quantification of its chemical defence, and discuss the protection this provides against natural enemies, particularly intraguild predators. We consider the role of infochemicals in location of prey, intraspecific communication and intraguild interactions. We also discuss the role of prey allelochemicals in relation to H. axyridis extreme dietary generalism. Harmonia axyridis poses a number of practical problems for human health and well-being, including “ladybug taint” wine contamination and problems resulting from large aggregations overwintering in buildings. We consider chemical insights into these issues and, in particular, how attractants and repellents might help manage H. axyridis populations through a push–pull strategy. We conclude by discussing future perspectives for research.     

Winter survival of Harmonia axyridis in The Netherlands

Since the establishment of Harmonia axyridis in Europe, populations of native ladybirds have decreased. Overwintering survival is one of the aspects of the biology of H. axyridis that may contribute to its firm establishment in and invasion of a new area. In this study winter survival of five wild H. axyridis populations was assessed under natural and semi-natural conditions, with a focus on the potential influence of location and orientation on winter survival.Overwintering survival of H. axyridis in the Netherlands is high: 70.8-88.2%. When overwintering at one central site, populations sampled at five locations showed statistically significant different mortality rates. Furthermore, winter survival of H. axyridis at the sample sites was higher when beetles were hibernating at the southwestern sides of buildings, where most aggregations of ladybirds were found. Survival was higher at sheltered sites compared to exposed sites.Harmonia axyridis has a comparable or higher overwintering survival than most common native ladybird species. A high overwintering survival results in a large post-hibernation population in spring, leading to a rapid population build-up. Thus, the high winter survival probably contributes to the success of the exotic H. axyridis.     

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.     

The aggregation behavior of <Emphasis Type="Italic">Harmonia axyridis</Emphasis> in its native range in Northeast China

Harmonia axyridis has become notorious as an urban pest in many of the regions where it has been introduced, despite its numerous contributions to the biological control of insects injurious to agriculture and horticulture. Aggregative behavior prior to overwintering leads to invasions of human habitations as beetles seek refuge from freezing temperatures. Here we describe the aggregation behavior of native H. axyridis populations of northeast China that breed in agricultural fields (mostly corn and rice) and shrub/forest habitats and then migrate through rural villages in autumn. More than 140,000 beetles were collected during direct observations in 16 villages in five townships in Jilin Province. Beetles aggregated on dwellings shortly after agricultural harvests, favoring white walls with southern exposures, the largest aggregations occurring in villages in mountainous townships at higher elevations. The sex ratio was consistently female-biased and succinic phenotypes were more than twice as abundant as melanic phenotypes in all locations. A special trap compared the relative attractiveness of different surface colors (white > yellow = black > green > red = natural wood) and potential baits (corn pollen = honey > caramel = cocoa > milk = blank control). All aggregations disappeared abruptly just prior to the first frost, whereupon beetles were discovered sheltering in montane caves with southern aspects at higher elevations. Villagers reported substantial expenditures on pesticides in efforts to eliminate beetles from their homes every autumn. Invasion of human habitations appears to be an intrinsic tendency of native H. axyridis populations in China, which is the result of behavioral adaptations for cold-avoidance.     

Horizontal transmission of a microsporidium from the convergent lady beetle, Hippodamia convergens Guérin-Méneville (Coleoptera: Coccinellidae), to three coccinellid species of Nova Scotia

Convergent lady beetles, Hippodamia convergens Guérin-Méneville, are a popular choice for aphid control in North America. An unidentified microsporidium was found in H. convergens adults that were purchased from a commercial insectary in 2004. This study examined egg cannibalism and egg predation as a means of horizontal transmission of the unidentified microsporidium among H. convergens larvae and three coccinellid species found in Nova Scotia: Coccinella septempunctata (seven-spotted lady beetle), C. trifasciata perplexa (three-banded lady beetle), and Harmonia axyridis (multicolored Asian lady beetle). The microsporidium was transmitted with 100% efficiency when first instars fed on microsporidia-infected eggs. Mean spore count data from smear preparations of infected beetles suggest that the infection was as heavy in C. trifasciata perplexa (a native coccinellid) (11.2 ± 0.96 spores/100 μm²) as it was in H. convergens (the natural host) (12.8 ± 1.16) but lighter in the introduced species C. septempunctata (7.5 ± 0.65) and H. axyridis (0.8 ± 0.11). For all of the beetle species examined, larval development was significantly longer for microsporidia-infected individuals than for their uninfected cohorts. The microsporidium had no effect on larval mortality. Based on the results of this study, field-collected H. convergens should be examined for microsporidia and uninfected individuals should be used to rear individuals for release in biological control programs. However, this is unlikely to happen because H. convergens are relatively easy and inexpensive to collect from their overwintering sites for redistribution.     

Do microsporidia function as “biological weapon” for Harmonia axyridis under natural conditions?

Invasive alien species, such as the multicoloured Asian ladybird Harmonia axyridis, are often regarded as major drivers of biodiversity loss. Therefore understanding which characteristics or mechanisms contribute to their invasive success is important. Here the role of symbiotic microsporidia in the hemolymph of H. axyridis was investigated in the context of intraguild predation between wild‐caught H. axyridis and the native ladybird species Coccinella septempunctata. The microsporidia were recently discussed to contribute to the unpalatability of Harmonia for other coccinellids during intraguild predation and to function as “biological weapons”. In the present study, visual detection of microsporidia in hemolymph samples revealed that 73.5 % of H. axyridis were infected. Intraguild predation experiments between larvae of the two species showed a significant competitive advantage for H. axyridis, even against larger larvae of C. septempunctata. Adult C. septempunctata always killed and fed on H. axyridis larvae. However only 11.4 % (4 of 47) of C. septempunctata that fed on infected H. axyridis died within 4 months. In contrast to previous studies this suggests that microsporidia or harmonine, the chemical defense compound of H. axyridis, do not lead to death of C. septempunctata preying on larvae of H. axyridis. Instead our results support the idea that competitive advantage during intraguild predation greatly facilitates the success of H. axyridis and that this may help this highly invasive species to outcompete native species. The impact of microsporidia on Harmonia itself as well as on interspecific interactions require further studies.