Chemical protection of Calvia quatuordecimguttata eggs against intraguild predation by the invasive ladybird Harmonia axyridis

Intraspecific and interspecific predation of eggs is a well documented phenomenon amongst aphidophagous coccinellids. The invasive species Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae) is known to be a top intraguild predator and reported to attack the eggs of many coccinellid species both in a laboratory setting and in the wild. A previous laboratory study highlighted that while many species’ eggs were highly palatable to H. axyridis, the eggs of Calvia quatuordecimguttata (Linnaeus) (Coleoptera: Coccinellidae) appeared to be extremely well protected from attack. Here we present the results of behavioural experiments testing the hypothesis that substances on the egg surface are responsible for this protection, and report preliminary results of GC-MS analysis of these compounds. When the coatings of C. quatuordecimguttata eggs were removed using hexane, they became significantly more susceptible to predation by neonate H. axyridis larvae. However, their overall palatability was not affected, in that complete consumption was never or rarely observed. This suggests that the surface compounds are a true indicator of unpalatability in this species. The effect of hexane-washing on already palatable conspecific eggs was also analysed but had no significant effect on the susceptibility of eggs to cannibalism. We conclude that the eggs of at least one European species are effectively protected by surface deterrents from intraguild predation by H. axyridis. This effect might be due to both the diversity and abundance of hydrocarbons present within the egg coating, the presence of alkenes and/or the presence of patches of a red substance on the eggs’ surface, which is thought to belong to the acid group. In conjunction with data on the susceptibility of other immature stages of C. quatuordecimguttata, this finding may indicate a decreased risk of the species falling victim to invasive H. axyridis, despite their coincident habitat ranges.     

Invasion history, habitat preferences and phenology of the invasive ladybird Harmonia axyridis in Belgium

Harmonia axyridis Pallas (Coleoptera: Coccinellidae) is native to Asia, and was widely introduced as a biocontrol agent of aphids and coccids in Europe and North America. In Europe, H. axyridis is considered to be an invasive alien species because of its potential to disrupt native ladybird communities. Since 1999, the Belgian Ladybird Working Group mapped all Belgian Coccinellidae and recorded data on substratum plants and habitat. The first feral H. axyridis population in Belgium was recorded in 2001, but the expansion rate is decreasing because it now colonised the whole country. Recorded occupancy in Belgium showed an average rate of increase of 189% between 2002 and 2006. In Belgium, H. axyridis occurred in a wide range of habitats, including those of high conservation value. However, habitat and land cover analysis showed that H. axyridis is more frequently found in urbanised landscapes than in semi-natural landscapes. A niche overlap analysis based on plant use data showed that the potential to affect native species is higher for generalist, deciduous and coniferous tree ladybird species than for heathland and wetland specialist species. Phenology data showed that H. axyridis is able to reproduce later in the year than native species. Based on recorded distribution, ecology and phenology, we discuss the potential of H. axyridis to disrupt native ladybird assemblages in Belgium.     

Intraguild predation involving Harmonia axyridis: a review of current knowledge and future perspectives

As an effective generalist predator of aphids and other hemipteran pests H. axyridis has been a successful biological control agent. However, the very functional traits that have contributed to its success in this regard also implicate it as an intraguild predator that poses a significant risk not only to the diversity of other natural enemies of Hemiptera (and their associated ecosystem services), but to biodiversity more widely. In this paper we will specifically review the existing data on intraguild predation involving H. axyridis, and consider the strength and symmetry of such interactions both within its native guild and within exotic guilds where it has established as an invasive alien. We will use these studies to interpret the observed population declines in predator diversity in the field, predict species at risk in regions not yet invaded and consider implications for resulting ecosystem services. We will also indicate gaps in our knowledge that require further study in order to identify opportunities for mitigation.     

Harmonia axyridis in Europe: spread and distribution of a non-native coccinellid

Native to Asia, Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae) is considered an invasive alien ladybird in Europe and North America, where it was widely introduced as a biological control agent of aphids and coccids. In Europe, H. axyridis was sold by various biological control companies from 1995 in France, Belgium and the Netherlands, and was also intentionally released in at least nine other countries. It has spread very rapidly, particularly since 2002, and is now regarded as established in thirteen European countries. The established range extends from Denmark in the north to southern France in the south, and from Czech Republic in the east to Great Britain in the west. In this paper we map the spread and distribution of H. axyridis in Europe, and examine the situation on a country-by-country basis. We report first records of the species in five countries; Spain, Sweden, Denmark, Czech Republic and Italy; and first evidence of H. axyridis establishment in the latter three countries. Despite releases of H. axyridis in Portugal, Spain and Greece, there is little evidence of establishment in southern Europe. It is predicted that the spread and increase within Europe will continue and that H. axyridis will become one of the most widely distributed coccinellids in the continent.     

A natural flightless mutation in the ladybird, Harmonia axyridis

A homozygous flightless strain, obtained by selection from a laboratory population of Harmonia axyridis Pallas (Coleoptera, Coccinellidae), has been reared under controlled conditions since 1982 after sampling in China. The adults of this flightless strain have normal elytra and wings but drop almost vertically when they attempt to fly. The origin of this mutation, whether laboratory or natural, is discussed. As the mutation does not affect the fitness of the beetles, this flightless strain can be used in biological control. Releasing flightless adults rather than larvae should provide the potential for more continuous control of aphids by both larvae and adults.     

Phenotypic variation in invasive and biocontrol populations of the harlequin ladybird, Harmonia axyridis

Despite numerous releases for biological control purposes during more than 20 years in Europe, Harmonia axyridis failed to become established until the beginning of the 21st century. Its status as invasive alien species is now widely recognised. Theory suggests that invasive populations should evolve toward greater phenotypic plasticity because they encounter differing environments during the invasion process. On the contrary, populations used for biological control have been maintained under artificial rearing conditions for many generations; they are hence expected to become specialised on a narrow range of environments and show lower phenotypic plasticity. Here we compared phenotypic traits and the extent of adaptive phenotypic plasticity in two invasive populations and two populations commercialized for biological control by (i) measuring six phenotypic traits related to fitness (eggs hatching rate, larval survival rate, development time, sex ratio, fecundity over 6 weeks and survival time of starving adults) at three temperatures (18, 24 and 30°C), (ii) recording the survival rate and quiescence aggregation behaviour when exposed to low temperatures (5, 10 and 15°C), and (iii) studying the cannibalistic behaviour of populations in the absence of food. Invasive and biocontrol populations displayed significantly different responses to temperature variation for a composite fitness index computed from the traits measured at 18, 24 and 30°C, but not for any of those traits considered independently. The plasticity measured on the same fitness index was higher in the two invasive populations, but this difference was not statistically significant. On the other hand, invasive populations displayed significantly higher survival and higher phenotypic plasticity when entering into quiescence at low temperatures. In addition, one invasive population displayed a singular cannibalistic behaviour. Our results hence only partly support the expectation of increased adaptive phenotypic plasticity of European invasive populations of H. axyridis, and stress the importance of the choice of the environmental parameters to be manipulated for assessing phenotypic plasticity variation among populations.     

Mating refusal and its significance in females of the ladybird beetle, Harmonia axyridis

ABSTRACT. . Mating receptivity in females of the ladybird, Harmonia axyridis Pallas (Coleoptera: Coccinellidae), was studied with reference to internal and environmental conditions. Females sometimes reject copulation: they run away, or shake off the male and bend the abdomen upwards. Newly emerged females show a greater likelihood of accepting copulation with an advance in the stage of ovarian development, this being related to age. Females usually mate repeatedly, but frequently show refusal behaviour under unfavourable food conditions. Dissections reveal that females which accept copulation have a greater number of mature eggs in the ovary than do rejectors. Results also suggest that the food conditions are a major environmental factor, directly controlling mating receptivity.     

Intraguild predation between the aphidophagous ladybird beetles Harmonia axyridis and Coccinella undecimpunctata (Coleoptera: Coccinellidae): The role of intra and extraguild prey densities

Laboratory cages were used to evaluate the influence of extraguild (EGprey) and intraguild prey (IGprey) densities on the direction, symmetry and magnitude of the intraguild predation (IGP) of the aphidophagous Harmonia axyridis Pallas on Coccinella undecimpunctata L. and vice versa. In order to understand the role of competition between IGprey, the experiments included treatments with one IGpredator, with one or four IGprey and EGprey (Aphis fabae Scopoli) ranging from zero to sufficient aphids to satiate the predators for 12, 24 or 48 h. Increases in EGprey and IGprey densities did not alter the direction, but decreased the magnitude and symmetry of IGP. Predation on one individual of IGprey decreased from more than 80%, in the absence of EGprey, to from 6% to 53%, at higher EGprey densities. Decrease in IGP was less when H. axyridis was the IGpredator. Even at high EGprey densities, eggs and 2nd larval stages of C. undecimpunctata were vulnerable to IGP and the level of predation was 40% and 53%. The presence of more than one IGprey increased the magnitude of IGP mainly at EGprey densities sufficient to satiate the predators for 12 and 24 h, suggesting that competition between the IGpredator and IGprey may be one of the processes promoting IGP. These results and those of other authors suggest that H. axyridis has the potential to be an IGpredator, mainly of the most vulnerable stages of IGprey. Thus, H. axyridis may negatively affect the survival of C. undecimpunctata, when these two species exploit the same resources.     

Predicting the potential geographical distribution of the harlequin ladybird, Harmonia axyridis, using the CLIMEX model

Harmonia axyridis (Pallas, 1773) (Coleoptera: Coccinellidae) is a ladybird beetle native to temperate and subtropical parts of Asia. Since 1916 populations of this species have been introduced throughout the world, either deliberately, or by accident through international transport. Harmonia axyridis was originally released as a classical biological control agent of aphid and coccid pests in orchards and forests, but since 1994 it is also available as a commercial product for augmentative control in field and greenhouse crops. It is a very voracious and effective natural enemy of aphids, psyllids and coccids in various agricultural and horticultural habitats and forests. During the past 20 years, however, it has successfully invaded non-target habitats in North America (since 1988), Europe (1999) and South America (2001) respectively in a short period of time, attacking a wide range of non-pest species in different insect orders. Becoming part of the agricultural commercial pathway, it is prone to being introduced into large areas across the world by accident. We use the CLIMEX programme (v2) to predict the potential geographical distribution of H. axyridis by means of matching the climate of its region of origin with other regions in the world and taking in account biological characteristics of the species. Establishment and spread seem likely in many regions across the world, including those areas which H. axyridis has already invaded (temperate Europe, North America). Based on the CLIMEX prediction a large part of Mediterranean Europe, South America, Africa, Australia and New Zealand seem highly suitable for long-term survival of H. axyridis as well. In addition we evaluate CLIMEX as a strategic tool for estimating establishment potential as part of an environmental risk assessment procedure for biological control agents we discuss biological and ecological aspects necessary to fine-tune its establishment and spread in areas after it has been introduced.     

Interactions between the fungal pathogen Beauveria bassiana and three species of coccinellid: Harmonia axyridis, Coccinella septempunctata and Adalia bipunctata

Harmonia axyridis is a predatory coccinellid, native to central and eastern Asia. It has been available in many countries for use as a biological control agent of pest aphids and scale insects. In many of these countries, including the USA, H. axyridis has established. It is now considered an invasive alien species for a number of reasons, including its impact on functional biodiversity. Beauveria bassiana is known to be a natural mortality agent of overwintering coccinellids and is a potential candidate for the biological control of H. axyridis. In this paper we compare the susceptibility of three species of coccinellid, H. axyridis (cultures derived from Japan and UK), Coccinella septempunctata and Adalia bipunctata to infection by B. bassiana (commercial strain GHA) after exposure at three doses (10⁵, 10⁷, 10⁹ conidia ml⁻¹). The two subpopulations of H. axyridis (Japan and UK) were more resistant to B. bassiana infection than either A. bipunctata or C. septempunctata. This is exemplified by the median lethal doses at 10 days post-inoculation (LD₅₀) of 10^6.2, 10^6.0, 10^8.3, 10^9.6 conidia ml⁻¹ for A. bipunctata, C. septempunctata, H. axyridis (Japan) and H. axyridis (UK), respectively. Only doses of 10⁹ conidia ml⁻¹ resulted in mortality of H. axyridis, in contrast, 80% of C. septempunctata and 70% of A. bipunctata exposed to 10⁷ conidia ml⁻¹ of B. bassiana succumbed to infection. The fecundity (cumulative mean egg production over 10 days) of A. bipunctata and H. axyridis (UK) was also assessed. The fecundity of C. septempunctata could not be assessed because this species requires diapause prior to the onset of reproduction and these studies were on beetles that had recently eclosed (2–8 weeks). Harmonia axyridis (Japan) produced no eggs in most treatments including the control and so was excluded from analysis. High dose (10⁹ conidia ml⁻¹) inoculation reduced the fecundity of A. bipunctata to zero but egg production was similar for individuals inoculated with doses of 10⁵, 10⁷ conidia ml⁻¹ and control individuals. In contrast, all doses of B. basssiana reduced H. axyridis (UK) egg production dramatically. We discuss these results in relation to the potential for control of H. axyridis using B. bassiana.     

Intraguild responses of aphid predators on apple to the invasion of an exotic species, Harmonia axyridis

The effects of the invasion ofan exotic predator, Harmonia axyridis(Pallas) (Coleoptera: Coccinellidae), wereinvestigated using three experiments on theecology of aphid predators on apple. Oneexperiment, 1992, was collected prior to theH. axyridis invasion, and two others,1996 to 1997 and 1999 to 2000, were collectedafter the invasion. Except for one year, 1999,H. axyridis was the dominant coccinellid,replacing the formerly dominant Coccinellaseptempunctata L. (Coleoptera: Coccinellidae),another exotic species. The dominance of H. axyridis was greater among larvae thanamong adults. There was no apparent effect ofthe H. axyridis invasion on abundance ofthe predator, Aphidoletes aphidimyza(Rondani) (Diptera: Cecidomyiidae) and apossible positive effect on the abundance ofchrysopids (Neuroptera: Chrysopidae). Principal component analysis indicated thatalthough individual species were affected, theoverall effect of H. axyridis invasion onthe predator guild as a whole was negligible. The data indicate that the interaction betweenthe two exotic species, H. axyridis andC. septempunctata, may be allowing nativecoccinellids to become more abundant on applethan when C. septempunctata was thedominant coccinellid.     

Locomotor Behavior of Flightless Harmonia axyridis Pallas (Col., Coccinellidae)

To improve the efficiency of the lady beetle H. axyridis as a biological control agent against aphids, a flightless population was obtained by feeding adults with a mutagen and selecting their progeny for nonflying but otherwise morphologically normal individuals. These flightless adults attempted to fly but immediately fell. They softened their fall by opening their elytrae and wings. The inability to fly could result from change in their flying behavior compared to control adults. The flight duration was very much shorter, and the wing beat frequency and, more particularly, the amplitude of the wing beats were clearly lower. More time was spent in the other components of the flight behavior such as wing rotation, wing immobility, and wing folding. The sequence of these patterns differed slightly, due mainly to change in their frequency. The locomotor behavior was not modified by the mutation, which affected only the wing muscles. Searching behavior of mutant adults differed from that of control adults only in that they took longer to encounter and ingest aphids. Nevertheless, the larval growth and reproductive rate remained unchanged. The behavioral and biological features of these flightless adults indicate that it should be possible to use them in biological control programs.     

Estimation of mortality by entomophages on exotic Harmonia axyridis versus native Adalia bipunctata in semi-field conditions in northern Italy

A semi-field experiment was carried out in two peach orchards in northern Italy to assess mortality due to predators and parasitoids on the exotic coccinellid Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae) in comparison with the native coccinellid Adalia bipunctata L. (Coleoptera: Coccinellidae). The experiments were conducted in cages to avoid the possible escape of the exotic ladybird (not yet established in Italy). Two kinds of cage experiments were included: ‘exclusion cages’ (access by walking predators impeded) and ‘free cages’ (walking predators free to enter). The cages, containing all the stages of the two ladybird species, were placed in two localities and left for 24 h. All ladybird stages used for the semi-field experiments came from a laboratory rearing. The eggs of H. axyridis experienced less mortality than those of A. bipunctata. The ant workers were the most frequent predators in ‘free cages’ but A. bipunctata cannibalism on eggs was also detected. Larvae of both coccinellid species were predated equally but larval predation of L1 and L2 was higher in comparison to predation of L3 and L4. Pupae and adults of both exotic and native ladybirds were never attacked by predators. Predation on younger larval stages was higher in the ‘free cages’ in comparison with ‘exclusion cages’. No ladybird parasitisation was observed. The ‘free cage’ technique seems to provide a standardised and realistic estimation of predation impact but more studies are needed to evaluate ladybird parasitisation in semi-field conditions.     

Overwintering, phenology and fecundity of Harmonia axyridis in comparison with native coccinellid species in Italy

Classical biological control is generallyunderstood as an environmentally safe practiceof insect pest management. However, questionshave been raised about possible negativeeffects for native species. As part of a riskassessment study a semi-field trial was carriedout in northern Italy in order to compare theoverwintering of the native species Propylea quatuordecimpunctata (L.) and Adalia bipunctata (L.) (Coleoptera:Coccinellidae) with the exotic Harmoniaaxyridis (Pallas) (Coleoptera: Coccinellidae).All the experiments were carried out from April1998 to April 2001. Harmonia axyridis wasable to overwinter successfully in northernItaly. Fecundity, oviposition rate, longevityand rate of increase of overwintering femaleswere calculated and the phenology of the threespecies was studied. Harmonia axyridisoverwintering mortality (31.9%) was lower incomparison with mortality of native species(68.9% for P. quatuordecimpunctata and61.3% for A. bipunctata). As aconsequence, post-overwintering rate ofincrease of the exotic species was higher.Longevity of overwintered females was similaramong the species. Mean fecundity of H. axyridis (783.8 eggs per female) was slightlyhigher than A. bipunctata (720.2 eggs perfemale), and much higher than P. quatuordecimpunctata (193.7 eggs per female). H. axyridis and A. bipunctatacompleted four generations and P. quatuordecimpunctata three generations in a year. The results show that H. axyridisappears to posses a high potential for establishment in Italy.     

Interaction between Podisus maculiventris and Harmonia axyridis two predators used in augmentative biological control in greenhouse crops

Intraguild predation (IGP) betweenthe pentatomid Podisus maculiventris(Say) and the coccinellid Harmoniaaxyridis (Pallas) in the absence or presenceof the extraguild prey Spodopteralittoralis (Boisduval) and Myzuspersicae (Sulzer) was studied in thelaboratory. Interactions were asymmetric infavor of the pentatomid. Podisusmaculiventris readily fed upon eggs and larvaeof H. axyridis, but adult beetles wererarely attacked. Success of attacks by P.maculiventris was stage dependent, fourthinstars and adults being more successful inkilling ladybeetle larvae than second instars.Attacks by H. axyridis on the pentatomidwere rare and none of them were successful. Theeffect of introducing extraguild prey on thelevel of IGP was tested both in petri dishesand on caged sweet pepper plants. Whensufficient numbers of S. littoralislarvae were present to satiate the pentatomid,predation on H. axyridis larvae decreasedsignificantly, indicating that the coccinellidis a less preferred or less vulnerable prey.When the aphid M. persicae was presentedas extraguild prey, levels of IGP were notaltered. Nymphs of P. maculiventrissuccessfully completed development whenexclusively fed on larvae of H. axyridis,but developmental time was longer than onlepidopteran prey. No pentatomid nymphs reachedadulthood on aphids alone. IGP by P.maculiventris on H. axyridis may be ofsome importance in greenhouse crops, where bothpredators are being used increasingly inaugmentative biological control programs.Nonetheless, it is expected that in practicelarger larvae and adults of H. axyridiswill escape most attacks by the pentatomid.     

Cold tolerance of the harlequin ladybird Harmonia axyridis in Europe

As an essential aspect of its invasive character in Europe, this study examined the cold hardiness of the harlequin ladybird Harmonia axyridis. This was done for field-collected populations in Belgium overwintering either in an unheated indoor or an outdoor hibernaculum. The supercooling point, lower lethal temperature and lower lethal time at 0 and −5 °C were determined. Possible seasonal changes were taken into account by monitoring the populations during each winter month. The supercooling point and lower lethal temperature remained relatively constant for the overwintering populations in the outdoor hibernaculum, ranging from −17.5 to −16.5 °C and −17.1 to −16.3 °C, respectively. In contrast, the supercooling point and lower lethal temperature of the population overwintering indoors clearly increased as the winter progressed, from −18.5 to −13.2 °C and −16.7 to −14.1 °C, respectively. A proportion of the individuals overwintering indoors could thus encounter problems surviving the winter due to premature activation at times when food is not available. The lower lethal time of field populations at 0 and −5 °C varied from 18 to 24 weeks and from 12 to 22 weeks, respectively. Morph type and sex had no influence on the cold hardiness of the overwintering adults. In addition, all cold tolerance parameters differed greatly between the laboratory population and field populations, implying that cold tolerance research based solely on laboratory populations may not be representative of field situations. We conclude from this study that the strong cold hardiness of H. axyridis in Europe may enable the species to establish in large parts of the continent.     

Entomopathogenic fungi recorded from the harlequin ladybird, Harmonia axyridis

Entomopathogenic fungi were recorded from field samples of the harlequin ladybird Harmonia axyridis, an invasive coccinellid that has recently arrived in Denmark. Larvae, pupae and adults were found to be infected by Isaria farinosa, Beauveria bassiana and species of Lecanicillium. This is the first record of entomopathogenic fungi infecting larvae and pupae. Winter mortality due to fungal infection reached 17.9% in adults collected at one location. The larval stage was most susceptible to fungal infection, as confirmed through bioassay with I. farinosa.     

Inferring the origin of populations introduced from a genetically structured native range by approximate Bayesian computation: case study of the invasive ladybird Harmonia axyridis

Correct identification of the source population of an invasive species is a prerequisite for testing hypotheses concerning the factors responsible for biological invasions. The native area of invasive species may be large, poorly known and/or genetically structured. Because the actual source population may not have been sampled, studies based on molecular markers may generate incorrect conclusions about the origin of introduced populations. In this study, we characterized the genetic structure of the invasive ladybird Harmonia axyridis in its native area using various population genetic statistics and methods. We found that native area of H. axyridis most probably consisted of two geographically distinct genetic clusters located in eastern and western Asia. We then performed approximate Bayesian computation (ABC) analyses on controlled simulated microsatellite data sets to evaluate (i) the risk of selecting incorrect introduction scenarios, including admixture between sources, when the populations of the native area are genetically structured and sampling is incomplete and (ii) the ability of ABC analysis to minimize such risks by explicitly including unsampled populations in the scenarios compared. Finally, we performed additional ABC analyses on real microsatellite data sets to retrace the origin of biocontrol and invasive populations of H. axyridis, taking into account the possibility that the structured native area may have been incompletely sampled. We found that the invasive population in eastern North America, which has served as the bridgehead for worldwide invasion by H. axyridis, was probably formed by an admixture between the eastern and western native clusters. This admixture may have facilitated adaptation of the bridgehead population.     

Interactions between the parasitoid wasp Dinocampus coccinellae and two species of coccinellid from Japan and Britain

The establishment, spread and increase of the invasive coccinellid Harmonia axyridis Pallas in North America and Britain have coincided with declines in native ladybirds. In Britain, this pattern was predicted soon after H. axyridis was first recorded. However, predictions of the impact that H. axyridis may have on the parasites, parasitoids and pathogens of native coccinellids have been less certain, largely because of uncertainty over whether H. axyridis would become an alternate host for such agents. Dinocampus coccinellae (Schrank) is a braconid parasitoid of ladybirds of the sub-family Coccinellinae. In Japan, this wasp uses both Coccinella septempunctata brucki Mulsant and H. axyridis as hosts, but successfully parasitizes a higher proportion of the former species. Data are here presented that indicate the same is true in Britain, but to a greater extent. This study investigates the interactions of D. coccinellae with C. septempunctata L. and H. axyridis from Japan and Britain. We show that coccinellid activity affects encounter rates with the wasp, with H. axyridis being more active than C. septempunctata in the tests. Escalation rates from investigation to ovipositional attack were higher for C. septempunctata than H. axyridis for both Japanese and British wasps. Wasp emergence was higher, relative to ovipositional attack, from C. septempunctata than from H. axyridis. Wasps did not discriminate between Japanese and British ladybirds of the same species. British wasp eclosion rates were higher from Japanese than from British H. axyridis. We consider both the effect D. coccinellae may have on H. axyridis population demography, and the effect H. axyridis may have on D. coccinellae in Europe.