Survival of the Colorado potato beetle on virus-infected tomato in relation to plant nitrogen and alkaloid content

Infection by tobacco mosaic virus improved the suitability of tomato, Lycopersicon esculentum Mill. for survival of Colorado potato beetle, Leptinotarsa decemlineata (Say), larvae. This improvement was due, at least in part, to the increase in total nitrogen content of virus-infected plants. The simultaneous increase in tomatine content had no discernable effect on L. decemlineata survival. Our results are consistent with the suggestions that virus infection may improve the suitability of partially resistant or non-preferred hosts, and that virus infection may facilitate the adaption of phytophagous insects to such marginal host plant species.

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Résumé Pour vérifier l'hypothèse selon laquelle une contamination par un virs végétal peut accroître la consommation de plantes délaissées ou partiellement résistantes, nous avons examiné les interactions entre Leptinotarsa decemlineata Say, Lycopersicon esculentum Mill., et le virus de la mosaïque du tabac (TMV). La survie des larves de L. decemlineata a augmenté avec la contamination par TMV; les teneurs en azote et en tomatine étaient toutes deux plus élevées dans les plantes contaminées par le virus. La survie a augmenté linéairement avec la teneur en azote, mais sans dépendre de la teneur en tomatine. L'influence positive pour L. decemlineata de la contamination par le virus peut être attribuée au moins en partie à la teneur en azote plus élevée. Nos résultats correspondent à l'hypothèse selon laquelle la consommation de plantes délaissées ou partiellement résistantes peut être augmentée par la contamination virale, et que cette contamination peut faciliter l'adaptation d'insectes phytophages sur des plantes marginalement consommables.

     

Citrus limonoid effects on Colorado potato beetle larval survival and development

The effects of citrus limonoids, applied topically to potato (Solanum tuberosum L. var. Katahdin) foliage, on Colorado potato beetle (Leptinotarsa decemlineata) (Say) (Chrysomelidae) larval development, growth, and survival were quantified in laboratory assays and a small-plot field test. In laboratory assays, survival, development rate, and body weight decreased with increasing limonoid concentration, however these measures of larval response did not significantly differ among varying periods of limonoid exposure (three, six, or nine days). Significant limonoid application concentration and frequency effects on survival, development rate, and defoliation were observed in the field test. These results indicate the potential utility of lethal and non-lethal effects of citrus limonoids for management of the Colorado potato beetle.     

Effects ofBacillus thuringiensis var.San diego on predation effectiveness,development and mortality ofColeomegilla maculata lengi (Col.: Coccinellidae) larvae

Laboratory experiments were conducted to determine the effects of M-One^™ (Bacillus thuringiensis var.san diego) on larval instars ofColeomegilla maculata lengi Timberlake. Coccinellid larval development (from egg hatch to adult), completed on pollen treated with suspensions of M-One^™ at 20 ml/litre (5.6×10⁸ CPBIU/litre) and 200 ml/litre, took respectively 29.3 and 38.5 days compared with 21.9 days for the control (water). M-One^™ did not cause larval mortality.C. maculata third instars did not show any preference between eggs ofLeptinotarsa decemlineata (Say) treated with water or with M-One^™ at 20 ml/litre. However, at 200 ml M-One^™/litre, the number of eggs attacked was 34.7% lower than the eggs treated with water only, 48 h after the beginning of the test. These results indicate that the use of M-One^™, at the manufacturer's recommended field rate of 20 ml/litre, does not cause a major threat to larvalC. maculata populations.

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Résumé Des bioessais en laboratoire ont été effectués afin de déterminer si les larves de la coccinelle maculée,Coleomegilla maculata lengi Timberlake (Col.: Coccinellidae), sont affectées par M-One^™, un insecticide biologique préparé à partir de la bactérieBacillus thuringiensis var.san diego Berliner et utilisé dans la lutte contre le doryphore de la pomme de terre,Leptinotarsa decemlineata (Say) (Col.: Chrysomelidae). Le développement larvaire, effectué sur du pollen traité avec des concentrations de 20 ml M-One^™/litre (5,6×10⁸ unités internationales de doryphore/litre) et 200 ml M-One^™/litre, a nécessité 29,3 et 38,5 jours respectivement, comparativement à 21,9 jours pour le témoin (eau). M-One^™ n'a pas causé de mortalité chez les larves. Au cours d'un test de 48 h, les larves de stade III n'ont montré aucune préférence entre des œufs traités avec 20 ml M-One^™/litre et des œufs traités avec de l'eau. Par contre avec 200 ml M-One^™/litre, le nombre d'œufs attaqués a diminué significativement de 34,7% par rapport au témoin, 48 h après le début du test. Ces résultats indiquent que l'utilisation de M-One^™ à la concentration recommandée de 20 ml/litre ne représente pas une menace pour les populations larvaires de la coccinelle maculée.

     

Antifeedant and toxic effects of drimanes on Colorado potato beetle larvae

Among the drimane compounds tested, the dialdehydes polygodial and warburganal were the most active as antifeedants against Colorado potato beetle larvae, Leptinotarsa decemlineata Say (Coleoptera: Chrysomelidae), in a dual-choice assay with potato, Solanum tuberosum L., leaf discs. Lactones were less effective. Direct observations showed that decreased feeding on leaf discs treated with polygodial and warburganal was accompanied by increased locomotry activity. Topical application of these two compounds on the insect's cuticle decreased food intake of untreated leaf discs, indicating that besides deterrent effects, toxic properties of these molecules influence feeding behaviour.     

Resistance of three wild tuber-bearing potatoes to the Colorado potato beetle

The resistance of Solanum okadae Hawkes & Hjert. (PI 458367), Solanum oplocense Hawkes (PI 473368), and Solanum tarijense Hawkes (PI 414150) to the Colorado potato beetle, Leptinotarsa decemlineata (Say) (Chrysomelidae: Chrysomelini), was studied. In replicated field trials all three accessions showed a high level of resistance to the beetle. No significant genetic variability between genotypes of the same species was found. Results from host acceptance behavior experiments, suitability for larval development tests, foliage consumption tests, and adult survival and oviposition tests supported the hypothesis that the mode of resistance differs between the three wild Solanum species. Solanum okadae and S. oplocense affected host acceptance and consumption. Because the beetle reacted differently to these two species it was hypothesized that the antifeedant chemical(s) differed in nature or quantity. S. tarijense contrasted with the other two species by affecting mostly adult colonization and oviposition.     

Sperm utilization and post-copulatory female-guarding in the Colorado potato beetle, Leptinotarsa decemlineata

Crosses between white and black color morphs of the Colorado potato beetle, Leptinotarsa decemlineata Say, used as genetic markers, showed that male beetles ride and guard females to achieve the three copulations required to fill the spermatheca. Multiple matings are necessary for the females to realize their full reproductive potential. Sperm mixing occurs in the spermatheca providing partial sperm precedence. The data support the competitive mate searching theory.

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Résumé Différentes combinaisons d'accouplements entre des mutants noirs et blancs du doryphore de la pomme de terre, Leptinotarsa decemlineata, servirent à étudier le comportement de guarde de la femelle par le mâle après un premier accouplement. Les résultats indiquent que le sperme de deux mâles accouplant la même femelle se mélange dans la spermathèque assurant ainsi un certain niveau de précédence du sperme du dernier mâle.La masse de sperme dans la spermathèque était inférieure à sa capacité chez les femelles ayant été accouplées une ou deux fois. Le doryphore mâle aurait donc avantage à demeurer sur la femelle après un premier accouplement afin d'obtenir le minimum de trois accouplements requis pour remplir la spermathèque avant que les autres mâles puissent accoupler cette femelle. Les autres mâles ont avantage à essayer de dérober cette femelle pour remplir eux mêmes la spermathèque ou tout au moins profiter d'une certaine précédence du sperme du dernier mâle pour fertiliser une partie des oeufs.Sans accouplements multiples, la femelle ne possède pas suffisamment de sperme pour féconder tous ses oeufs. II semble aussi que les accouplements répétés puissent augmenter sa fécondité.Les résultats supportent une stratégie reproductive de compétition plutôt qu'une simple maximisation du nombre d'accouplements.

     

Suppression of larval Colorado potato beetle growth and development by digestive proteinase inhibitors

Evidence is presented that the Colorado potato beetle (CPB) (Leptinotarsa decemlineata (Say)) depends, at least partially, on cysteine proteinases for protein digestion. Midgut homogenates of CPB larvae have a mildly acidic pH and exhibit major proteolytic activity in the mildly acidic pH range. This proteolytic activity is activated by reducing agents, is inhibited by E-64 (a specific cysteine proteinase inhibitor), and is not inhibited by serine proteinase inhibitors. In addition, consumption of E-64 treated potato leaves by CPB larvae at rates as low as 0.8 g/cm² of leaf tissue has a deleterious effect on larval growth and development.

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Résumé Les protéinases intestinales des larves de Leptinotarsa decemlineata sont en partie caractérisées par les pH limitant leur activité, et par l'effet d'inhibiteurs de protéinases de spécificités connues. Les protéinases ont été testées avec la méthémoglobine tritiée comme substrat et nous avons déterminé les taux relatifs de peptides radioactives TCA solubles, libérées, dans des conditions précises, pour des pH compris entre 2,0 et 12,0. Le taux le plus élevé d'activité protéolytique a été observé pour des pH 5,0 à 6,0, bien qu'il y ait eu une activité appréciable aux pH 4,0 et 8,0. Parmi les inhibiteurs examinés, le E-64, inhibiteur protéinase cystéine très sélectif, a été le plus efficace sur l'activité protéinase. La pepstatine, inhibiteur protéinase aspartique, a été actif, mais seulement dans une gamme plus moin de pH. Les inhibiteurs protéinase sérine, ont été pratiquement inactifs sur l'activité protéinase.Chez des larves de doryphores nourries de feuilles de pommes de terre traitées avec différentes concentrations de E-64 ou de pepstatine, la consommation de E-64 a retardé fortement la croissance larvaire et le développement, la pepstatine a provoqué un retard du développement plus limitée.Nos résultats suggèrent que le doryphore dépend, tout au moins partiellement, des protéinases cystéine pour l'assimilation des protéines.

     

Ultrastructure of the neurosecretory system of the Colorado potato beetle,Leptinotarsa decemlineata (Say)

Summary The ultrastructure of seven types of neurosecretory cells (NSC) in the medial and lateral groups of the protocerebrum is described. The differences among cell types established earlier by light microscopy parallel differences in size and appearance of the neurosecretory particles observed in electron micrographs. No relationship was found between the affinity for Gomori's paraldehyde fuchsin stain and the nature of the particles.The secretions of the A-, A₁-, and C-types of NSC of the medial group are characterized by electron-dense neurosecretory granules of 1250 Å dia., medium-dense granules of 2100 Å, and electron-lucent vesicles of 1700 Å, respectively. The L-type NSCof the lateral group contain smaller (1300 Å) or larger (1700 Å) neurosecretory granules. The medial B- and E-types of NSC and the lateral L_B-type contain granulated vesicles (1200 Å) of the same appearance. These cell types differ in other respects and most likely have separate functions.The author wishes to thank the Laboratory of Virology of the Agricultural University for the use of the electron microscope, Mr. J. Groenewegen and Miss J. van Rinsum for technical assistance, and Professor J. Lattin for correcting the English text. Part of the work has been done while the author was in the service of the Netherlands Organization for the Advancement of Pure Research (ZWO, grant 942-48), and the National Council for Agricultural Research (TNO).     

A proctolin-like peptide and its immunocytochemical localization in the Colorado potato beetle,Leptinotarsa decemlineata

Summary Evidence is presented that neurons in the adult Colorado potato beetle contain a proctolin-like substance. By use of immunocytochemical methods the location of immunoreactive neurons in the central and stomatogastric nervous systems is described. No such neurons were found in the proto- and deutocerebrum or optic lobe. Few immunoreactive neurons are present in the tritocerebrum and numerous proctolin-immunoreactive neurons occur in all ventral ganglia and in the frontal ganglion. Two groups of neurosecretory cells in the suboesophageal ganglion contain a proctolin-immunoreactive substance. In these cells this material is co-localized with a bovine pancreatic polypeptide/FMRF amide-like substance and with a vasopressin/vasotocin/oxytocin-like substance. Proctolin-immunoreactive axon terminals were found on the musculature of the fore- and hindgut and of the vas deferens, and on some segmental muscles. Furthermore, proctolin-immunoreactive neurosecretory axon terminals were found in the corpus cardiacum. The proctolin-like substance may therefore function both as a neurotransmitter/neuromodulator and as a neurohormone. The presence of a proctolin-like substance was also demonstrated with a sensitive bioassay. On fractionation of extracts of the nervous systems of Leptinotarsa decemlineata with high performance liquid chromatography most of the proctolin-like bioactive material comigrated with authentic proctolin. This shows that a proctolin-like substance in this insect is very similar to, if not identical with, the known pentapeptide proctolin.     

Disrupting spring colonization of Colorado potato beetle to nonrotated potato fields

Overwintering Colorado potato beetles (Leptinotarsa decemlineata (Say)) were concentrated primarily within woody borders, and mortality was lower in borders than in potato fields. After overwintering, only 15–44% of live beetles were in the potato fields. In experiments with small plots, colonization of fields from woody borders was reduced ∼60% by a trap crop, either treated with adulticide or with beetles collected daily. Such trap crops, or simply pitfall traps to prevent colonization from woody borders, could significantly reduce early-season adult numbers and subsequent larval populations. However, success is dependent on the local densities of overwintered beetles, and the prevalence of arrestment behavior in the case of trap crops.     

Structure-activity studies of modified citrus limonoids as antifeedants for Colorado potato beetle larvae, Leptinotarsa decemlineata

Limonin and ten structurally modified limonins were evaluated as antifeedants against 4th instar larvae of the Colorado potato beetle, Leptinotarsa decemlineata Say, in no-choice leaf dics assays. The epoxide and furan groups were shown to be essential structural requirements for high antifeedant activity.

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Résumé Le pouvoir phagodissuasif de la limonine et de 10 limonines dont la structure avait été modifiée a été évalué sur des disques de feuilles de Solanum tuberosum offerts à des larves du 4ème stade de L. decemlineata. Aucune activité dissuasive n'ayant été observée avec le tétrahydrolimonine ou le déoxylimonine, l'anneau furane et le groupe époxy sont donc nécessaires pour obtenir cette activité. Ni la réduction de la fonction 7-kéto, ni la rupture de l'anneau A de la limonine n'ont eu d'effêt sur cette activité. La réduction des fonctions 16-carbonyl et 7-kéto de la limonine ont diminué l'activité, mettant en évidence une action possible de l'anneau D-lactone dans l'activité phagodissuasive. Tandis que la déépoxydation de la limonine entraîne une perte d'activité totale, une restauration partielle de cette activité a pu être obtenu par une réduction ultérieure de la fonction 7-kéto.

     

Functional Response ofColeomegilla maculata(Coleoptera: Coccinellidae) to Colorado Potato Beetle Eggs (Coleoptera: Chrysomelidae)

A comparative study of the functional response ofColeomegilla maculataDeGeer (Coleoptera: Coccinellidae) fourth instars was conducted under laboratory, greenhouse, and field conditions. In the laboratory, individual larvae were placed in 9-cm petri dishes for 24 h, with 1, 3, 5, or 7 Colorado potato beetle (Leptinotarsa decemlineata[Say]) (Coleoptera: Chrysomelidae) egg masses. Each egg mass was standardized at 15 eggs. In the greenhouse and field,C. maculatalarvae were provided with an equivalent of 0.5 to 35L. decemlineataegg masses/m²of potato leaf. Fourth instars ofC. maculataexhibited a type II functional response toL. decemlineataeggs under laboratory, greenhouse, and field conditions. Predator search efficiency was inversely related with prey density. The maximum mean attack rate (8.7 eggs) byC. maculatalarvae in the field was about half the mean attack rate in the laboratory (17.6 eggs) and greenhouse (20.1 eggs). The difference in prey density between the laboratory and field seems to have been a major contributing factor in determining the rate of predation, whereas differences in environmental conditions (e.g., temperature and possible alternate food) may explain the differences observed in the predation rate in the greenhouse and field.     

Potential impact of Coleomegilla maculata predation on adaptation of Leptinotarsa decemlineata to Bt-transgenic potatoes

The relationship between Leptinotarsa decemlineata Say egg density and Coleomegilla maculata DeGeer predatory behavior was investigated at different spatial scales (plant-to-plant and plot-to-plot). Both adult C. maculata location and daily egg consumption rates were monitored over time in greenhouse and field tests. Despite aggregation in areas of highest prey density by C. maculata, egg consumption was inversely related to egg mass density at the smallest and the largest spatial scales tested. The experimental data on predation rates in high and low density field treatments were included in a mathematical model to simulate impact of natural enemies on the rate of L. decemlineata adaptation to Bt-toxin-expressing transgenic potato plants when Bt-expressing plants are mixed at the plot-to-plot level with normal potato plants. Results showed that C. maculata predatory behavior could decrease the rate at which L. decemlineata adapted to Bt-toxins if plot-to-plot mixed-plantings were used.     

Compatibility of a natural enemy, Coleomegilla maculata lengi (Col., Coccinellidae) and four insecticides used against the Colorado potato beetle (Col., Chrysomelidae)

Abstract:   The toxicity of four insecticides used to control the Colorado potato beetle, Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae), imidacloprid (Admire^®), cryolite (Kryocide^®), cyromazine (Trigard^®), and Bacillus thuringiensis var. tenebrionis (Novodor^®), to one of its natural enemies, the 12-spotted lady beetle, Coleomegilla maculata lengi Timberlake (Coleoptera: Coccinellidae) was determined in the laboratory. Toxicity assays against C. maculata adults and larvae consisted of (1) topical applications and (2) exposures to treated foliage and prey, using concentrations up to 10 times the manufacturer's recommendations. Over a 6-day period, cyromazine (insect growth regulator) and B. t . var. tenebrionis (microbial insecticide) had no lethal effects on first and third instars C. maculata . For both larval and adult stages, cryolite (inorganic insecticide) caused very low predator mortality when topically applied and moderate mortality when ingested through contaminated eggs of Colorado potato beetles. Imidacloprid (systemic organic insecticide) was highly toxic to adult and larval C. maculata . Its estimated LD₅₀ at 6 days following treatment, corresponded to 0.02–0.09 times the recommended field concentration, depending on the developmental stage and mode of contamination. These results indicate that integrated pest management programmes for Colorado potato beetles using imidacloprid or, to a lesser degree, cryolite, would be detrimental to C. maculata . Cyromazine and B. t . var. tenebrionis seem to present a better compatibility with the protection of C. maculata populations.     

Behavioral responses of Colorado potato beetle larvae to combinations of temperature and insolation, under field conditions

In short-term field trials at combinations of ambient temperature (°C) and insolation (W·m⁻²), larval Colorado potato beetles (Leptinotarsa decemlineata [Say] [Coleoptera: Chrysomelidae]) were observed after their release on the adaxial surface of leaflets on potato plants (Solanum tuberosum L. Solanaceae). The larvae either began feeding or moved under the leaflet; mean interval from release to expression of these behaviors (2.9±0.05 min [n =358]) was independent of air temperature and insolation. Proportion of larvae moving under the leaflet increased logistically with both air temperature and insolation. A 1 W·m⁻² change in insolation (P) evoked the same effect on this proportion as a 0.0838 °C change in air temperature (T _a ), so the two quantities were combined as T^*=T _a +P·0.0838 °C/(W·m⁻²), which has units of °C. The proportion of larvae moving under the leaflet increased logistically with T^*. In 1-day field trials we monitored air temperature, insolation and proportion of larvae under the leaflet, and compared the latter to predictions from the logistic regression derived from the short-term trials. Consistently more larvae occurred under leaflets than predicted from the logistic regression; this bias diminished as T^* increased until at T^*≥40 °C, observed and predicted proportions were equal. This pattern of deviation from the predictions of the logistic regression is consistent with a thermoregulatory strategy in which larvae move away from hostile conditions, rather than seek optimal conditions.     

Differential prey utilization by the generalist predator Coleomegilla maculata lengi according to prey size and species

Prey utilization by the generalist predator Coleomegilla maculata lengi Timb. (Coleoptera: Coccinellidae) of three sympatric lepidopterous species was quantified in relation with prey size (age) and prey species. Based on optimal foraging theory, we argued that costs associated with the utilization of small and large prey are higher than those of intermediate prey size. As a result, we expected a higher prey consumption rate on intermediate prey size leading to a convex prey consumption curve. Laboratory experiments showed that, within a given prey instar, Coleomegilla maculata lengi preyed more on Plutella xylostella (L.) compared to Artogeia rapae (L.) and Trichoplusia ni (Hübner). Generally, prey consumption rate by Coleomegilla maculata lengi on the three prey species decreased with increasing immature prey size (age). The predation efficacy of Coleomegilla maculata lengi adults and fourth instar larvae was higher compared to younger coccinellids (L2). Although, Coleomegilla maculata lengi showed a higher level of predation on smaller immature prey, we demonstrated that it is not the optimal size range for this predator. As predicted, prey weight consumption rate by Coleomegilla maculata lengi was higher at intermediate prey size leading to a convex prey utilization curve. The beneficial impact of Coleomegilla maculata lengi predation on the host plant was also estimated by using a Protection Index that considers the differential predation caused by the coccinellids and the relative importance of each pest species in terms of plant injury. Coleomegilla maculata lengi has a more significant beneficial impact when it preys on T. ni immatures.     

Evaluation of dietary effects of transgenic corn pollen expressing Cry3Bb1 protein on a non-target ladybird beetle, Coleomegilla maculata

A transgenic corn event (MON 863) has been recently developed by Monsanto Company for control of corn rootworms, Diabrotica spp. (Coleoptera: Chrysomelidae). This transgenic corn event expresses the cry3Bb1 gene derived from Bacillus thuringiensis (Berliner), which encodes the insecticidal Cry3Bb1 protein for corn rootworm control. A continuous feeding study was conducted in the laboratory to evaluate the dietary effect of MON 863 pollen expressing the Cry3Bb1 protein on the survival, larval development, and reproductive capacity of the non-target species, Coleomegilla maculata DeGeer (Coleoptera: Coccinellidae). First instar C. maculata (less than 24 h old) and newly emerging adults (less than 72 h old) were fed individually on a diet mixture containing 50% of MON 863 pollen, non-transgenic (control) corn pollen, bee pollen (a component of normal rearing diet), or potassium arsenate-treated control corn pollen. In the larval tests, 96.7%, 90.0%, and 93.3% of C. maculata larvae successfully pupated and then emerged as adults when fed on MON 863 pollen, non-transgenic corn pollen, and bee pollen (normal rearing) diets, respectively. Among the larvae completing their development, there were no significant differences in the developmental time to pupation and adult emergence among the transgenic corn pollen, non-transgenic corn pollen, and bee pollen diet treatments. All larvae fed on arsenate treated corn pollen diet died as larvae. For tests with adults, 83.3%, 80.0%, and 100% of adult C. maculata survived for the 30 days of the test period when reared on diets containing 50% of MON 863 pollen, non-transgenic corn pollen, and bee pollen respectively. While the adult survival rate on MON 863 pollen diet was significantly less than that on the bee pollen diet, there was no significant difference between the MON 863 and non-transgenic corn pollen treatments. During the period of adult testing, an average of 77, 80, and 89 eggs per female were laid by females fed on the MON 863 pollen, control corn pollen, and bee pollen, respectively; no significant differences were detected in the number of eggs laid among these treatments. These results demonstrate that when offered at 50% by weight of the dietary component, transgenic corn (MON 863) pollen expressing Cry3Bb1 protein had no measurable negative effect on the survival and development of C. maculata larvae to pupation and adulthood nor any adverse effect on adult survival and reproductive capacity. Relevance of these findings to ecological impacts of transgenic Bt crops on non-target beneficial insects is discussed.     

Intraspecific mitochondrial DNA variation in the Colorado potato beetle,Leptinotarsa decemlineata (Coleoptera: Chrysomelidae)

Restriction endonuclease analyses of mitochondrial DNA (mtDNA) were used to examine genetic variability and population structure inLeptinotarsa decemlineata (Say). A group of three enzymes, EcoRI, HpaI, and PstI, was used to reveal polymorphism both within and among some of the 10 populations tested, yielding 16 haplotypes in combination. The frequencies of these 16 haplotypes differed significantly across geographic regions, indicating some partitioning of mtDNA haplotypes. Estimates of mtDNA sequence divergence (δ) between haplotypes ranged from 0.016 to 0.135%, suggesting local differentiation of mtDNA in some populations. Analysis of these data suggests that Texas was colonized by more than one mtDNA lineage, most likely originating in Mexico. We hypothesize that a larger founder size for the initial introductions or high levels of variability in the parent population at the edge of the CPB expanding range led to the initial partitioning of haplotypes observed in samples from Texas.