Global invasion history of the agricultural pest butterfly Pieris rapae revealed with genomics and citizen science.

The small cabbage white butterfly, Pieris rapae, is a major agricultural pest of cruciferous crops and has been introduced to every continent except South America and Antarctica as a result of human activities. In an effort to reconstruct the near-global invasion history of P. rapae, we developed a citizen science project, the "Pieris Project," and successfully amassed thousands of specimens from 32 countries worldwide. We then generated and analyzed nuclear (double-digest restriction site-associated DNA fragment procedure [ddRAD]) and mitochondrial DNA sequence data for these samples to reconstruct and compare different global invasion history scenarios. Our results bolster historical accounts of the global spread and timing of P. rapae introductions. We provide molecular evidence supporting the hypothesis that the ongoing divergence of the European and Asian subspecies of P. rapae (∼1,200 y B.P.) coincides with the diversification of brassicaceous crops and the development of human trade routes such as the Silk Route (Silk Road). The further spread of P. rapae over the last ∼160 y was facilitated by human movement and trade, resulting in an almost linear series of at least 4 founding events, with each introduced population going through a severe bottleneck and serving as the source for the next introduction. Management efforts of this agricultural pest may need to consider the current existence of multiple genetically distinct populations. Finally, the international success of the Pieris Project demonstrates the power of the public to aid scientists in collections-based research addressing important questions in invasion biology, and in ecology and evolutionary biology more broadly.

The genome sequence of the grape phylloxera provides insights into the evolution, adaptation, and invasion routes of an iconic pest.

BACKGROUND: Although native to North America, the invasion of the aphid-like grape phylloxera Daktulosphaira vitifoliae across the globe altered the course of grape cultivation. For the past 150 years, viticulture relied on grafting-resistant North American Vitis species as rootstocks, thereby limiting genetic stocks tolerant to other stressors such as pathogens and climate change. Limited understanding of the insect genetics resulted in successive outbreaks across the globe when rootstocks failed. Here we report the 294-Mb genome of D. vitifoliae as a basic tool to understand host plant manipulation, nutritional endosymbiosis, and enhance global viticulture. RESULTS: Using a combination of genome, RNA, and population resequencing, we found grape phylloxera showed high duplication rates since its common ancestor with aphids, but similarity in most metabolic genes, despite lacking obligate nutritional symbioses and feeding from parenchyma. Similarly, no enrichment occurred in development genes in relation to viviparity. However, phylloxera evolved > 2700 unique genes that resemble putative effectors and are active during feeding. Population sequencing revealed the global invasion began from the upper Mississippi River in North America, spread to Europe and from there to the rest of the world. CONCLUSIONS: The grape phylloxera genome reveals genetic architecture relative to the evolution of nutritional endosymbiosis, viviparity, and herbivory. The extraordinary expansion in effector genes also suggests novel adaptations to plant feeding and how insects induce complex plant phenotypes, for instance galls. Finally, our understanding of the origin of this invasive species and its genome provide genetics resources to alleviate rootstock bottlenecks restricting the advancement of viticulture.

Correction to: The genome sequence of the grape phylloxera provides insights into the evolution, adaptation, and invasion routes of an iconic pest.

An amendment to this paper has been published and can be accessed via the original article.

Deciphering the worldwide invasion of the Asian long-horned beetle: A recurrent invasion process from the native area together with a bridgehead effect.

Retracing introduction routes is crucial for understanding the evolutionary processes involved in an invasion, as well as for highlighting the invasion history of a species at the global scale. The Asian long-horned beetle (ALB) Anoplophora glabripennis is a xylophagous pest native to Asia and invasive in North America and Europe. It is responsible for severe losses of urban trees, in both its native and invaded ranges. Based on historical and genetic data, several hypotheses have been formulated concerning its invasion history, including the possibility of multiple introductions from the native zone and secondary dispersal within the invaded areas, but none have been formally tested. In this study, we characterized the genetic structure of ALB in both its native and invaded ranges using microsatellites. In order to test different invasion scenarios, we used an approximate Bayesian "random forest" algorithm together with traditional population genetics approaches. The strong population differentiation observed in the native area was not geographically structured, suggesting complex migration events that were probably human-mediated. Both native and invasive populations had low genetic diversity, but this characteristic did not prevent the success of the ALB invasions. Our results highlight the complexity of invasion pathways for insect pests. Specifically, our findings indicate that invasive species might be repeatedly introduced from their native range, and they emphasize the importance of multiple, human-mediated introductions in successful invasions. Finally, our results demonstrate that invasive species can spread across continents following a bridgehead path, in which an invasive population may have acted as a source for another invasion.

Biases of STRUCTURE software when exploring introduction routes of invasive species.

Population genetic methods are widely used to retrace the introduction routes of invasive species. The unsupervised Bayesian clustering algorithm implemented in STRUCTURE is amongst the most frequently used of these methods, but its ability to provide reliable information about introduction routes has never been assessed. We simulated microsatellite datasets to evaluate the extent to which the results provided by STRUCTURE were misleading for the inference of introduction routes. We focused on an invasion scenario involving one native and two independently introduced populations, because it is the sole scenario that can be rejected when obtaining a particular clustering with a STRUCTURE analysis at K = 2 (two clusters). Results were classified as "misleading" or "non-misleading". We investigated the influence of effective size, bottleneck severity and number of loci on the type and frequency of misleading results. We showed that misleading STRUCTURE results were obtained for 10% of all simulated datasets. Our results highlighted two categories of misleading output. The first occurs when the native population has a low level of diversity. In this case, the two introduced populations may be very similar, despite their independent introduction histories. The second category results from convergence issues in STRUCTURE for K = 2, with strong bottleneck severity and/or large numbers of loci resulting in high levels of differentiation between the three populations. Overall, the risk of being misled by STRUCTURE in the context of introduction routes inferences is moderate, but it is important to remain cautious when low genetic diversity or genuine multimodality between runs are involved.

Multiple introductions, admixture and bridgehead invasion characterize the introduction history of Ambrosia artemisiifolia in Europe and Australia.

Admixture between differentiated populations is considered to be a powerful mechanism stimulating the invasive success of some introduced species. It is generally facilitated through multiple introductions; however, the importance of admixture prior to introduction has rarely been considered. We assess the likelihood that the invasive Ambrosia artemisiifolia populations of Europe and Australia developed through multiple introductions or were sourced from a historical admixture zone within native North America. To do this, we combine large genomic and sampling data sets analysed with approximate Bayesian computation and random forest scenario evaluation to compare single and multiple invasion scenarios with pre- and postintroduction admixture simultaneously. We show the historical admixture zone within native North America originated before global invasion of this weed and could act as a potential source of introduced populations. We provide evidence supporting the hypothesis that the invasive populations established through multiple introductions from the native range into Europe and subsequent bridgehead invasion into Australia. We discuss the evolutionary mechanisms that could promote invasiveness and evolutionary potential of alien species from bridgehead invasions and admixed source populations.

Cannibalism in invasive, native and biocontrol populations of the harlequin ladybird.

BACKGROUND: Cannibalism is widespread in both vertebrates and invertebrates but its extent is variable between and within species. Cannibalism depends on population density and nutritional conditions, and could be beneficial during colonisation of new environments. Empirical studies are needed to determine whether this trait might facilitate invasion of a new area in natural systems. We investigated whether the propensity for cannibalism in H. axyridis differs both between native and invasive populations and between invasive populations from the core and from the front of the invasive area in Western Europe. We also compared the propensity for cannibalism of these natural populations with that of laboratory-reared biocontrol populations. We measured the cannibalism rates of eggs by first instar larvae and adult females at two different individual densities of ladybirds from three types of population (invasive, native and biocontrol), in laboratory-controlled conditions. RESULTS: Cannibalism was significantly greater in larvae from invasive populations compared to native or biocontrol populations, but there was no difference in cannibalism rates between populations from the core or front of the invaded range. Cannibalism was significantly lower in larvae from biocontrol populations compared to wild (invasive and native) populations. No differences in cannibalism rates of adult females were found between any populations. While high population density significantly increased cannibalism in both larvae and adults, the norm of reaction of cannibalism to individual density did not change significantly during the invasion and/or laboratory rearing processes. CONCLUSION: This study is the first to provide evidence for a higher propensity for cannibalism in invasive populations compared to native ones. Our experiments also shed light on the difference in cannibalism evolution with respect to life stages. However, we are still at an early stage in understanding the underlying mechanisms and several different research perspectives are needed to determine whether the higher propensity for cannibalism is a general feature of the invasion process.

Complementarity of statistical treatments to reconstruct worldwide routes of invasion: the case of the Asian ladybird Harmonia axyridis.

Inferences about introduction histories of invasive species remain challenging because of the stochastic demographic processes involved. Approximate Bayesian computation (ABC) can help to overcome these problems, but such method requires a prior understanding of population structure over the study area, necessitating the use of alternative methods and an intense sampling design. In this study, we made inferences about the worldwide invasion history of the ladybird Harmonia axyridis by various population genetics statistical methods, using a large set of sampling sites distributed over most of the species' native and invaded areas. We evaluated the complementarity of the statistical methods and the consequences of using different sets of site samples for ABC inferences. We found that the H. axyridis invasion has involved two bridgehead invasive populations in North America, which have served as the source populations for at least six independent introductions into other continents. We also identified several situations of genetic admixture between differentiated sources. Our results highlight the importance of coupling ABC methods with more traditional statistical approaches. We found that the choice of site samples could affect the conclusions of ABC analyses comparing possible scenarios. Approaches involving independent ABC analyses on several sample sets constitute a sensible solution, complementary to standard quality controls based on the analysis of pseudo-observed data sets, to minimize erroneous conclusions. This study provides biologists without expertise in this area with detailed methodological and conceptual guidelines for making inferences about invasion routes when dealing with a large number of sampling sites and complex population genetic structures.

Extending approximate Bayesian computation with supervised machine learning to infer demographic history from genetic polymorphisms using DIYABC Random Forest.

Simulation-based methods such as approximate Bayesian computation (ABC) are well-adapted to the analysis of complex scenarios of populations and species genetic history. In this context, supervised machine learning (SML) methods provide attractive statistical solutions to conduct efficient inferences about scenario choice and parameter estimation. The Random Forest methodology (RF) is a powerful ensemble of SML algorithms used for classification or regression problems. Random Forest allows conducting inferences at a low computational cost, without preliminary selection of the relevant components of the ABC summary statistics, and bypassing the derivation of ABC tolerance levels. We have implemented a set of RF algorithms to process inferences using simulated data sets generated from an extended version of the population genetic simulator implemented in DIYABC v2.1.0. The resulting computer package, named DIYABC Random Forest v1.0, integrates two functionalities into a user-friendly interface: the simulation under custom evolutionary scenarios of different types of molecular data (microsatellites, DNA sequences or SNPs) and RF treatments including statistical tools to evaluate the power and accuracy of inferences. We illustrate the functionalities of DIYABC Random Forest v1.0 for both scenario choice and parameter estimation through the analysis of pseudo-observed and real data sets corresponding to pool-sequencing and individual-sequencing SNP data sets. Because of the properties inherent to the implemented RF methods and the large feature vector (including various summary statistics and their linear combinations) available for SNP data, DIYABC Random Forest v1.0 can efficiently contribute to the analysis of large SNP data sets to make inferences about complex population genetic histories.

Local dispersal pathways during the invasion of the cactus moth, Cactoblastis cactorum, within North America and the Caribbean.

Cactoblastis cactorum, a species of moth native to Argentina, feeds on several prickly pear cactus species (Opuntia) and has been successfully used as a biological control of invading Opuntia species in Australia, South Africa and native ruderal Opuntia species in some Caribbean islands. Since its introduction to the Caribbean its spread was uncontrolled, invading successfully Florida, Texas and Louisiana. Despite this long history of invasion, we are still far from understanding the factors determining the patterns of invasion of Cactoblastis in North America. Here, we explored three non-mutually exclusive explanations: a) a stepping stone model of colonization, b) long distance colonization due to hurricanes, and/or c) hitchhiking through previously reported commercial routes. Genetic diversity, genetic structure and the patterns of migration among populations were obtained by analyzing 10 nuclear microsatellite loci. Results revealed the presence of genetic structure among populations of C. cactorum in the invaded region and suggest that both marine commercial trade between the Caribbean islands and continental USA, as well as recurrent transport by hurricanes, explain the observed patterns of colonization. Provided that sanitary regulations avoiding human-mediated dispersal are enforced, hurricanes probably represent the most important agent of dispersal and future invasion to continental areas.

Dispersal strategies of phytophagous insects at a local scale: adaptive potential of aphids in an agricultural environment.

BACKGROUND: The spread of agriculture greatly modified the selective pressures exerted by plants on phytophagous insects, by providing these insects with a high-level resource, structured in time and space. The life history, behavioural and physiological traits of some insect species may have evolved in response to these changes, allowing them to crowd on crops and to become agricultural pests. Dispersal, which is one of these traits, is a key concept in evolutionary biology but has been over-simplified in most theoretical studies. We evaluated the impact of the local-scale dispersal strategy of phytophagous insects on their fitness, using an individual-based model to simulate population dynamics and dispersal between leaves and plants, by walking and flying, of the aphid Aphis gossypii, a major agricultural pest, in a melon field. We compared the optimal values for dispersal parameters in the model with the corresponding observed values in experimental trials. RESULTS: We show that the rates of walking and flying disperser production on leaves were the most important traits determining the fitness criteria, whereas dispersal distance and the clustering of flying dispersers on the target plant had no effect. We further show that the effect of dispersal parameters on aphid fitness depended strongly on plant characteristics. CONCLUSION: Parameters defining the dispersal strategies of aphids at a local scale are key components of the fitness of these insects and may thus be essential in the adaptation to agricultural environments that are structured in space and time. Moreover, the fact that the effect of dispersal parameters on aphid fitness depends strongly on plant characteristics suggests that traits defining aphid dispersal strategies may be a cornerstone of host-plant specialization.

Conflicts of Interest in GM Bt Crop Efficacy and Durability Studies.

Public confidence in genetically modified (GM) crop studies is tenuous at best in many countries, including those of the European Union in particular. A lack of information about the effects of ties between academic research and industry might stretch this confidence to the breaking point. We therefore performed an analysis on a large set of research articles (n = 672) focusing on the efficacy or durability of GM Bt crops and ties between the researchers carrying out these studies and the GM crop industry. We found that ties between researchers and the GM crop industry were common, with 40% of the articles considered displaying conflicts of interest (COI). In particular, we found that, compared to the absence of COI, the presence of a COI was associated with a 50% higher frequency of outcomes favorable to the interests of the GM crop company. Using our large dataset, we were able to propose possible direct and indirect mechanisms behind this statistical association. They might notably include changes of authorship or funding statements after the results of a study have been obtained and a choice in the topics studied driven by industrial priorities.

Mealybug species from Chilean agricultural landscapes and main factors influencing the genetic structure of Pseudococcus viburni.

The present study aimed to characterize the distribution of mealybug species along Chilean agro-ecosystems and to determine the relative impact of host plant, management strategy, geography and micro-environment on shaping the distribution and genetic structure of the obscure mealybug Pseudococcus viburni. An extensive survey was completed using DNA barcoding methods to identify Chilean mealybugs to the species level. Moreover, a fine-scale study of Ps. viburni genetic diversity and population structure was carried out, genotyping 529 Ps. viburni individuals with 21 microsatellite markers. Samples from 16 localities were analyzed using Bayesian and spatially-explicit methods and the genetic dataset was confronted to host-plant, management and environmental data. Chilean crops were found to be infested by Ps. viburni, Pseudococcus meridionalis, Pseudococcus longispinus and Planococcus citri, with Ps. viburni and Ps. meridionalis showing contrasting distribution and host-plant preference patterns. Ps. viburni samples presented low genetic diversity levels but high genetic differentiation. While no significant genetic variance could be assigned to host-plant or management strategy, climate and geography were found to correlate significantly with genetic differentiation levels. The genetic characterization of Ps. viburni within Chile will contribute to future studies tracing back the origin and improving the management of this worldwide invader.

The tomato borer, Tuta absoluta, invading the Mediterranean Basin, originates from a single introduction from Central Chile.

The Lepidopteran pest of tomato, Tuta absoluta, is native to South America and is invasive in the Mediterranean basin. The species' routes of invasion were investigated. The genetic variability of samples collected in South America, Europe, Africa and Middle East was analyzed using microsatellite markers to infer precisely the source of the invasive populations and to test the hypothesis of a single versus multiple introductions into the old world continents. This analysis provides strong evidence that the origin of the invading populations was unique and was close to or in Chile, and probably in Central Chile near the town of Talca in the district of Maule.

Secondary contact and admixture between independently invading populations of the western corn rootworm, Diabrotica virgifera virgifera in Europe.

The western corn rootworm, Diabrotica virgifera virgifera (Coleoptera: Chrysomelidae), is one of the most destructive pests of corn in North America and is currently invading Europe. The two major invasive outbreaks of rootworm in Europe have occurred, in North-West Italy and in Central and South-Eastern Europe. These two outbreaks originated from independent introductions from North America. Secondary contact probably occurred in North Italy between these two outbreaks, in 2008. We used 13 microsatellite markers to conduct a population genetics study, to demonstrate that this geographic contact resulted in a zone of admixture in the Italian region of Veneto. We show that i) genetic variation is greater in the contact zone than in the parental outbreaks; ii) several signs of admixture were detected in some Venetian samples, in a bayesian analysis of the population structure and in an approximate bayesian computation analysis of historical scenarios and, finally, iii) allelic frequency clines were observed at microsatellite loci. The contact between the invasive outbreaks in North-West Italy and Central and South-Eastern Europe resulted in a zone of admixture, with particular characteristics. The evolutionary implications of the existence of a zone of admixture in Northern Italy and their possible impact on the invasion success of the western corn rootworm are discussed.

Estimation of demo-genetic model probabilities with Approximate Bayesian Computation using linear discriminant analysis on summary statistics.

Comparison of demo-genetic models using Approximate Bayesian Computation (ABC) is an active research field. Although large numbers of populations and models (i.e. scenarios) can be analysed with ABC using molecular data obtained from various marker types, methodological and computational issues arise when these numbers become too large. Moreover, Robert et al. (Proceedings of the National Academy of Sciences of the United States of America, 2011, 108, 15112) have shown that the conclusions drawn on ABC model comparison cannot be trusted per se and required additional simulation analyses. Monte Carlo inferential techniques to empirically evaluate confidence in scenario choice are very time-consuming, however, when the numbers of summary statistics (Ss) and scenarios are large. We here describe a methodological innovation to process efficient ABC scenario probability computation using linear discriminant analysis (LDA) on Ss before computing logistic regression. We used simulated pseudo-observed data sets (pods) to assess the main features of the method (precision and computation time) in comparison with traditional probability estimation using raw (i.e. not LDA transformed) Ss. We also illustrate the method on real microsatellite data sets produced to make inferences about the invasion routes of the coccinelid Harmonia axyridis. We found that scenario probabilities computed from LDA-transformed and raw Ss were strongly correlated. Type I and II errors were similar for both methods. The faster probability computation that we observed (speed gain around a factor of 100 for LDA-transformed Ss) substantially increases the ability of ABC practitioners to analyse large numbers of pods and hence provides a manageable way to empirically evaluate the power available to discriminate among a large set of complex scenarios.

Biological invasions in agricultural settings: insights from evolutionary biology and population genetics.

Invasion biology and agriculture are intimately related for several reasons and in particular because many agricultural pest species are recent invaders. In this article we suggest that the reconstruction of invasion routes with population genetics-based methods can address fundamental questions in ecology and practical aspects of the management of biological invasions in agricultural settings. We provide a brief description of the methods used to reconstruct invasion routes and describe their main characteristics. In particular, we focus on a scenario--the bridgehead invasion scenario --which had been overlooked until recently. We show that this scenario, in which an invasive population is the source of other invasive populations, is evolutionarily parsimonious and may have played a crucial role in shaping the distribution of many recent agricultural pests.

Can things get worse when an invasive species hybridizes? The harlequin ladybird Harmonia axyridis in France as a case study.

So far, only a few studies have explicitly investigated the consequences of admixture for the adaptative potential of invasive populations. We addressed this question in the invasive ladybird Harmonia axyridis. After decades of use as a biological control agent against aphids in Europe and North America, H. axyridis recently became invasive in four continents and has now spread widely in Europe. Despite this invasion, a flightless strain is still sold as a biological control agent in Europe. However, crosses between flightless and invasive individuals yield individuals able to fly, as the flightless phenotype is caused by a single recessive mutation. We investigated the potential consequences of admixture between invasive and flightless biological control individuals on the invasion in France. We used three complementary approaches: (i) population genetics, (ii) a mate-choice experiment, and (iii) a quantitative genetics experiment. The invasive French population and the biological control strain showed substantial genetic differentiation, but there are no reproductive barriers between the two. Hybrids displayed a shorter development time, a larger size and a higher genetic variance for survival in starvation conditions than invasive individuals. We discuss the potential consequences of our results with respect to the invasion of H. axyridis in Europe.

Inbreeding depression is purged in the invasive insect Harmonia axyridis.

Bottlenecks in population size reduce genetic diversity and increase inbreeding, which can lead to inbreeding depression. It is thus puzzling how introduced species, which typically pass through bottlenecks, become such successful invaders. However, under certain theoretical conditions, bottlenecks of intermediate size can actually purge the alleles that cause inbreeding depression. Although this process has been confirmed in model laboratory systems, it has yet to be observed in natural invasive populations. We evaluate whether such purging could facilitate biological invasions by using the world-wide invasion of the ladybird (or ladybug) Harmonia axyridis. We first show that invasive populations endured a bottleneck of intermediate intensity. We then demonstrate that replicate introduced populations experience almost none of the inbreeding depression suffered by native populations. Thus, rather than posing a barrier to invasion as often assumed, bottlenecks, by purging deleterious alleles, can enable the evolution of invaders that maintain high fitness even when inbred.

Bridgehead effect in the worldwide invasion of the biocontrol harlequin ladybird.

Recent studies of the routes of worldwide introductions of alien organisms suggest that many widespread invasions could have stemmed not from the native range, but from a particularly successful invasive population, which serves as the source of colonists for remote new territories. We call here this phenomenon the invasive bridgehead effect. Evaluating the likelihood of such a scenario is heuristically challenging. We solved this problem by using approximate Bayesian computation methods to quantitatively compare complex invasion scenarios based on the analysis of population genetics (microsatellite variation) and historical (first observation dates) data. We applied this approach to the Harlequin ladybird Harmonia axyridis (HA), a coccinellid native to Asia that was repeatedly introduced as a biocontrol agent without becoming established for decades. We show that the recent burst of worldwide invasions of HA followed a bridgehead scenario, in which an invasive population in eastern North America acted as the source of the colonists that invaded the European, South American and African continents, with some admixture with a biocontrol strain in Europe. This demonstration of a mechanism of invasion via a bridgehead has important implications both for invasion theory (i.e., a single evolutionary shift in the bridgehead population versus multiple changes in case of introduced populations becoming invasive independently) and for ongoing efforts to manage invasions by alien organisms (i.e., heightened vigilance against invasive bridgeheads).