White Butterfly Research Articles

Sex‐linked inheritance of diapause induction in the butterfly Pieris napi

AbstractMany temperate insects survive harsh environmental conditions, such as winter, by entering a state of developmental arrest. This diapause state is predominantly induced by photoperiod. The photoperiod varies with latitude and has led to local adaptation in the photoperiodic induction of diapause in many insects. To understand the rapid evolution of the photoperiodic threshold, it is important to investigate and understand the underlying genetic mechanisms. In the present study, the genetic basis of photoperiodic diapause induction is investigated in the green‐veined white butterfly Pieris napi (Lepidoptera, Pieridae) by assaying diapause induction in a range of conditions for a Swedish and Spanish population. Furthermore, the inheritance of diapause induction is assessed in reciprocal F1 hybrids and backcrosses between the two populations. The southern population shows a clear photoperiodic threshold determining diapause or direct development, whereas the northern populations show a high incidence of diapause, regardless of photoperiod. The hybrid crosses reveal that the inheritance of diapause induction is strongly sex‐linked, and that diapause incidence in the genetic crosses is highly dependent on photoperiod. This emphasizes the importance of assaying a range of conditions in diapause inheritance studies. The results indicate a strongly heritable diapause induction with a major component on the Z‐chromosome, as well as a minor effect of the autosomal background.

Fine-scale frequency differentiation along a herbivory gradient in the trichome dimorphism of a wild Arabidopsis.

Geographic variation is commonly observed in plant resistance traits, where plant species might experience different selection pressure across a heterogeneous landscape. Arabidopsis halleri subsp. gemmifera is dimorphic for trichome production, generating two morphs, trichome‐producing (hairy) and trichomeless (glabrous) plants. Trichomes of A. halleri are known to confer resistance against the white butterfly, cabbage sawfly, and brassica leaf beetle, but not against flea beetles. We combined leaf damage, microclimate, and microsatellite loci data of 26 A. halleri populations in central Japan, to explore factors responsible for fine‐scale geographic variation in the morph frequency. We found that hairy plants were less damaged than glabrous plants within populations, but the among‐site variation was the most significant source of variation in the individual‐level damage. Fixation index (Gst″) of a putative trichome locus exhibited a significant divergence along population‐level damage with an exception of an outlier population, inferring the local adaptation to herbivory. Notably, this outlier was a population wherein our previous study reported a balancing role of the brassica leaf beetle Phaedon brassicae on the morph frequency. This differentiation of the trichome locus was unrelated to neutral genetic differentiation (evaluated by Gst″ of microsatellite loci) and meteorological factors (including temperature and solar radiation). The present findings, combined with those of our previous work, provide suggestive evidence that herbivore‐driven divergence and occasional outbreak of a specific herbivore have jointly contributed to the ecogeographic pattern in the frequency of two morphs.

UV reflectance is associated with environmental conditions in Palaearctic Pieris napi (Lepidoptera: Pieridae).

The subject of our investigation was the visual features of wing color with special focus on the UV reflectance in the green-veined white butterfly (Pieris napi). Previous studies had concluded that UV reflectance on dorsal wing surfaces is found only in the female P. napi. Based on UV sensitive photography, we analyzed a correlation between 12 geographic and environmental factors and UV reflectance patterns on 3 patches on the forewings of 407 P. napi specimens from the Palaearctic region. Results had shown that females significantly differ from males: they exhibit a 25% higher UV reflectance. To investigate whether and how UV reflectance levels on the forewings and hindwings of both sexes are influenced by the environment, we performed a principal component analysis (PCA) with several environmental variables. For several variables (in particular, latitude and longitude, mean annual temperature and precipitation, and temperature annual range and altitude), the generalized linear model (GLM) model revealed a significant correlation in both sexes. This suggests a link between UV reflectance levels and the environment and distribution of P. napi. We found that stronger UV reflectance is associated with generally more hostile environments and concluded that large-scale environmental factors influence the UV reflectance on the forewings of both male and female green-veined white butterflies.

Effect of Planting Dates and Varieties on The Incidence of Cabbage Butterfly (Pieris Brassicae) in Nagaland

Field experiment conducted in 2013–2015 at the experimental cum research farm, School of Agricultural Sciences and Rural Development, Nagaland University, Medziphema campus, evaluated the effect of sowing dates and varieties against the large white butterfly (Pieris brassicae) on cabbage. The results revealed that late planting on 19th December harboured the maximum incidence, in terms of number of caterpillars/plant (8.50–2013–2014; 6.96–2014–2015, respectively). The pooled data showed a mean incidence of 7.3. Early planting on 19th November was observed with the least incidence. Among the varieties, the maximum incidence was observed in “Pride of India” (9.45–2013–2014; 7.47–2014–2015, respectively), with the pooled mean being 8.46. The lowest population was observed in “Green Express” (2.85–2013–2014; 1.78–2014–2015) and the pooled data showed an incidence of 2.32. The interaction of dates of planting and varieties was observed to be significant.

Trade-offs between fecundity and choosiness in ovipositing butterflies

Being choosy can allow animals to find and identify the best resources or safest locations to rear offspring. Despite these benefits, individuals vary in the degree to which they are choosy. One explanation is that choosiness represents a costly form of offspring investment and is part of a suite of life history trade-offs. We examined trade-offs between choosiness and fecundity in the cabbage white butterfly, Pieris rapae. To test the prediction that choosiness is negatively correlated with fecundity, we presented female cabbage white butterflies with an array of host plant leaves that varied in nutrient content and conspecific density. Butterflies preferentially laid eggs on leaves with higher nutrient content and fewer conspecific models. In addition, butterflies that were choosier with regard to plant nutrition also tended to be choosy in terms of conspecific density. Choosy females were less fecund, with fecundity measured as the sum of eggs laid over 3 days plus all remaining mature eggs in their abdomen. These results are consistent with the idea that life history trade-offs include investment in costly behavioural traits that require time and energy, such as choosiness or cognition.

Efficacy of aqueous extracts of black alder (Alnus glutinosa GAERTN.) and black elderberry (Sambucus nigra L.) in reducing the occurrence of Phyllotreta spp., some lepidopteran pests and diamondback moth parasitoids on white cabbage

AbstractThe insecticidal and antifeedant effects of aqueous extracts from two common plants (black alder Alnus glutinosa and black elderberry Sambucus nigra) on the occurrence of flea beetles (Phyllotreta spp.), as well as lepidopteran pests like the diamondback moth Plutella xylostella, small white butterfly Pieris rapae, large white butterfly Pieris brassicae, cabbage moth Mamestra brassicae and the Plutella xylostella parasitoid complex were investigated on white cabbage in 2008-2009. Aqueous extracts of black elderberry Sambucus nigra significantly reduced numbers of flea beetles Phyllotreta spp., eggs of Pieris rapae and P. brassicae, and larvae and pupae of Plutella xylostella on the cabbage. The aqueous extract of Alnus glutinosa was less effective. Parasitoids had the greatest impact on reducing numbers of diamondback moth: the dominant species was Diadegma fenestralis. The smallest number of parasitoids was reared from pupae collected from plots where the cabbages had been sprayed with the Sambucus nigra extract.

Structure and function of adipokinetic hormones of the large white butterfly Pieris brassicae

AbstractThe large white butterfly Pieris brassicae L. (also called cabbage white) is very common in Europe, Asia and the northern region of Africa, and has also been found in South Africa during approximately the last 20 years. The species is considered a pest insect, with larvae attacking brassicaceous crops. The adult is a strong migratory flyer and new territory can be infested this way. As a first step to investigate methods for combating this pest species, the present study aims to determine the complement of adipokinetic peptides, here generically referred to as adipokinetic hormones (AKHs), which are required to regulate the mobilization of fuels for insect flight. Biological assays, as well as mass spectrometry, reveal information about the presence, structure and function of AKHs in P. brassicae: a methanolic extract of the corpora cardiaca has hypertrehalosaemic activity in cockroaches, does not cause hyperlipaemia in locusts, and has adipokinetic activity in P. brassicae itself. Liquid‐chromatography electrospray ion trap mass spectrometry reveals three peptides that can be associated with the AKH family: the non‐amidated undecapeptide Vanca‐AKH (pELTFTSSWGGK‐OH), the nonapeptide Manse‐AKH (pELTFTSSWG amide) and the novel octapeptide Piebr‐AKH (pELTFSSGW amide). Sequence confirmation of all three assigned structures is obtained from matching mass spectrometry spectra from synthetic and native peptides. Moreover, the synthetic peptides Manse‐AKH and Piebr‐AKH have significant hyperlipaemic (=adipokinetic) activity when injected into newly‐emerged adult cabbage white butterflies. The non‐amidated Vanca‐AKH is, apparently, incompletely processed Manse‐AKH without hormonal activity. Simulated dispersal flight is able to release AKHs, as indicated by the higher concentration of lipids in the haemolymph of adult P. brassicae after activity and rest periods.

Differentiation in the marbled white butterfly species complex driven by multiple evolutionary forces

AbstractAimGenetic and phenotypic data may show convergent or contrasting spatial patterns. Discrepancies between markers may develop in response to different evolutionary forces. In this study we analyse inter‐ and intraspecific differentiation of closely related taxa in the marbled white butterfly species group. Based on genetic and phenotypic characters we test for potential evolutionary drivers and propose a taxonomic revision.LocationWestern Palaearctic (including north‐western Africa).MethodsWe compared distributions of mitochondrial cytochrome c oxidase subunit I gene (COI) sequences, of several allozyme loci, and of the shape of wings and genitalia obtained by applying landmark‐based techniques for the three butterfly species Melanargia galathea (central and eastern Europe), M. lachesis (Iberia) and M. lucasi (North Africa).ResultsAll studied markers showed a strong spatial structure, although discordance among their patterns was detected. COI sequences, wing shape and genitalia indicated a main split between M. galathea and M. lucasi. A lower differentiation between M. galathea and M. lachesis was found in wing shape and reflected in two mutations of the COI gene, while allozymes indicated a strong divergence. Within M. galathea, allozyme data and COI, but not morphology, revealed the existence of a slightly differentiated lineage in the Italian Peninsula, France and Switzerland. Based on COI, Melanargia lucasi was split into two subgroups, a western and an eastern Maghreb lineage.Main conclusionsLong‐term isolation of Melanargia populations between North Africa and Europe led to divergence between M. galathea and M. lucasi. This was followed by a recent differentiation among populations isolated during the cold periods of the Pleistocene, such as M. lachesis in Iberia. These lineages are characterized by a tendency not to overlap in secondary sympatry. The different patterns of the four markers may arise from divergent evolutionary processes and pressures: wings may be mainly affected by natural selection, genital structures by sexual selection, whereas long‐term isolation and drift may have driven divergence of mitochondrial DNA and allozymes.

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Developmental lead exposure has mixed effects on butterfly cognitive processes

While the effects of lead pollution have been well studied in vertebrates, it is unclear to what extent lead may negatively affect insect cognition. Lead pollution in soils can elevate lead in plant tissues, suggesting it could negatively affect neural development of insect herbivores. We used the cabbage white butterfly (Pieris rapae) as a model system to study the effect of lead pollution on insect cognitive processes, which play an important role in how insects locate and handle resources. Cabbage white butterfly larvae were reared on a 4-ppm lead diet, a concentration representative of vegetation in polluted sites; we measured eye size and performance on a foraging assay in adults. Relative to controls, lead-reared butterflies did not differ in time or ability to search for a food reward associated with a less preferred color. Indeed, lead-treated butterflies were more likely to participate in the behavioral assay itself. Lead exposure did not negatively affect survival or body size, and it actually sped up development time. The effects of lead on relative eye size varied with sex: lead tended to reduce eye size in males, but increase eye size in females. These results suggest that low levels of lead pollution may have mixed effects on butterfly vision, but only minimal impacts on performance in foraging tasks, although follow-up work is needed to test whether this result is specific to cabbage whites, which are often associated with disturbed areas.

Range, genetic diversity and future of the threatened butterfly, Pieris virginiensis

Abstract Pieris virginiensis, the West Virginia White butterfly, faces severe potential habitat loss and degradation of existing suitable habitat in the near future from climate change and plant invasion. Increasing isolation and local extinction events resulting from deforestation and climate change have a chance to significantly impact the future of this butterfly. We used genetic analysis and spatial modelling techniques to estimate the current spatial and genetic scope of P. virginiensis. We used linear modelling and machine learning to predict the occurrence of P. virginiensis through space and time using occurrence records between 1879 and 2014 and environmental predictors from a world climate dataset. We predicted the future occurrence of P. virginiensis with climate projections for the year 2070. We sequenced the cytochrome oxidase I, and the internal transcribed spacer region I, in the mitrochondrial and nuclear genomes, respectively, then used a combination of genetic diversity measures and phylogeny construction to evaluate the genetic diversity of P. virginiensis. We found that latitude accounts for nearly one‐fifth of the variation in emergence date. Spatial models predicted that P. virginiensis may lose up to 60% of available habitat in the next 50 years. Genetic data indicated some isolation in an Ohio population's COI gene, and low genetic diversity across the range of P. virginiensis in ITS. These may be indicative of either the last glacial expansion or a more recent disturbance. A large effective population size indicates that P. virginiensis are not yet nearing extinction.

De Novo Assembly and Developmental Transcriptome Analysis of the Small White Butterfly Pieris rapae.

The small white butterfly Pieris rapae is one of the most destructive pests of Brassicaceae. Yet little is understood about its genes involved in development. To facilitate research on P. rapae, we sequenced the transcriptome of P. rapae during six developmental stages, including the egg, three larval stages, the pupa, and the adult. In total, 240 million high-quality reads were obtained. De novo assembly generated 96,069 unigenes with an average length of 1353 nt. Of these, 31,629 unigenes had homologs as determined by a blastx search against the NR database with a cut-off e-value of 10−5. Clusters of Orthologous Groups of proteins (COG), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were conducted to functionally annotate those genes. Then, 849 genes involved in seven canonical development signaling pathway were identified, including dozens of key genes such as Hippo, Notch, and JAK2. A total of 21,883 differentially expressed (cut-off of 2-fold) unigenes were detected across the developmental stages, most of which were found between the egg and first larval stages. Interestingly, only 34 differentially expressed unigenes, most of which are cuticle protein related genes, were detected with a cut-off of 210-fold. Furthermore, we identified 32 heat shock protein (Hsp) genes that were expressed with complete open reading frames. Based on phylogenetic trees of the Hsp genes, we found that Hsp genes with close evolutionary relationships had similar expression pattern. Additionally, partial pattern recognition receptors genes were found to be developmental regulated. This study provides comprehensive sequence resources for P. rapae and numerous differential expressed genes, and these findings will lay the foundation for future functional genomics studies on this species.

Multiple cues influence multiple traits in the phenotypically plastic melanization of the cabbage white butterfly

Phenotypic plasticity, or the ability of organisms to produce different phenotypes depending upon environmental factors, may be adaptive in varying environments. However, because environments differ in many ways and organisms consist of many traits perfect phenotype-environment matches are unlikely. Studies that investigate multiple interacting environmental factors and the plastic responses of multiple traits should increase our understanding of the limits of adaptive plasticity. We experimentally examined the effects of variation in temperature and photoperiod on the seasonally plastic, and likely adaptive, melanization of a temperate butterfly, Pieris rapae. Although several melanin-based traits changed in response to temperature and photoperiodic variation, these traits tended to fall into two 'trait groups' consisting of traits covarying positively. However, these two trait groups responded to environmental factors, particularly temperature, in independent and sometimes opposing ways, with one increasing and the other decreasing in melanization with increased temperature. In some cases, plastic responses were complex and non-linear. Furthermore, when temperature and photoperiod were manipulated orthogonally, we sometimes detected interactive effects on melanization. These complex responses to two environmental cues may reflect sub-optimal responses or may occur if the two cues together provide more reliable information about future conditions than would either cue alone. Our results highlight the limits of studies of phenotypic plasticity that consider only single environmental factors and limit treatments to just two levels.

In vivo real-time monitoring of aphrodisiac pheromone release of small white cabbage butterflies (Pieris rapae)

The study of insect behavior is of practical importance for developing possible control methods in Integrated Pest Management. Currently, one model of butterfly mating behavior suggests that the initial location of potential mates occurs visually followed by the release of one or more short-range male aphrodisiac pheromones. This model is supported by data obtained from field observations and inferences based on the behavioral effects of chemicals extracted or isolated using indirect and offline techniques. In this study, we performed in vivo real-time monitoring of the male aphrodisiac pheromones released by the small white cabbage male butterfly (Pieris rapae Linnaeus) using confined direct analysis in real time (cDART) mass spectrometry. cDART is a new method easily adapted to the study in real time of chemicals released into the environment by virtually any insect. The major compound released by the male Pieris rapae was identified as ferrulactone. The experimental results reported here indicate that the release of ferrulactone occurs less than 1s after the male visualizes its partner, and reaches a maximum after about one half minute. This study is the first reported in vivo detection and monitoring of butterfly male aphrodisiac pheromones in real time.

Peierls substitution for magnetic Bloch bands

We consider the Schr\"odinger operator in two dimensions with a periodic potential and a strong constant magnetic field perturbed by slowly varying non-periodic scalar and vector potentials, $\phi(\epsilon x)$ and $A(\epsilon x)$, for $\epsilon\ll 1$. For each isolated family of magnetic Bloch bands we derive an effective Hamiltonian that is unitarily equivalent to the restriction of the Schr\"odinger operator to a corresponding almost invariant subspace. At leading order, our effective Hamiltonian can be interpreted as the Peierls substitution Hamiltonian widely used in physics for non-magnetic Bloch bands. However, while for non-magnetic Bloch bands the corresponding result is well understood, for magnetic Bloch bands it is not clear how to even define a Peierls substitution Hamiltonian beyond a formal expression. The source of the difficulty is a topological obstruction: magnetic Bloch bundles are generically not trivializable. As a consequence, Peierls substitution Hamiltonians for magnetic Bloch bands turn out to be pseudodifferential operators acting on sections of non-trivial vector bundles over a two-torus, the reduced Brillouin zone. Part of our contribution is the construction of a suitable Weyl calculus for such pseudos. As an application of our results we construct a new family of canonical one-band Hamiltonians $H^B_{\theta,q}$ for magnetic Bloch bands with Chern number $\theta\in \mathbb{Z}$ that generalizes the Hofstadter model $H^B_{\rm Hof} = H^B_{0,1}$ for a single non-magnetic Bloch band. It turns out that $H^B_{\theta,q}$ is isospectral to $H^{q^2B}_{\rm Hof}$ for any $\theta$ and all spectra agree with the Hofstadter spectrum depicted in his famous black and white butterfly. However, the resulting Chern numbers of subbands, corresponding to Hall conductivities, depend on $\theta$ and $q$, and thus the models lead to different colored butterflies.

Effect ofSolenopsis invicta(Hymenoptera: Formicidae) on Flower-Visiting Behavior of Insects onBrassica napus(Brassicales: Brassicaceae)

Abstract This study used field trials to investigate how the fire ant Solenopsis invicta Buren (Hymenoptera: Formicidae) affects the flower-visiting behavior of insects on rapeseed Brassica napus L. (Brassicales: Brassicaceae). Chemicals from fire ants were identified, and their electrophysiological effects on the cabbage white butterfly Pieris rapae (L.) (Lepidoptera: Pieridae) were analyzed by gas chromatography—electroantennogram detection (GC-EAD) bioassays. The results showed that S. invicta significantly reduced the abundance of pollinators. The number of striped flea beetles (Phyllotreta striolata [F.]; Coleoptera: Chrysomelidae) and hover flies (Eristalinus quinquestriatus [F.] and E. quinquelineatus [F.]; Diptera: Syrphidae) in fire ant—included and fire-ant-and-aphid-included plots was 43.5 and 39.4% lower, respectively, than in control plots. The visiting duration at a single flower for P. rapae in the plots that had fire ants and aphids excluded was significantly longer than in the fire ant—in...

Assessing the Role of Wing Spots in Intraspecific Communication in the Cabbage White Butterfly (Pieris rapae L.) Using a Simple Device to Increase Butterfly Responses

Butterflies are regularly used as model systems for understanding the role of coloration in communication because of their highly variable and conspicuous phenotypes. Most research showing a role for color in communication has focused on aspects of brightness or hue of entire wings or large color patches. However, evidence is accumulating that butterflies sometimes use smaller wing pattern elements in communication. Here we provide evidence that both male and female cabbage white butterflies (Pieris rapae L.) discriminate among conspecifics on the basis of the number of small but conspicuous black wing spots of the dorsal forewing. Male butterflies were more interactive with model butterflies with two spots, which resemble female butterflies, than with model butterflies with only one spot, which resemble male butterflies. Female butterflies showed the opposite response, being more interactive with male-like (one-spot) models than with female-like (two-spot) models. Some of our experiments were conducted with an electronic device designed to create a realistic and controlled fluttering behavior of the models. We describe the design and function of this device and provide evidence that it increased butterfly responses compared to a non-fluttering model. This device could prove useful for others addressing questions of communication in butterflies or other flying insects.

Potential of Ethiopian mustard, Brassica carinata as a trap crop for large white butterfly, Pieris brassicae infesting Indian mustard, Brassica juncea

The large white butterfly, Pieris brassicae (L.), is an important pest of Indian mustard, Brassica juncea (L.) Czern., and inflicts heavy damage to all the above ground plant parts with strong yield reducing impacts. Farmers have few practical options other than to spray insecticides to protect their crop. In this study, an attempt was made during 2012–2013 and 2013–2014 crop seasons at Ludhiana, India, to evaluate Ethiopian mustard, Brassica carinata A. Braun as a trap crop to manage this pest as an alternate pest management strategy. B. carinata borders surrounding B. juncea were compared to chemically protected B. juncea and control (without B. carinata borders) plots for their effects on P. brassicae infestation and grain yield. Oviposition preference of adult butterflies was studied in a two-choice test in field cages, while the effect of two host plants on larval performance was studied under laboratory conditions under no choice conditions. B. juncea plots bordered with B. carinata harbored significantly lower larval population compared to control B. juncea plots during both the seasons. The grain yield in the bordered plots was also significantly higher than that from control plots and was statistically non-significant from that obtained in chemically protected plots. Female butterflies showed distinct oviposition preference for B. carinata over B. juncea and the larvae reared on B. carinata completed development in shorter period and grew bigger and heavier than those reared on B. juncea. Results indicated that B. carinata has potential to be used as trap crop to manage P. brassicae.

Black and Garlic Mustard Plants Are Highly Suitable for the Development of Two Native Pierid Butterflies.

In multivoltine insects that oviposit and develop on short-lived plants, different herbivore generations across a growing season often exploit different plant species. Here, we compare the development time, pupal mass, and survival of two closely related oligophagous herbivore species on two species of brassicaceous plants that grow in different habitats and which exhibit little overlap in temporal growth phenology. In central Europe, the green-veined white butterfly, Pieris napi L., is bivoltine, whereas the small cabbage white butterfly, Pieris rapae L., has two to three generations a year. Moreover, P. napi is primarily found in moist, open (e.g., meadow), and forest habitats, whereas P. rapae prefers drier, open habitats. Both butterflies were reared on Garlic mustard (Alliaria petiolata), which is shade-tolerant and grows early in spring in forest undergrowth, and Black mustard (Brassica nigra), which prefers open disturbed habitats and is most common in summer. Both host plant species differ in other traits such as secondary chemistry. We hypothesized that, owing to habitat preference, P. napi would develop equally well on both plants but that P. rapae would perform better on B. nigra. The results provide partial support for this hypothesis, as both herbivores performed equally well on A. petiolata and B. nigra. However, there were differences in these parameters that were species-specific: on both plants P. rapae developed faster and had larger pupae than P. napi. Our results show that specialized herbivores can exploit different species of related plants that grow at different times of the season, enabling them to have multiple generations.

Characterization and cytotoxic activity of apoptosis-inducing pierisin-5 protein from white cabbage butterfly

In this study, caspase-dependent apoptosis-inducing pierisin-5 gene was identified and characterized from cabbage white butterfly, Pieris canidia. A thousand-fold increase in expression of pierisin-5 gene was observed from second to third instar larvae, gradually decreasing before pupation. Pierisin-5 was purified from the fifth-instar larvae and was found to exhibit cytotoxicity against HeLa and HepG2 human cancer cell lines. Pierisin-5 showed growth inhibition and several morphological changes such as cell shrinkage, chromatin condensation and apoptotic body formation with programmed cell death in HeLa and HepG2 cells. Moreover, DNA fragmentation was observed after gel electrophoresis analysis. Caspase substrate assay showed further cleavage of Ac-DEVD-pNA, suggesting the activation of Caspase-3. Flow cytometry analysis revealed the cell cycle arrest at G1 phase and increased the percentage of apoptotic cells in cancer cell lines treated with pierisin-5. These findings suggest that pierisin-5 could significantly induce apoptosis in cancer cell lines and is mediated by activation of caspase-3 in the mitochondrial pathway. Phylogenetic analysis using pierisin proteins from Pierid butterflies, ADP-ribosylating toxins from bacteria, human, rat, and mouse indicated the possibility of horizontal transfer of pierisin genes from bacteria to butterflies. The single copy of pierisin gene unlike other insect toxin genes also supports lateral transfer.