Dynamics of genomic change during evolutionary rescue in the seed beetle Callosobruchus maculatus

Rapid adaptation can prevent extinction when populations are exposed to extremely marginal or stressful environments. Factors that affect the likelihood of evolutionary rescue from extinction have been identified, but much less is known about the evolutionary dynamics (e.g., rates and patterns of allele frequency change) and genomic basis of successful rescue, particularly in multicellular organisms. We conducted an evolve‐and‐resequence experiment to investigate the dynamics of evolutionary rescue at the genetic level in the cowpea seed beetle, Callosobruchus maculatus, when it is experimentally shifted to a stressful host plant, lentil. Low survival (~1%) at the onset of the experiment caused population decline. But adaptive evolution quickly rescued the population, with survival rates climbing to 69% by the F5 generation and 90% by the F10 generation. Population genomic data showed that rescue likely was caused by rapid evolutionary change at multiple loci, with many alleles fixing or nearly fixing within five generations of selection on lentil. Selection on these loci was only moderately consistent in time, but parallel evolutionary changes were evident in sublines formed after the lentil line had passed through a bottleneck. By comparing estimates of selection and genomic change on lentil across five independent C. maculatus lines (the new lentil‐adapted line, three long‐established lines and one case of failed evolutionary rescue), we found that adaptation on lentil occurred via somewhat idiosyncratic evolutionary changes. Overall, our results suggest that evolutionary rescue in this system can be caused by very strong selection on multiple loci driving rapid and pronounced genomic change.     

Consequences of inbreeding for the cowpea seed beetle, Callosobruchus maculatus (F.)(Coleoptera: Bruchidae)

Inbreeding is said to reduce vigour and fitness. It may also determine how a population responds to selection. Local populations of Callosobruchus maculatus , the cowpea seed beetle, are established annually from small numbers of founders and the species has been distributed to many parts of the world where isolated populations may have been founded by very small numbers of individuals. After more than 20 generations of inbreeding, inbred lines have been shown to diverge from a common ancestral stock in similar directions with respect of some variables such as developmental speed, but haphazardly in respect of other parameters such as male weight. The respective roles of drift and of selection as effective evolutionary forces in inbred lines are discussed in the light of these results. It is argued that some intraspecific differences in C. maculatus may be explained as a product of periodic inbreeding, but that the process does not impair the ability to adapt to local conditions so contributing to the status of the species as a pest of international importance.     

Intra-specific variation in the morphology and the benefit of large genital sclerites of males in the adzuki bean beetle (Callosobruchus chinensis)

Rapid evolution has led to a large diversity in the sizes and morphology of male genitals across taxa, but the mechanisms driving this evolution remain controversial. In this study, we investigated the function of male genital sclerites in the adzuki bean beetle (Callosobruchus chinensis) and compared the length and morphology of genital sclerites between two populations that vary in their degree of polyandry. We found that the length of male genital sclerites was negatively correlated with copulation duration but positively correlated with the speed of matings with multiple females. Additionally, we found that the average length and number of genital sclerite spines of males from the more polyandrous population were larger than those from the less polyandrous population. We suggest that the genital sclerite of male adzuki bean beetles evolved by sexual selection, and a larger genital sclerite has a selective advantage because it allows for rapid copulations with multiple females.     

Gaseous ozone: A potent pest management strategy to control Callosobruchus maculatus (Coleoptera: Bruchidae) infesting green gram

Extending the storage life of legumes by protecting it from the Callosobruchus maculatus infestation is a major concern for the producers, processors and exporters. Legume processing industry requires “greener” alternatives to the conventional fumigants. Gaseous ozone has a great potential as an insect management strategy that is suited for this niche. Nevertheless, the efficacy of ozone against C. maculatus is yet unknown. A laboratory study was conducted to test the insecticidal effect of ozone in controlling the infestation of C. maculatus in green gram. We have determined the concentration of ozone exposure time–mortality relationship for all the stages of C. maculatus that were exposed to 500–1,500 ppmv ozone. The percentages of mortality for different stages of C. maculatus increased with the increase in ozone concentration and exposure time. It was documented that adult stage is least tolerant to ozone (500 ppmv for 274.40 min exposure required to kill 90%), whereas the most tolerant stage is pupa (500 ppmv for 1816.54 min is required to kill 90%). The results indicate that gaseous ozone is the attractive alternative to the synthetic fumigants.     

Individual and synergistic effects of male external genital traits in sexual selection

Male genital traits exhibit extraordinary interspecific phenotypic variation. This remarkable and general evolutionary trend is widely considered to be the result of sexual selection. However, we still do not have a good understanding of whether or how individual genital traits function in different competitive arenas (episodes of sexual selection), or how different genital traits may interact to influence competitive outcomes. Here, we use an experimental approach based on high‐precision laser phenotypic engineering to address these outstanding questions, focusing on three distinct sets of micron‐scale external (nonintromittent) genital spines in male Drosophila kikkawai Burla (Diptera: Drosophilidae). Elimination of the large pair of spines on the male secondary claspers sharply reduced male ability to copulate, yet elimination of the other sets of spines on the primary and secondary claspers had no significant effects on copulation probability. Intriguingly, both the large spines on the secondary claspers and the cluster of spines on the primary claspers were found to independently promote male competitive fertilization success. Moreover, when large and small secondary clasper spines were simultaneously shortened in individual males, these males suffered greater reductions in fertilization success relative to males whose traits were altered individually, providing evidence for synergistic effects of external genital traits on fertilization success. Overall, the results are significant in demonstrating that a given genital trait (the large spines on the secondary claspers) can function in different episodes of sexual selection, and distinct genital traits may interact in sexual selection. The results offer an important contribution to evolutionary biology by demonstrating an understudied selective mechanism, operating via subtle trait interactions in a post‐insemination context, by which genital traits may be co‐evolving.     

The genetic architecture of sexual conflict: male harm and female resistance in Callosobruchus maculatus

Males harm females during mating in a range of species. This harm is thought to evolve because it is directly or indirectly beneficial to the male, despite being costly to his mate. The resulting sexually antagonistic selection can cause sexual arms races. For sexually antagonistic co-evolution to occur, there must be genetic variation for traits involved in female harming and susceptibility to harm, but even then intersexual genetic correlations could facilitate or impede sexual co-evolution. Male Callosobruchus maculatus harm their mates during copulation by damaging the female's reproductive tract. However, there have been no investigations of the genetic variation in damage or in female susceptibility to damage, nor has the genetic covariance between these characters been assessed. Here, we use a full-sib/half-sib breeding design to show that male damage is heritable, whereas female susceptibility to damage is much less so. There is also a substantial positive genetic correlation between the two, suggesting that selection favouring damaging males will increase the prevalence of susceptible females. We also provide evidence consistent with intralocus sexual conflict in this species.     

Comparative seed resistance in eighty cowpea accessions to the seed bruchid,Callosobruchus maculatus (Fabricius) (Coleoptera: Bruchidae)

Eighty cowpea accessions comprising both improved and local were assessed for their resistance to Callosobruchus maculatus (F.) attack under laboratory conditions (29 ± 3?°C and 65 ± 5% R.H.). Sixty-nine accessions were derived from the International Institute for Tropical Agriculture breeding programmes, four were local cultivars, while seven were improved cultivars from national research institutes. Results showed that there were significant differences among the accessions in terms of number of initial bruchid eggs laid on them, adult emergence, number of seeds damaged and bruchid-induced seed weight loss at three months after artificial infestation with C. maculatus. On IT04K-334-2, IT04K-343-1, IT06K-141, IT99K-216-48-1, IT99K-494-6 and IT99K-529-2, fewer initial eggs were laid and no adult emergence, seed damage or weight loss and were considered highly resistant to C. maculatus. On the other hand, IT03K-337, IT04K-217-51, IT04K-332-1, IT04K-339-1, IT06K-188-49, IT81D-1137, IT845-2231-15, IT93K-452-1, IT04K-154-1, IT96K-610, IT97K-1069-6, IT97K-390-2, Ife Bpc, IT99K-573-2-1, Ife Modupe, IT845-2246, Drum, Ife Brown and Oloyin were the most susceptible with the highest damage indices values (p?<?0.05). The range of each of the resistance indices measured in the susceptible varieties was: number of initial eggs laid (61–100), adult emergence (40.0–62.10%), number of seeds damaged (53.0–85.5) and seed weight loss (58.0–66.25%).     

Sexual dimorphism and allometry in two seed beetles (Coleoptera: Bruchidae)

Male Callosobruchus chinensis (Coleoptera: Bruchidae) have elaborated, pectinate antennae, which are absent from conspecific females and both sexes of a congener, Callosobruchus maculatus. To begin to unravel the mechanisms producing this striking dimorphism, we examined which morphological traits best explain body size variation in bruchid beetles and quantified sexual dimorphism of antenna size through allometric analyses. Using principal component analyses, we found that elytron length and pronotum width were significantly correlated with the first principal component, which was interpreted as explaining variation in body size. Regressions of log‐transformed body size measures on log‐transformed antenna length revealed that males of both species had longer antennae than conspecific females for any given body size, although most of this effect was attributable to higher intercepts, rather than increased allometry, in males. Comparisons among heterospecific males revealed that C. maculatus males have noticeably longer antennae than C. chinensis males at large body sizes. Callosobruchus chinensis males, thus, appear to have increased the receptive area of their antennae by adding to the width of, rather than further elongating, their antennae. Finally, we found evidence for positive allometry between log‐transformed antenna length and log‐transformed antenna width in C. chinensis males. We discuss our results in the context of evidence supporting the presence of an additional, and potentially unique, sex pheromone in C. chinensis females.     

Influence of body and genital morphology on relative male fertilization success in oriental beetle

Although the frequently large variance in relative male fertilizationsuccess when females are mated by more than 1 male has beenappreciated for some time, the factors that influence relativepaternity are still poorly understood. Recently, experimentalevidence that morphology of male genitalia influences fertilizationsuccess has been documented in 2 water striders, a dung beetle,and a leaf beetle. We explored the role of male genital morphologyin postcopulatory sexual selection in the oriental beetle. Wemated females to 2 males in succession and assessed relativepaternity by the sterile male technique. Morphology of the malegenitalia was found to strongly influence relative paternitybut only for the first male to mate. Male body size influencedrelative fertilization success as well, but again, only forthe first male; surprisingly, smaller males achieved higherpaternity when mating first. We also found suggestive evidencethat copula duration of both the first and second male to mateinfluenced paternity. Other factors, including female size anddegree of asymmetry of hind tibiae length of males had no effecton relative fertilization success. Our results for the orientalbeetle are novel among sperm precedence studies for 2 reasons:1) traits of the first male appear to be more important in influencingpaternity than those of the second, and 2) smaller, not larger,males achieved greater relative paternity. Our results alsocontribute to the growing body of empirical evidence in supportof the hypothesis that male genitalia evolve by postcopulatorysexual selection.     

Distinctive developmental variability of genital parts in the sexually dimorphic beetle, Prosopocoilus inclinatus (Coleoptera: Lucanidae)

Recent comparative studies have revealed that the rapid diversity of genitalia is closely related to sexual selection and that genital development interacts with the development of different body parts. Hypotheses about developmental stability due to selection to genital parts were tested by estimating allometric relations in a sexually dimorphic stag beetle Prosopocoilus inclinatus . All genital parts of males scaled to body size with a slope of less than 1 and all but the median lobe (male intromittent organ) showed smaller variability than other body parts. This supported the 'one-size-fits-all' hypothesis, which suggests broad copulation opportunity by males of any size with females within a population. Nevertheless, we found large variation among different genital parts in coefficients of variation and in values of the switch point where the allometric relations varied significantly. These results strongly support the view that developmental trajectories of genital traits are not necessarily integrated. Among the genitalic traits, male intromittent organ and female genitalia exhibited large variability, suggesting a high responsiveness to the selective regimes and physical interaction during copulation. This may account for rapid diversification of genital morphology, even in closely-related populations in beetle species.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 90 , 573–581.     

No evidence for divergence in male harmfulness or female resistance in response to changes in the opportunity for dispersal

The outcome of sexual conflict can depend on the social environment, as males respond to changes in the inclusive fitness payoffs of harmfulness and harm females less when they compete with familiar relatives. Theoretical models also predict that if limited male dispersal predictably enhances local relatedness while maintaining global competition, kin selection can produce evolutionary divergences in male harmfulness among populations. Experimental tests of these predictions, however, are rare. We assessed rates of dispersal in female and male seed beetles Callosobruchus maculatus, a model species for studies of sexual conflict, in an experimental setting. Females dispersed significantly more often than males, but dispersing males travelled just as far as dispersing females. Next, we used experimental evolution to test whether limiting dispersal allowed the action of kin selection to affect divergence in male harmfulness and female resistance. Populations of C. maculatus were evolved for 20 and 25 generations under one of three dispersal regimens: completely free dispersal, limited dispersal and no dispersal. There was no divergence among treatments in female reproductive tract scarring, ejaculate size, mating behaviour, fitness of experimental females mated to stock males or fitness of stock females mated to experimental males. We suggest that this is likely due to insufficient strength of kin selection rather than a lack of genetic variation or time for selection. Limited dispersal alone is therefore not sufficient for kin selection to reduce male harmfulness in this species, consistent with general predictions that limited dispersal will only allow kin selection if local relatedness is independent of the intensity of competition among kin.     

Biological activity of tree marigold, Tithonia diversifolia, on cowpea seed bruchid, Callosobruchus maculatus (Coleoptera: Bruchidae)

The powder and ethanol extract of Tithonia diversifolia leaves were tested for their efficacy at five different concentrations (0.0%, 0.5%, 1.0%, 1.5% and 2.0%) on bruchid mortality, oviposition and adult emergence of cowpea seed beetle Callosobruchus maculatus at ambient tropical storage conditions of temperature and relative humidity. Results showed that the leaf extract had a higher bioactivity on oviposition, adult emergence and mortality of C. maculatus. The mean number of eggs laid on seeds treated with extract was reduced from 20.7 in the solvent-treated to 4.7 at 2%Tithonia extract concentration while adult emergence reduced from a mean of 92.2 in the solvent-treated seeds to 72.2 at 2.0% extract treatment. Mortality was 100% at higher concentrations of 3%, 4% and 5% within 24 h of extract application but at lower concentrations mortality was 73.3% and 93.3% at 1% and 2%, respectively after 24 h. Fourty-eight hours after application, 100% mortality of adult C. maculatus was obtained at all concentrations. The mean number of eggs laid was reduced from 41.3 in the untreated to 17.3 at 2.0% powder concentration while adult emergence dropped from 98.5% in the control to 74.2% at the highest powder concentration. The powder was effective at higher concentrations and longer exposure time. At 3%, 4% and 5%, 63–75% mortalities occurred within 24 h of application while at 48 h, mortality was 76–98% at 3–5% powder application rates. The results of this study revealed T. diversifolia as a potential candidate for bioinsecticide preparations because of antiovipositional, ovicidal and knockdown properties of its products, which have some volatile components.     

Allometry of male genitalia in a species of soldier beetle: support for the one-size-fits-all hypothesis

Hypotheses about the functions of the male genitalia and the male scape in insects were tested by measuring the slopes of allometric relations in six populations of Chauliognathus scutellaris. All allometric relations used elytron length as the indicator of overall body size. Male genitalia have lower slopes than male pronota (a structure not involved in reproduction), male scapes (secondary sexual characters) have higher slopes than male pronota, and female scapes have slopes that are not different from the slopes of female pronota. These results support Eberhard's one-size-fits-all hypothesis regarding the size of male genitalia in insects, and they raise questions about the role of the male scape in reproductive activities.     

Polymorphism of competition type and its genetics in Callosobruchus maculatus (Coleoptera: Bruchidae)

Callosobruchus maculatus has both contest and scramble competition strategies. The currently existing theoretical models using game theory suggest that the contest strategy should be selected for. However, most geographic strains of C. maculatus show scramble competition. We experimentally crossed the representative strains of contest and scramble. We expressed the degree of contest competition by a continuous value named the C-value, which ranges from zero (pure scramble) to unity (pure contest). The competition types expressed in the C-value were genetically additive. Their larval developmental rates were negatively correlated with C-values. Multiple-generation experiments of the mixed strains confirmed that there were no overwhelming advantages of contest over scramble type. Most of the mixed strains remained in the intermediate states. We discuss the results in terms of the resource size necessary for developmental success and developmental speed. Received: April 19, 2001 / Accepted: November 2, 2001     

Complex mitonuclear interactions and metabolic costs of mating in male seed beetles

The lack of evolutionary response to selection on mitochondrial genes through males predicts the evolution of nuclear genetic influence on male‐specific mitochondrial function, for example by gene duplication and evolution of sex‐specific expression of paralogs involved in metabolic pathways. Intergenomic epistasis may therefore be a prevalent feature of the genetic architecture of male‐specific organismal function. Here, we assess the role of mitonuclear genetic variation for male metabolic phenotypes [metabolic rate and respiratory quotient (RQ)] associated with ejaculate renewal, in the seed beetle Callosobruchus maculatus, by assaying lines with crossed combinations of distinct mitochondrial haplotypes and nuclear lineages. We found a significant increase in metabolic rate following mating relative to virgin males. Moreover, processes associated with ejaculate renewal showed variation in metabolic rate that was affected by mitonuclear interactions. Mitochondrial haplotype influenced mating‐related changes in RQ, but this pattern varied over time. Mitonuclear genotype and the energy spent during ejaculate production affected the weight of the ejaculate, but the strength of this effect varied across mitochondrial haplotypes showing that the genetic architecture of male‐specific reproductive function is complex. Our findings unveil hitherto underappreciated metabolic costs of mating and ejaculate renewal, and provide the first empirical demonstration of mitonuclear epistasis on male reproductive metabolic processes.     

The effects of copulation duration in the bruchid beetle Callosobruchus maculatus

Control over copulation duration is a potentially importantgenerator of sexual conflict that has received little empiricalattention. The copulatory behavior of the bruchid beetle Callosobruchusmaculatus may reflect a sexual conflict over copulation duration.Males have spines on their intromittent organs that puncturethe female reproductive tract, and females kick their matesduring copulation. If females are prevented from kicking, copulationslast longer and the injuries females sustain are more severe.Males supposedly use the spines as anchors to prolong copulationduration, and females kick to terminate copulations. We manipulatedcopulation duration experimentally and quantified its effectson male and female fitness components to test whether or notthere is a conflict over copulation duration in C. maculatus.Females did not suffer from long copulations but instead experiencedincreased lifetime fecundity. Ejaculate size increased withcopulation duration, and females apparently derive materialbenefits from the ejaculates. Males that mated first and hadlong copulations were relatively unsuccessful when competingwith sperm from other males. However, there was a trend forfemale remating propensity to decrease with long copulationdurations, and first males may therefore also benefit from longcopulations. The copulation duration of the second male to matedid not have a significant effect on sperm precedence. We concludethat even though it seems likely that the male spines have evolvedto act as an anchor during copulation, there seems to be littleconflict over copulation duration per se in C. maculatus.     

Evolution of clutch size in insects. II. A test of static optimality models using the beetle Callosobruchus maculatus (Coleoptera: Bruchidae)

Wilson and Lessells (1993) analysed the effect of constraint assumptions on the predictions of static optimality models for insect clutch size. They concluded that the models could be reliably distinguished between (and hence the main constraints identified) only after precise quantitative predictions had been examined. The present paper describes a series of laboratory experiments, using the bruchid beetle Callosobruchus maculatus, that allow these quantitative predictions to be made and tested. Experiments in which female encounter rate with hosts was altered gave qualitative support for 3 out of 6 basic (single oviposition) models, but the quantitative fit of them all was poor. However, when the (a priori) condition was included in these models that several other females would oviposit on the same hosts (the multiple oviposition models), the time limiting multiple oviposition model alone produced quantitative predictions that were supported by observations. In other words, the results suggest that the main constraints on bruchid oviposition behaviour are the amount of time available for laying eggs and the number of other females ovipositing. However, additional qualitative predictions indicate that the number of eggs available to the female may also constrain clutch size evolutionarily. The usefulness of static optimality models for examining clutch size decisions in insects is discussed in the context of these results.     

'Shouldering' exaggerated genitalia: a unique behavioural adaptation for the retraction of the elongate intromittant organ by the male rove beetle (Aleochara tristis Gravenhorst)

Complex genitalia are ubiquitous among arthropods, but little attention has been given to the fact that the evolution of such elaborate structures may have led to biomechanical constraints that hinder their usage. In the rove beetle, Aleochara tristis, the male's intromittant organ consists of a long flagellum that is more than twice the body length. It is introduced into the spermathecal duct of the female during copulation. The flagellum apparently functions as a guiding rod for a tube growing from the spermatophore that the male deposits in the female's genital chamber. The extraordinary length of the intromittant organ poses a unique physical challenge for the male. During its retraction from the female after mating, the flagellum is under considerable tension. Any sudden release of this tension would result in the flagellum becoming severely entangled, preventing the male from mating again. In response to this novel physical challenge, males have apparently evolved a specialized behavioural adaptation that prevents entanglement after copulation. While retracting the flagellum from the female, the male secures it between a wing shoulder and the pronotum ('shouldering'), holding it taut for about one half of its length. This allows the stepwise retraction of the flagellum from the female and allows it to be retracted back into the male's body in an orderly fashion. This is, to our knowledge, the first demonstration of a behavioural adaptation that has evolved to ameliorate the biomechanical problems caused by exaggerated genital morphology.  © 2005 The Linnean Society of London, Biological Journal of the Linnean Society , 2005, 84 , 307–312.     

四纹豆象不同地理种群的遗传分化

【目的】通过对四纹豆象Callosobruchus maculatus Fabricius不同地理种群mtDNA-Cytb和COⅠ基因部分序列进行比较, 分析其不同地理种群间的遗传分化情况, 为揭示其与生物入侵的关系及入侵过程中种群系统发育地理格局与演变机制提供依据。【方法】用PCR产物直接测序法对分别来自中国海南、喀麦隆、韩国和泰国的四纹豆象4个地理种群的mtDNA-Cytb和COⅠ序列进行测序, 运用软件MEGA3.1对四纹豆象不同地理种群mtDNA-Cytb和COⅠ序列进行序列分析, 以绿豆象C. chinensis为外群构建了不同单倍型的分子系统树。【结果】34条420 bp Cytb序列中共检测到14个多态位点和5种单倍型, 33条822 bp COⅠ序列中检测到28个多态位点和9种单倍型, 其中4种单倍型为独享单倍型, 其余为全部或部分种群的共享单倍型。AMOVA分析结果显示, 四纹豆象4个地理种群间的遗传结构差异并不明显, 遗传差异主要发生在地理种群内。对4个地理种群进行了Fst值和基因流动统计, 结果表明4个地理种群间既存在着一定数量的基因交流, 也存在一定程度的遗传分化。【结论】根据单倍型分布格局初步推测, 中国不可能是四纹豆象的原产地, 而喀麦隆有可能是原产地之一, 并且喀麦隆种群与泰国种群之间的基因交流比较充分, 而中国种群与其他种群之间的遗传分化相对较大。     

Sperm competition risk and male genital anatomy: comparative evidence for reduced duration of female sexual receptivity in primates with penile spines

Selection pressures influencing the way in which males stimulate females during copulation are not well understood. In mammals, copulatory stimulation can influence female remating behaviour, both via neuroendocrine mechanisms mediating control of sexual behaviour, and potentially also via effects of minor injury to the female genital tract. Male adaptations to increase copulatory stimulation may therefore function to reduce sperm competition risk by reducing the probability that females will remate. This hypothesis was tested using data for primates to explore relationships between male penile anatomy and the duration of female sexual receptivity. It was predicted that penile spines or relatively large bacula might function to increase copulatory stimulation and hence to reduce the duration of female sexual receptivity. Results of the comparative analyses presented show that, after control for phylogenetic effects, relatively high penile spinosity of male primates is associated with a relatively short duration of female sexual receptivity within the ovarian cycle, although no evidence was found for a similar relationship between baculum length and duration of female sexual receptivity. The findings presented suggest a new potential function for mammalian penile spines in the context of sexual selection, and add to growing evidence that sperm competition and associated sexual conflict are important selection pressures in the evolution of animal genitalia.