Interspecific gene flow obscures phylogenetic relationships in an important insect pest species complex.

As genomic data proliferates, the prevalence of post-speciation gene flow is making species boundaries and relationships increasingly ambiguous. Although current approaches inferring fully bifurcating phylogenies based on concatenated datasets provide simple and robust answers to many species relationships, they may be inaccurate because the models ignore inter-specific gene flow and incomplete lineage sorting. To examine the potential error resulting from ignoring gene flow, we generated both a RAD-seq and a 500 protein-coding loci highly multiplexed amplicon (HiMAP) dataset for a monophyletic group of 12 species defined as the Bactrocera dorsalis sensu lato clade. With some of the world's worst agricultural pests, the taxonomy of the B. dorsalis s.l. clade is important for trade and quarantines. However, taxonomic confusion confounds resolution due to intra- and interspecific phenotypic variation and convergence, mitochondrial introgression across half of the species, and viable hybrids. We compared the topological convergence of our datasets using concatenated phylogenetic and various multispecies coalescent approaches, some of which account for gene flow. All analyses agreed on species delimitation, but there was incongruence between species relationships. Under concatenation, both datasets suggest identical species relationships with mostly high statistical support. However, multispecies coalescent and multispecies network approaches suggest markedly different hypotheses and detected significant gene flow. We suggest that the network approaches are likely more accurate because gene flow violates the assumptions of the concatenated phylogenetic analyses, but the data-reductive requirements of network approaches resulted in reduced statistical support and could not unambiguously resolve gene flow directions. Our study highlights the importance of testing for gene flow, particularly with phylogenomic datasets, even when concatenated approaches receive high statistical support.

Using the rDNA Internal Transcribed Spacer 1 to Identify the Invasive Pest Rhagoletis cerasi (Diptera: Tephritidae) in North America.

The cherry-infesting fruit fly Rhagoletis cerasi Loew is a significant commercial pest in Europe that has recently invaded North America. To date, it has been trapped only in Canada and northwestern counties of New York. It has the potential to spread further and threaten production and movement of cherry commodities. Timely diagnosis of the pest will facilitate surveys and quick response to new detections. Adult morphology of the pest is distinct from other flies in North America. However, when flies are significantly damaged on traps or the immature life stages are found in fruits, molecular methods of identification are important to confirm presence and host-use records. Other than DNA sequencing of genes from flies which takes over a day to complete, there are no timely methods of molecular identification for this pest. In this study, we report the first sequence record of the internal transcribed spacer 1 (ITS1) from R. cerasi and develop two diagnostic tests for the pest based on ITS1 differences among species in North America. The tests use loop-mediated isothermal amplification (LAMP) and multiplex, conventional polymerase chain reaction (mcPCR) technologies that target the same region of the R. cerasi ITS1 sequence. Both tests performed well when tested against collections of R. cerasi from North America and Europe, generating Diagnostic Sensitivity estimates of 98.4-99.5%. Likewise, the tests had relatively high estimates of Diagnostic Specificity (97.8-100%) when tested against Rhagoletis Loew species present in North America that also use cherry as a developmental host.

Highly variable COI haplotype diversity between three species of invasive pest fruit fly reflects remarkably incongruent demographic histories.

Distance decay principles predict that species with larger geographic ranges would have greater intraspecific genetic diversity than more restricted species. However, invasive pest species may not follow this prediction, with confounding implications for tracking phenomena including original ranges, invasion pathways and source populations. We sequenced an 815 base-pair section of the COI gene for 441 specimens of Bactrocera correcta, 214 B. zonata and 372 Zeugodacus cucurbitae; three invasive pest fruit fly species with overlapping hostplants. For each species, we explored how many individuals would need to be included in a study to sample the majority of their haplotype diversity. We also tested for phylogeographic signal and used demographic estimators as a proxy for invasion potency. We find contrasting patterns of haplotype diversity amongst the species, where B. zonata has the highest diversity but most haplotypes were represented by singletons; B. correcta has ~7 dominant haplotypes more evenly distributed; Z. cucurbitae has a single dominant haplotype with closely related singletons in a 'star-shape' surrounding it. We discuss how these differing patterns relate to their invasion histories. None of the species showed meaningful phylogeographic patterns, possibly due to gene-flow between areas across their distributions, obscuring or eliminating substructure.

Worldwide Phylogeography of Ceratitis capitata (Diptera: Tephritidae) Using Mitochondrial DNA.

The Mediterranean fruit fly, Ceratitis capitata (Weidemann), is one of the most economically important tephritid species worldwide. It has spread across six geographic regions as a result of successful invasions and continues to cause substantial losses to agricultural communities. Our study examined 1,864 flies originating from 150 localities, using mitochondrial DNA sequencing methods. We tested for population structure and revealed the genetic diversity for 1,592 specimens gathered from 144 wild fly collections from 46 countries representing the entire geographic range for this species. We also include in this study 272 Sterile Insect Technique (SIT) specimens from four SIT facilities. We recovered 202 haplotypes from the current sampling and updated previously published work to reveal a total of 231 haplotypes for this pest. These data show population structure at and below the regional level for these collections, shedding light on the current demographics for this species. We observed four common haplotypes, seen among 62% of the samples sequenced that have worldwide distribution. Three haplotypes were seen in SIT flies, with one seen as the predominant haplotype. Our work showed that two of the haplotypes were private to SIT flies, not present among wild fly collections. However, a third haplotype common among wild fly collections was also seen in one SIT facility but at a low frequency based on the current sampling. We provide guidance on the interpretation of these methods for the source estimation of current and future infestations.

Range-wide population genomics of the Mexican fruit fly: Toward development of pathway analysis tools.

Recurrently invading pests provide unique challenges for pest management, but also present opportunities to utilize genomics to understand invasion dynamics and inform regulatory management through pathway analysis. In the southern United States, the Mexican fruit fly Anastrepha ludens is such a pest, and its incursions into Texas and California represent major threats to the agricultural systems of those regions. We developed a draft genome assembly for A. ludens, conducted range-wide population genomics using restriction site-associated DNA sequencing, and then developed and demonstrated a panel of highly differentiated diagnostic SNPs for source determination of intercepted flies in this system. Using 2,081 genomewide SNPs, we identified four populations across the range of A. ludens, corresponding to western Mexico, eastern Mexico/Texas, Guatemala/Belize/Honduras, and Costa Rica/Panama, with some intergradation present between clusters, particularly in Central America. From this population genomics framework, we developed a diagnostic panel of 28 highly differentiated SNPs that were able to recreate the genomewide population structure in this species. We demonstrated this panel on a set of test specimens, including specimens intercepted as part of regular trapping surveillance in Texas and California, and we were able to predict populations of origin for these specimens. This methodology presents a highly applied use of genomic techniques and can be implemented in any group of recurrently invading pests.

The Genetic Diversity of Bactrocera dorsalis (Diptera: Tephritidae) in China and Neighboring Countries: A Review From Published Studies.

For more than a decade, various research groups have tracked the population genetics of the oriental fruit fly, Bactrocera dorsalis (Hendel) in China and neighboring countries using mitochondrial cytochrome c oxidase subunit I (COI) DNA. Although most research has reported high levels of mtDNA variation, to date no efforts have been made to integrate and compare the results from these studies simultaneously. Here, we show that: 1) despite the fact that a large portion of the sampling effort has focused on the Yunnan province beginning in 2005, each subsequent study recovers only a small number of previously sampled haplotypes; 2) new haplotypes of B. dorsalis remain to be found, a projection of new haplotypes versus the number of individuals sampled suggest that sampling the species mtDNA diversity is far from reaching an asymptote; 3) it is unlikely that the observed genetic variation is the result of NUMTs (nuclear mitochondrial DNA), as most differences between haplotypes are silent substitutions; and 4) although all studies employed the 3' end of COI, the length of COI fragment sequenced differs among studies, making comparisons challenging. Therefore, we offer these results with the caveat that mtDNA diversity might be underestimated in China.

Genetic structure and diversity in Brazilian populations of Anastrepha obliqua (Diptera: Tephritidae).

Anastrepha obliqua (Macquart), the West Indian fruit fly, is one of the most economically important pest species in the Neotropical region. It infests an extensive range of host plants that include over 60 species. The geographic range of A. obliqua is from northern Mexico to southern Brazil and includes the Caribbean Islands. Previous molecular studies have revealed significant genetic structure among populations. We used sequences from a fragment of the mitochondrial protein-coding gene cytochrome c oxidase I to estimate structure and genetic diversity of A. obliqua populations from Brazil. We analyzed a total of 153 specimens from the Amazon Forest, Atlantic Forest, Cerrado, and Caatinga biomes. Our study revealed weak genetic structure among the A. obliqua Brazilian populations sampled. Collections from the Amazon Forest had similar haplotype diversity compared to previously reported estimates for collections from the Caribbean and both populations are also closely related to each other, thus challenging the hypothesis that A. obliqua originated in the Caribbean and then moved to other regions of the Americas. Therefore, further evidence is necessary to draw a definite conclusion about the putative center of origin for A. obliqua. Additionally, we suggest a putative historical migration from the west to the east for the A. obliqua Brazilian populations, which could explain the high genetic diversity for this fly in the Amazon Forest and low genetic diversity in the other Brazilian biomes.

Tracking the Origins of Fly Invasions; Using Mitochondrial Haplotype Diversity to Identify Potential Source Populations in Two Genetically Intertwined Fruit Fly Species (Bactrocera carambolae and Bactrocera dorsalis [Diptera: Tephritidae]).

Bactrocera carambolae Drew and Hancock and Bactrocera dorsalis (Hendel) (Diptera: Tephritidae) are important pests of many fruits. These flies have been spread across the world through global travel and trade, and new areas are at risk of invasion. Whenever new invasive populations are discovered, quick and accurate identification is needed to mitigate the damage they can cause. Determining invasive pathways can prevent further spread of pests as well as subsequent reinvasions through the same pathway. Molecular markers can be used for both species identification and pathway analysis. We analyzed 1,601 individuals from 19 populations using 765 base pairs of the mitochondrial cytochrome oxidase I (COI) gene to infer the haplotype diversity and population structure within these flies from across their native and invasive ranges. We analyzed these samples by either grouping by species or geographic populations due to the genetic similarity in the mitochondrial genome. We found no genetic structure between B. dorsalis and B. carambolae and our findings suggest recent and most likely ongoing, genetic exchange between these two species in the wild. Hyper-diverse mitochondrial genetic diversity in the native range suggests large population sizes and relatively high mutation rates. Only 52% of the haplotypes found in the trap captures from California are shared with haplotypes from flies found in our global survey, indicating significant genetic diversity in the native range that is missing from our samples. However, these results provide a foundation for the accurate determination of the provenance of invasive populations around the world.

Identifying Anastrepha (Diptera; Tephritidae) Species Using DNA Barcodes.

Molecular identification of fruit flies in the genus Anastrepha (Diptera; Tephritidae) is important to support plant pest exclusion, suppression, and outbreak eradication. Morphological methods of identification of this economically important genus are often not sufficient to identify species when detected as immature life stages. DNA barcoding a segment of the mitochondrial cytochrome oxidase I gene has been proposed as a method to identify pests in the genus. The identification process for these fruit flies, however, has not been explained in prior DNA barcode studies. DNA barcode methods assume that available DNA sequence records are biologically meaningful. These records, however, can be limited to the most common species or lack population-level measurements of diversity for pests. In such cases, the available data used as a reference are insufficient for completing an accurate identification. Using 539 DNA sequence records from 74 species of Anastrepha, we demonstrate that our barcoding data can distinguish four plant pests: Anastrepha grandis (Macquart) (Diptera; Tephritidae), Anastrepha ludens (Loew), Anastrepha serpentina (Wiedemann), and Anastrepha striata Schiner. This is based on genetic distances of barcode records for the pests and expert evaluation of species and population representation in the data set. DNA barcoding of the cytochrome oxidase I gene alone cannot reliably diagnose the pests Anastrepha fraterculus (Wiedemann), Anastrepha obliqua (Macquart), and Anastrepha suspensa (Loew).

Incongruence between molecules and morphology: A seven-gene phylogeny of Dacini fruit flies paves the way for reclassification (Diptera: Tephritidae).

Molecular and morphological research often suggest conflicting results. Selective pressure on certain morphologies can confound understanding of evolutionary relationships. Dacini is one of the most diverse tribes of tephritid flies and contains many economically important pest species. Their black and yellow patterned body markings are presumed to act as wasp mimicry, and the characters separating species and groups are limited and in some cases phenotypically plastic. The traditional taxonomy of the tribe is controversial because groupings are based on unique combinations of morphological characters without the use of cladistic methods, though recent phylogenetic and taxonomic analyses have resulted in significant changes to their taxonomy. The monophyly of the three largest genera in the tribe has been tested with only small numbers of representatives per genus and a limited number of genes. To further understand the taxonomy and evolution of Dacini we sequenced seven genes from 167 Dacini species and five dipteran outgroups to construct a robust phylogeny and test phylogenetic relationships between genera, subgenera, and species complexes. Our phylogeny confirms the monophyly of Dacus, Bactrocera, and Zeugodacus. However, most groups below the genus level are not monophyletic, and only through further revision will we be able to understand their evolution and clarify the taxonomy within this tribe.

A Real-Time PCR Assay for the Separation of Autographa gamma (Noctuidae: Plusiinae) From Morphologically Similar Species in North America.

The silver Y moth, Autographa gamma L. (Noctuidae: Plusiinae), is a pest of major economic importance in its native range of Europe, Asia, and North Africa. Although not present in North America, larvae of A. gamma are commonly intercepted in commodity shipments at U.S. ports, and adult surveys are conducted each year in more than 20 states. Because of the similarity of A. gamma to several native North American species that are attracted to the same pheromone lure, morphological identification of adults is difficult and requires dissection. In 2010, a specimen of Autographa californica (Speyer, 1875) (Lepidoptera: Noctuidae) from Pennsylvania was incorrectly identified as A. gamma, signaling the need for an alternative method of rapid identification. Here we detail a real-time PCR assay capable of identifying A. gamma specimens in approximately 45 min using extracted DNA. The assay uses a hydrolysis probe that targets a species-specific segment of the CO1 DNA barcode region, while a control probe targets a conserved region of 18S rDNA. The assay was tested with two independent runs of 452 specimens of Plusiinae representing 23 different species. The assay provided unambiguous data 99.7% of the time and did not result in any false positives; these data were used to develop a rule set for interpreting the real-time PCR results. In addition, the same diagnostic probe was tested in bulk sample simulations using real-time PCR and droplet digital PCR where A. gamma could be detected in concentrations as low as 1:1,000,000 (gamma:californica). These experiments provide baseline data for developing a bulk sample assay.

A New Diagnostic Resource for Ceratitis capitata Strain Identification Based on QTL Mapping.

The Mediterranean fruit fly Ceratitis capitata (Wiedemann) is a destructive agricultural pest and the subject of exclusion efforts in many countries. Suppression and eradication of invasive populations to prevent its establishment is facilitated by the release of sterile males using the sterile insect technique (SIT). In SIT release areas, it is critical to accurately discriminate between released sterile males and wild individuals to detect extremely rare invasive individuals in areas inundated with millions of sterile male flies. Current methods for discrimination exist but are not always definitive, and a more reliable method is necessary. To address this, we developed a genotyping assay that can be used to discriminate between sterile males from the SIT strain and wild individuals. This was achieved by identifying single nucleotide polymorphisms (SNPs) linked to the maintained traits that facilitate male-only releases, white pupae (wp) and temperature-sensitive lethal (tsl), via QTL mapping. This resulted in the identification of one SNP that was in near-perfect linkage disequilibrium between genotype at this locus and the pupal color phenotype. Medfly from many SIT colonies and wild individuals from across its geographic range were genotyped for this locus, and results show its consistency in identifying SIT flies. In addition, linkage and QTL mapping of wp and tsl have larger impacts as they can serve as foundational tools to identify the genetic basis of traits that facilitate the separation of males from female flies, which can be used to develop SIT programs in related species.

Phylogenetic relationships of the tribe Toxotrypanini (Diptera: Tephritidae) based on molecular characters.

Current hypotheses of relationship among the species of the fruit fly genera Anastrepha and Toxotrypana are tested using sequence data from six DNA regions: the mitochondrial regions 16S, CAD, and COI, and the nuclear regions EF1a, PER, and PGD. DNA sequences were obtained from 146 species of Anastrepha, representing 19 of the 21 species groups as well as five of the six clades of the robusta group, and four species of Toxotrypana in addition to species of Hexachaeta, Pseudophorellia, Alujamyia, and 13 other tephritid genera used as outgroups. The results indicate that Hexachaeta is more closely related to the Molynocoelia group than to Toxotrypana and Anastrepha, and it is removed from the tribe Toxotrypanini. The group Anastrepha+Toxotrypana and the genus Toxotrypana are strongly supported as monophyletic, consistent with previous studies, but Toxotrypana arises within Anastrepha, confirming that Anastrepha as currently defined is paraphyletic. The placement of Toxotrypana within Anastrepha is clearly defined for the first time with high support, as the sister group to the cryptostrepha clade of the robusta group of Anastrepha. Within Anastrepha, the daciformis, dentata, leptozona, raveni, and striata species groups are highly supported clades. The serpentina group is recognized with lower support, and the fraterculus and pseudoparallela groups are supported with minor alterations. The robusta group is resolved as polyphyletic, but four of the six species clades within it are recovered monophyletic (one clade is not represented and another is represented by one species). The punctata and panamensis groups are resolved together in a clade. At least some species of the mucronota group are related, however this group requires further study. The benjamini, grandis, and spatulata groups appear to be polyphyletic. Relationships among the species groups are generally poorly resolved, with the following exceptions: (1) the lineage including Toxotrypana, the cryptostrepha clade, and the tripunctata group; (2) the sister group relationship of the daciformis+dentata groups; (3) a clade comprising the punctata and panamensis groups; and (4) the large clade comprising the pseudoparallela+spatulata+ramosa+grandis+serpentina+striata+fraterculus groups.

A Multiplex Real-Time PCR Assay to Diagnose and Separate Helicoverpa armigera and H. zea (Lepidoptera: Noctuidae) in the New World.

The Old World bollworm, Helicoverpa armigera (Hübner), and the corn earworm, H. zea (Boddie), are two of the most important agricultural pests in the world. Diagnosing these two species is difficult-adults can only be separated with a complex dissection, and larvae cannot be identified to species using morphology, necessitating the use of geographic origin for identification in most instances. With the discovery of H. armigera in the New World, identification of immature Helicoverpa based on origin is no longer possible because H. zea also occurs in all of the geographic regions where H. armigera has been discovered. DNA barcoding and restriction fragment length polymorphism (RFLP) analyses have been reported in publications to distinguish these species, but these methods both require post-PCR processing (i.e., DNA sequencing or restriction digestion) to complete. We report the first real-time PCR assay to distinguish these pests based on two hydrolysis probes that bind to a segment of the internal transcribed spacer region 2 (ITS2) amplified using a single primer pair. One probe targets H. armigera, the second probe targets H. zea, and a third probe that targets a conserved segment of 18S rDNA is used as a control of DNA quality. The assay can be completed in 50 minutes when using isolated DNA and is successfully tested on larvae intercepted at ports of entry and adults captured during domestic surveys. We demonstrate that the assay can be run in triplex with no negative effects on sensitivity, can be run using alternative real-time PCR reagents and instruments, and does not cross react with other New World Heliothinae.

Phylogeographic Structure in Anastrepha ludens (Diptera: Tephritidae) Populations Inferred With mtDNA Sequencing.

Anastrepha ludens (Loew) (Diptera: Tephritidae), the Mexican fruit fly, is a major pest of citrus and mango. It has a wide distribution in Mexico and Central America, with infestations occurring in Texas, California, and Florida with origins believed to have been centered in northeastern Mexico. This research evaluates the utility of a sequence-based approach for two mitochondrial (COI and ND6) gene regions. We use these markers to examine genetic diversity, estimate population structure, and identify diagnostic information for A. ludens populations. We analyzed 543 individuals from 67 geographic collections and found one predominant haplotype occurring in the majority of specimens. We observed 68 haplotypes in all and see differences among haplotypes belonging to northern and southern collections. Mexico haplotypes differ by few bases possibly as a result of a recent bottleneck event. In contrast to the hypothesis suggesting northeastern Mexico as the origin of this species, we see that specimens from two southern collections show high genetic variability delineating three mitochondrial groups. These data suggest that Central America is the origin for A. ludens. We show that COI and ND6 are useful for phylogeographic studies of A. ludens.

A phylogenetic assessment of the polyphyletic nature and intraspecific color polymorphism in the Bactrocera dorsalis complex (Diptera, Tephritidae).

The Bactrocera dorsalis complex (Tephritidae) comprises 85 species of fruit flies, including five highly destructive polyphagous fruit pests. Despite significant work on a few key pest species within the complex, little has been published on the majority of non-economic species in the complex, other than basic descriptions and illustrations of single specimens regarded as typical representatives. To elucidate the species relationships within the Bactrocera dorsalis complex, we used 159 sequences from one mitochondrial (COI) and two nuclear (elongation factor-1α and period) genes to construct a phylogeny containing 20 described species from within the complex, four additional species that may be new to science, and 26 other species from Bactrocera and its sister genus Dacus. The resulting concatenated phylogeny revealed that most of the species placed in the complex appear to be unrelated, emerging across numerous clades. This suggests that they were placed in the Bactrocera dorsalis complex based on the similarity of convergent characters, which does not appear to be diagnostic. Variations in scutum and abdomen color patterns within each of the non-economic species are presented and demonstrate that distantly-related, cryptic species overlap greatly in traditional morphological color patterns used to separate them in keys. Some of these species may not be distinguishable with confidence by means other than DNA data.

Genetic Diversity of Bactrocera dorsalis (Diptera: Tephritidae) on the Hawaiian Islands: Implications for an Introduction Pathway Into California.

Population genetic diversity of the oriental fruit fly, Bactrocera dorsalis (Hendel), on the Hawaiian islands of Oahu, Maui, Kauai, and Hawaii (the Big Island) was estimated using DNA sequences of the mitochondrial cytochrome c oxidase subunit I gene. In total, 932 flies representing 36 sampled sites across the four islands were sequenced for a 1,500-bp fragment of the gene named the C1500 marker. Genetic variation was low on the Hawaiian Islands with >96% of flies having just two haplotypes: C1500-Haplotype 1 (63.2%) or C1500-Haplotype 2 (33.3%). The other 33 flies (3.5%) had haplotypes similar to the two dominant haplotypes. No population structure was detected among the islands or within islands. The two haplotypes were present at similar frequencies at each sample site, suggesting that flies on the various islands can be considered one population. Comparison of the Hawaiian data set to DNA sequences of 165 flies from outbreaks in California between 2006 and 2012 indicates that a single-source introduction pathway of Hawaiian origin cannot explain many of the flies in California. Hawaii, however, could not be excluded as a maternal source for 69 flies. There was no clear geographic association for Hawaiian or non-Hawaiian haplotypes in the Bay Area or Los Angeles Basin over time. This suggests that California experienced multiple, independent introductions from different sources.

Molecular diagnosis of populational variants of Anthonomus grandis (Coleoptera: Curculionidae) in North America.

The utility of the cytochrome oxidase I (COI) DNA sequence used for DNA barcoding and a Sequence Characterized Amplified Region for diagnosing boll weevil, Anthonomus grandis Boheman, variants was evaluated. Maximum likelihood analysis of COI DNA sequences from 154 weevils collected from the United States and Mexico supports previous evidence for limited gene flow between weevil populations on wild cotton and commercial cotton in northern Mexico and southern United States. The wild cotton populations represent a variant of the species called the thurberia weevil, which is not regarded as a significant pest. The 31 boll weevil COI haplotypes observed in the study form two distinct haplogroups (A and B) that are supported by five fixed nucleotide differences and a phylogenetic analysis. Although wild and commercial cotton populations are closely associated with specific haplogroups, there is not a fixed difference between the thurberia weevil variant and other populations. The Sequence Characterized Amplified Region marker generated a larger number of inconclusive results than the COI gene but also supported evidence of shared genotypes between wild and commercial cotton weevil populations. These methods provide additional markers that can assist in the identification of pest weevil populations but not definitively diagnose samples.

Phylogeography of Anastrepha obliqua inferred with mtDNA sequencing.

Anastrepha obliqua (Macquart) (Diptera: Tephritidae), the West Indian fruit fly, is a frugivorous pest that occasionally finds its way to commercial growing areas outside its native distribution. It inhabits areas in Mexico, Central and South America, and the Caribbean with occasional infestations having occurred in the southern tier states (California, Florida, and Texas) of the United States. This fly is associated with many plant species and is a major pest of mango and plum. We examine the genetic diversity of the West Indian fruit fly based on mitochondrial COI and ND6 DNA sequences. Our analysis of 349 individuals from 54 geographic collections from Mexico, Central America, the Caribbean, and South America detected 61 haplotypes that are structured into three phylogenetic clades. The distribution of these clades among populations is associated with geography. Six populations are identified in this analysis: Mesoamerica, Central America, Caribbean, western Mexico, Andean South America, and eastern Brazil. In addition, substantial differences exist among these genetic types that warrants further taxonomic review.

Patterns of mitochondrial haplotype diversity in the invasive pest Epiphyas postvittana (Lepidoptera: Tortricidae).

The light brown apple moth, Epiphyas postvittana (Walker) (Lepidoptera: Tortricidae), is a horticultural pest of Australia and New Zealand that has more recently invaded Hawaii, Europe, and California. A 2,216-bp region of the mitochondrial genome containing the cytochrome oxidase I and II genes was sequenced from 752 individuals. Haplotype network analyses revealed a major split between a predominantly Western Australian clade and all other samples, suggestive of either a deep genetic divergence or a cryptic species. Nucleotide and haplotype diversity were highest in the country of origin, Australia, and in New Zealand populations, with evidence of haplotype sharing between New Zealand and Tasmania. Nucleotide and haplotype diversity were higher in California than within the British Isles or Hawaii. From the total of 96 haplotypes, seven were found in California, of which four were private. Within California, there have been at least two introductions; based on genetic diversity we were unable to assign a likely source for a single moth found and eradicated in Los Angeles in 2007; however, our data suggest it is unlikely that Hawaii and the British Isles are sources of the major E. postvittana population found throughout the rest of the state since 2006.