Mimetic Females Research Articles

Phenotypic Plasticity of the Mimetic Swallowtail Butterfly Papilio polytes: Color Pattern Modifications and Their Implications in Mimicry Evolution.

Butterfly wing color patterns are sensitive to environmental stress, such as temperature shock, and this phenotypic plasticity plays an important role in color pattern evolution. However, the potential contributions of phenotypic plasticity to mimicry evolution have not been evaluated. Here, we focused on the swallowtail butterfly Papilio polytes, which has nonmimetic and mimetic forms in females, to examine its plastic phenotypes. In the nonmimetic form, medial white spots and submarginal reddish spots in the ventral hindwings were enlarged by cold shock but were mostly reduced in size by heat shock. These temperature-shock-induced color pattern modifications were partly similar to mimetic color patterns, and nonmimetic females were more sensitive than males and mimetic females. Unexpectedly, injection of tungstate, a known modification inducer in nymphalid and lycaenid butterflies, did not induce any modification, but fluorescent brightener 28, another inducer discovered recently, induced unique modifications. These results suggest that phenotypic plasticity in nonmimetic females might have provided a basis of natural selection for mimetic color patterns during evolution.

Doublesex Controls Both Hindwing and Abdominal Mimicry Traits in the Female-Limited Batesian Mimicry of Papilio memnon.

Papilio butterflies are known to possess female-limited Batesian mimicry polymorphisms. In Papilio memnon, females have mimetic and non-mimetic forms, whereas males are monomorphic and non-mimetic. Mimetic females are characterized by color patterns and tails in the hindwing and yellow abdomens. Recently, an analysis of whole-genome sequences has shown that an approximately 160 kb region of chromosome 25 is responsible for mimicry and has high diversity between mimetic (A) and non-mimetic (a) alleles (highly diversified region: HDR). The HDR includes three genes, UXT, doublesex (dsx), and Nach-like, but the functions of these genes are unknown. Here, we investigated the function of dsx, a gene involved in sexual differentiation, which is expected to be functionally important for hindwing and abdominal mimetic traits in P. memnon. Expression analysis by reverse transcription quantitative PCR (RT-qPCR) and RNA sequencing showed that mimetic dsx (dsx-A) was highly expressed in the hindwings in the early pupal stage. In the abdomen, both dsx-A and dsx-a were highly expressed during the early pupal stage. When dsx was knocked down using small interfering RNAs (siRNAs) designed in the common region of dsx-A and dsx-a, a male-like pattern appeared on the hindwings of mimetic and non-mimetic females. Similarly, when dsx was knocked down in the abdomen, the yellow scales characteristic of mimetic females changed to black. Furthermore, when dsx-a was specifically knocked down, the color pattern of the hindwings changed, as in the case of dsx knockdown in non-mimetic females but not mimetic females. These results suggest that dsx-a is involved in color pattern formation on the hindwings of non-mimetic females, whereas dsx-A is involved in hindwing and abdominal mimetic traits. dsx was involved in abdominal and hindwing mimetic traits, but dsx expression patterns in the hindwing and abdomen were different, suggesting that different regulatory mechanisms may exist. Our study is the first to show that the same gene (dsx) regulates both the hindwing and abdominal mimetic traits. This is the first functional analysis of abdominal mimicry in butterflies.

Chemical Identity of Cuticular Lipid Components in the Mimetic Swallowtail Butterfly Papilio polytes.

The swallowtail Papilio polytes shows Batesian and female-limited polymorphic mimicry. In Japan, P. polytes females have two different forms: the cyrus form is non-mimetic and resembles males, whereas the polytes form mimics Pachliopta aristolochiae and Byasa (Atrophaneura) alcinous as unpalatable models. During mating, P. polytes males use cuticular lipids to distinguish non-mimetic females from conspecific males and sympatric sister species. In this study, we investigated whether compositional differences in cuticular lipids exist between mimetic and non-mimetic females of P. polytes and between mimetic females and their model species. The mimetic and non-mimetic females had nearly identical cuticular lipid profiles, which differed from those of males. The two model species exhibited sexually dimorphic and species-specific cuticular lipid compositions, which were distinctly different from those of mimetic P. polytes females. These results strongly suggest that P. polytes females maintain the identity of cuticular lipid profiles regardless of the mimicry type, and this feature helps males recognize mimetic females as the correct mating partners.

Batesian mimicry has evolved with deleterious effects of the pleiotropic gene doublesex

Dimorphic female-limited Batesian mimicry in the swallowtail butterfly Papilio polytes is regulated by the supergene locus H, harbouring the mimetic (H) and non-mimetic (h) doublesex (dsx) gene. In the present study, we demonstrated that dsx-H negatively affects the number of eggs laid, hatching rate, larval survival rate, and adult lifespan. When crossed with hh males, the number of eggs laid of mimetic females (genotype HH) was lower than that of non-mimetic females (hh). Moreover, hh and Hh females laid fewer eggs when crossed with HH males. The hatching and larval survival rates were lower when both female and male parents harboured dsx-H. The adult lifespan of HH females was shorter than that of hh females, while it was similar in males regardless of the genotype. These findings suggest the presence of a cost–benefit balance of Batesian mimicry, which is evolved to avoid predation but is accompanied by physiological deficits, in this species.

Mimicry genes reduce pre-adult survival rate in Papilio polytes: A possible new mechanism for maintaining female-limited polymorphism in Batesian mimicry.

Batesian mimicry, in which harmless organisms resemble unpalatable or harmful species, is a well-studied adaptation for predation avoidance. The females of some Batesian mimic species comprise mimetic and nonmimetic individuals. Mimetic females of such polymorphic species clearly have a selective advantage due to decreased predation pressure, but the selective forces that maintain nonmimetic females in a population remain unclear. In the swallowtail butterfly, Papilio polytes, female polymorphism is controlled by the H (mimetic) and h (nonmimetic) alleles at a single autosomal locus. Here, we examined whether the dominant H allele has a deleterious effect on the pre-adult survival rate (egg-to-adult emergence rate). We repeated an assortative mating-like treatment-that is breeding of males and females whose mothers had the same phenotype (mimetic or nonmimetic)-for three consecutive generations, while avoiding inbreeding. Results showed that pre-adult survival rate decreased over generations only in lines derived from mothers with the mimetic phenotype (hereafter, mimetic-assorted lines). This lowered survival was due to an increased mortality at the final instar larval stage and the pupal stages. Interestingly, the pre-adult mortality in the mimetic-assorted lines seemed to be associated with a male-biased sex ratio at adult emergence. These results suggest that the dominant H allele displays a mildly deleterious effect that is expressed more strongly in females and homozygous individuals than in heterozygous individuals. We propose that this cost of mimicry in larval and pupal stages contributes to the maintenance of female-limited polymorphism in P.polytes.

Behaviour before beauty: signal weighting during mate selection in the butterfly Papilio polytes.

Mating displays often contain multiple signals. Different combinations of these signals may be equally successful at attracting a mate, as environment and signal combination may influence relative signal weighting by choosy individuals. This variation in signal weighting among choosy individuals may facilitate the maintenance of polymorphic displays and signalling behaviour. One group of animals known for their polymorphic patterning are Batesian mimetic butterflies, where the interaction of sexual selection and predation pressures are hypothesized to influence the maintenance of polymorphic wing patterning and behaviour. Males in the female-limited polymorphic Batesian mimetic butterfly Papilio polytes use female wing pattern and female activity levels when determining whom to court. They court stationary females with mimetic wing patterns more often than stationary females with non-mimetic, male-like wing patterns, and active females more often than inactive females. It is unclear whether females modify their behaviour to increase (or decrease) their likelihood of receiving male courtship, or whether non-mimetic females spend more time in cryptic environments than mimetic females, to compensate for their lack of mimicry-driven predation protection (at the cost of decreased visibility to males). In addition, relative signal weighting of female wing pattern and activity to male mate selection is unknown. To address these questions, we conducted a series of observational studies of a polymorphic P. polytes population in a large butterfly enclosure. We found that males exclusively courted active females, irrespective of female wing pattern. However, males did court active non-mimetic females significantly more often than expected given their relative abundance in the population. Females exhibited similar activity levels, and selected similar resting environments, irrespective of wing pattern. Our results suggest that male preference for non-mimetic females may play an active role in the maintenance of the non-mimetic female form in natural populations, where males are likely to be in the presence of active, as well as inactive, mimetic and non-mimetic females.

Assessing ecological and physiological costs of melanism in North American Papilio glaucus females: two decades of dark morph frequency declines.

Polymorphisms for melanic form of insects may provide various selective advantages. However, melanic alleles may have significant/subtle pleiotrophic “costs.” Several potential pleiotrophic effects of the W (=Y)‐linked melanism gene in Papilio glaucus L. (Lepidoptera) showed no costs for melanic versus yellow in adult size, oviposition preferences, fecundity, egg viability, larval survival/growth rates, cold stress tolerance, or postdiapause emergence times. Sexual selection (males choosing yellow rather than mimetic dark females) had been suggested to provide a balanced polymorphism in P. glaucus, but spermatophore counts in wild females and direct field tethering studies of size‐matched pairs of virgin females (dark and yellow), show that male preferences are random or frequency‐dependent from Florida to Michigan, providing no yellow counter‐advantages. Recent frequency declines of dark (melanic/mimetic) females in P. glaucus populations are shown in several major populations from Florida (27.3°N latitude) to Ohio (38.5° N). Summer temperatures have increased significantly at all these locations during this time (1999–2018), but whether dark morphs may be more vulnerable (in any stage) to such climate warming remains to be determined. Additional potential reasons for the frequency declines in mimetic females are discussed: (i) genetic introgression of Z‐linked melanism suppressor genes from P. canadensis (R & J) and the hybrid species, P. appalachiensis (Pavulaan & Wright), (ii) differential developmental incompatibilities, or Haldane effects, known to occur in hybrids, (iii) selection against intermediately melanic (“dusty”) females (with the W‐linked melanic gene, b+) which higher temperatures can cause.

Does male preference play a role in maintaining female limited polymorphism in a Batesian mimetic butterfly?

Female-limited polymorphism occurs in multiple butterfly species with Batesian mimicry. While frequency-dependent selection is often argued as the driving force behind polymorphism in Batesian mimicry systems, male preference and alternative female mating strategies may also influence the maintenance of multiple female forms. Through a series of behavioural assays with the female-limited Batesian mimetic butterfly Papilio polytes, we show that males prefer stationary mimetic females over stationary non-mimetic females, but weigh female activity levels more heavily than female wing pattern when choosing between active mimetic and active non-mimetic females. Male preference for mimetic vs. non-mimetic females is independent of male genotype at the locus responsible for the female wing pattern, the autosomal gene doublesex. However male genotype does influence their response to active females. Male emphasis on female behaviour instead of appearance may reduce sexual selection pressures on female morphology, thereby facilitating frequency-dependent natural selection due to predation risk and toxic model abundance.

Behavioural mimicry in flight path of Batesian intraspecific polymorphic butterfly Papilio polytes.

Batesian mimics that show similar coloration to unpalatable models gain a fitness advantage of reduced predation. Beyond physical similarity, mimics often exhibit behaviour similar to their models, further enhancing their protection against predation by mimicking not only the model's physical appearance but also activity. In butterflies, there is a strong correlation between palatability and flight velocity, but there is only weak correlation between palatability and flight path. Little is known about how Batesian mimics fly. Here, we explored the flight behaviour of four butterfly species/morphs: unpalatable model Pachliopta aristolochiae, mimetic and non-mimetic females of female-limited mimic Papilio polytes, and palatable control Papilio xuthus. We demonstrated that the directional change (DC) generated by wingbeats and the standard deviation of directional change (SDDC) of mimetic females and their models were smaller than those of non-mimetic females and palatable controls. Furthermore, we found no significant difference in flight velocity among all species/morphs. By showing that DC and SDDC of mimetic females resemble those of models, we provide the first evidence for the existence of behavioural mimicry in flight path by a Batesian mimic butterfly.

Molecular basis of wing coloration in a Batesian mimic butterfly, Papilio polytes

Batesian mimicry protects animals from predators through resemblance with distasteful models in shape, color pattern, or behavior. To elucidate the wing coloration mechanisms involved in the mimicry, we investigated chemical composition and gene expression of the pale yellow and red pigments of a swallowtail butterfly, Papilio polytes, whose females mimic the unpalatable butterfly Pachliopta aristolochiae. Using LC/MS, we showed that the pale yellow wing regions in non-mimetic females consist of kynurenine and N-β-alanyldopamine (NBAD). Moreover, qRT-PCR showed that kynurenine/NBAD biosynthetic genes were upregulated in these regions in non-mimetic females. However, these pigments were absent in mimetic females. RNA-sequencing showed that kynurenine/NBAD synthesis and Toll signaling genes were upregulated in the red spots specific to mimetic female wings. These results demonstrated that drastic changes in gene networks in the red and pale yellow regions can switch wing color patterns between non-mimetic and mimetic females of P. polytes.

Behavioral Batesian Mimicry Involving lntraspecific Polymorphism in the ButterflyPapilio polytes

Batesian mimics gain protection from predation by their similarity to distasteful models. In butterflies, it has been thought that distasteful species and Batesian mimics fly slowly and in a straight line, but few studies have demonstrated their behavioral similarity, and no studies have been conducted on behavioral mimicry Involving Batesian intraspecific polymorphism. Here, we compared the wing stroke among various butterflies: palatable non-mimetic Papilio xuthus, unpalatable Pachliopta aristolochiae, and palatable polymorphic Papilio polytes (cyrus form, non-mimetic females; polytes form, Batesian mimetic females) to clarify whether the wing stroke of unpalatable butterflies is different from that of palatable species, whether that of the non-mimetic females of Pap. polytes is different from the mimetic females, and whether that of the mimetic females resembles that of the model. We found that the minimum positional angle (phi(min)) of Pach. aristolochiae and mimetic females of Pap. polytes was significantly larger than that of Pap. xuthus and non-mimetic females. We did not detect significant differences between that of Pach. aristolochiae and mimetic females of Pap. polytes. These results show that phi(min) differed between the mimicry group and palatable butterflies. In addition, the wingbeat frequency (WBF) of Pach. aristolochiae and mimetic females tended to differ from that of Pap. xuthus and non-mimetic females. This result suggests that there may be convergence of WBF in Batesian mimicry groups, as in the case of Müllerian mimicry groups, and serves as the first evidence of behavioral mimicry in Batesian intraspecific polymorphism.

A common mechanism explaining the evolution of female‐limited and both‐sex Batesian mimicry in butterflies

Summary Batesian mimicry describes the situation in which a palatable mimic resembles an unpalatable model. In some species of butterflies, both sexes mimic, but in others only females do. Two mechanisms have been proposed to generate female‐limited mimicry: sexual selection via female choice, and sexual selection via male–male competition. Each is not satisfactory because of too many exceptions. I hypothesized that reductions in physiological life span because of mimicry constituted the costs, while extensions of ecological life span because of mimicry constituted the benefits. Mimicry would result from the balance of a costs/benefits relationship; when balances are favourable, mimicry occurs. Non‐mimetic females of female‐limited mimicry butterfly Papilio polytes lived longer than its mimetic females in a butterfly farm greenhouse. Therefore, reduction in physiological life span must be a cost of mimicry. Male‐biased sex ratios were found in 11 of the 14 non‐mimetic and non‐model species netted in the Kakamega tropical rainforest in west Kenya, in other one species almost equivalent, and in remaining two species female‐biased. For each species, the sexes netted less frequently had more beak marks than those netted more frequently. That is, the biased sex ratios could be responsible for the higher predation rates of the opposite sex. Predators may have selectively attacked females with wider thoraxes, which flew at higher levels and flew more quickly. However, attack rates on males of some species may be high or low regardless of thorax width. As the benefits accrue largely to females with wider thorax due to female‐biased predation, balances of costs/benefits relationships divide participating species into three theoretical groups: those in which the adaptation is favourable for both sexes; favourable only for females; and unfavourable for both sexes. In female‐limited mimics of five species, thorax widths varied, whereas thoraxes tended to be wider in both‐sex mimics of four species.

GENETICS OF MIMICRY IN THE TIGER SWALLOWTAIL BUTTERFLIES, PAPILIO GLAUCUS AND P. CANADENSIS (LEPIDOPTERA: PAPILIONIDAE).

The tiger swallowtail butterfly, Papilio glaucus, exhibits a female-limited polymorphism for Batesian mimicry; the Canadian tiger swallowtail, Papilio canadensis, lacks the mimetic (dark) form entirely. The species hybridize to a limited extent where their ranges overlap. Field collections and censuses indicate that mimetic females occur throughout the range of P. glaucus but at lowest frequencies in populations at the latitudinal edges of its geographic range such as the southernmost part of Florida and along the entire northern edge of its distribution from Massachusetts to Minnesota. Frequencies of mimetic females have remained relatively stable over time. Inheritance of the mimetic form is controlled primarily by two interacting sex-linked loci. The typical matrilineal pattern of inheritance in P. glaucus can be explained by polymorphism at a Y-linked locus, b. Analysis of P. glaucus × P. canadensis crosses has also revealed an X-linked locus, s, which controls the expression of the mimetic phenotype. The P. canadensis allele, scan , suppresses the mimetic phenotype in hybrid and backcross females. Results from more than 12 yr of rearing tiger swallowtails, including interspecies hybrids, indicate that the absence of mimetic P. canadensis females is due to both a high frequency of the "suppressing" allele scan and low frequency of the black-pigment-determining b + allele. The frequency of scan (or other suppressing alleles of s) in P. glaucus populations outside the hybrid zone is low. Some males heterozygous at the s locus and some suppressed mimetic females occur within the hybrid zone. A simple genetic model predicts the frequency of daughters that differ in phenotype from their mothers.

Preferential predation of female butterflies and the evolution of batesian mimicry

BATESIAN mimicry, in which a palatable mimic resembles an unpalatable model, functions to protect insect mimics from birds. In butterflies that show batesian mimicry, female-limited mimicry is common1–3. The orthodox theory to explain this is sexual selection against males4–6. In these theoretical arguments, no difference in predation pressure between the sexes was assumed, but the existence of female-biased predation would enhance the evolution of sex-limited mimicry. To test for differences in attack rate between the sexes, I examined the rates of beak marks on wings of palatable butterflies of Papilionidae and Pieridae, and unpalatable Danaidae. Here I report that females were attacked more frequently than males, though danaids were generally attacked less. The papilionid and pierid males had low attack rates similar to those of danaid females. Analysis of a mathematical model highlighted these tendencies. Comparing a batesian mimetic species and its 'model' species, non-mimetic females were selectively attacked and the males, mimetic females and 'models' were attacked less. Therefore females benefit greatly when they become mimetic, whereas males will benefit much less should they become mimetic. Thus female-limited mimicry will be favoured even if the costs of mimicry to both sexes are the same.

The genetics of the mimetic butterfly Papilio polytes L.

Papilio polytesL. is a mimetic Swallowtail butterfly widely distributed in South East Asia. It has four female forms, three mimetic, and one non-mimetic resembling the monomorphic male in appearance. The various female forms are now shown to be controlled by allelomorphs at a single autosomal locus and not by independent genes as previously thought. The effects of the allelomorphs controlling the mimetic patterns are sex-limited to the female. There is some evidence that the locus is a complex one consisting of two or more tightly linked genes. As in previous investigations into mimicry in Swallowtails the dominance tends to be complete between sympatric forms. The accuracy of the mimicry depends not only on the presence of the appropriate major genes but also on the rest of the gene complex. Thus within a race there is an integrated genetic system and on outcrossing this becomes disturbed, leading to poorer mimicry. The system of modifiers controlling the accuracy of the mimetic pattern is closely paralleled by that inP. dardanus. In particular, inP. polytesf.theseusappears to differ from f.polytesonly as a result of the presence of a modifier system, as does f.hippocoonfrom f.hippocoonidesinP. dardanus. InP. dardanusfrom Ethiopia it was found that specific modifiers adjusting the tail length of mimetic females have been selected for, thus improving the mimicry. An analogous situation has been found inP. polytesalthough here the control is more effective in that the resulting difference in tail length between the mimetic and non-mimetic forms can be as much as 10 mm, whereas inP. dardanusit rarely exceeds 3 mm. The great similarity in the genetic structure ofP. polytesandP. dardanus(as well asP. memnon) strongly suggests that selection for a mimetic polymorphism results in the evolution of very similar genetic control mechanisms in different species - that is to say it is the nature of the selection rather than the species involved which determines the genetic architecture.