Pharmacist assessment of drug-gene interactions and drug-induced phenoconversion in major depressive disorder: a case report.

BACKGROUND: Response to antidepressant therapy is highly variable among individuals. Pharmacogenomic (PGx) testing presents an opportunity to guide drug selection while optimizing therapy outcomes and/or decreasing the risk for toxicity. CASE PRESENTATION: A patient with multiple comorbidities, including severe major depressive disorder (MDD), experienced adverse drug events and undesirable response to multiple antidepressant medications (i.e., bupropion, escitalopram, and venlafaxine). A clinical pharmacist assessed significant drug-gene, drug-drug, and drug-drug-gene interactions as well as other clinical factors to provide recommendations for antidepressant therapy optimization. CONCLUSION: This case highlights the importance of PGx testing and the key role of pharmacists in identifying and mitigating drug-related problems and optimizing drug therapy in patients with MDD.

The Kessler Abbreviated Psychological Distress Scale (K6) in Canadian population surveys : Report on psychometric assessment practices and analysis of the performance of several reliability coefficients.

OBJECTIVES: The objective of Study 1 is to identify the psychometric assessment and reporting practices of authors who have analyzed data from the Kessler Abbreviated Psychological Distress Scale (K6), collected in Canadian population surveys. The goal of Study 2 is to compare the performance of six reliability coefficients estimated from K6 data. METHODS: In Study 1, 71 publications using the K6 were reviewed and synthesized using an analysis grid. In Study 2, analyses were performed to compare the performance of the alpha coefficient to five other reliability coefficients using data from the 2009-2010 and 2013-2014 Canadian Community Health Survey (CCHS). Specifically, we estimated all six coefficient values, as well as their confidence intervals, regarding all respondents and respondent subgroups. RESULTS: Out of the 71 publications identified in Study 1, only nine reported a reliability coefficient drawn from their own sample. Even though no condition essential to use of the alpha coefficient was mentioned, it was the only coefficient presented. In Study 2, the values of all the other coefficients were found to be higher than those of the alpha coefficient. Significant variations were found in some respondent subgroups. CONCLUSION: Existing recommendations for the use of reliability coefficients are poorly implemented. It behooves authors to provide more information in their manuscripts, thereby enabling assessment of the psychometric qualities of the K6. The presentation of reliability coefficients for relevant subgroups and confidence intervals must also become standard practice, so that results can be more precisely interpreted.

How societal responses to COVID-19 could contribute to child neglect.

BACKGROUND: The ecosystemic approach to children's needs demands a cohesive response from societies, communities, and families. During the COVID-19 pandemic, the choices societies made to protect their community members from the virus could have created contexts of child neglect. With the closure of services and institutions, societies were no longer available to help meet the needs of children. OBJECTIVE: The purpose of this study is to examine parents' reports on the response their children received to their needs during the COVID-19 crisis. METHODS: During the period of the spring 2020 lockdown, 414 parents in the province of Quebec, Canada, completed an online questionnaire about the impact of the crisis on the response their children received to their needs. RESULTS: Compared to parents of younger children, parents of older children reported less fulfillment of their child's needs in three measured domains, namely cognitive and affective, security, and basic care needs. CONCLUSION: These results are discussed in light of the policies and the resources societies have put in place during the crisis to help families meet the needs of their children. Societies must learn from this crisis to put children at the top of their priorities in the face of a societal crisis. Thoughtful discussions and energy must be given to ensure that, while facing a crisis, the developmental trajectories of children are not sacrificed.

Anti-LG3 Antibodies Aggravate Renal Ischemia-Reperfusion Injury and Long-Term Renal Allograft Dysfunction.

Pretransplant autoantibodies to LG3 and angiotensin II type 1 receptors (AT1R) are associated with acute rejection in kidney transplant recipients, whereas antivimentin autoantibodies participate in heart transplant rejection. Ischemia-reperfusion injury (IRI) can modify self-antigenic targets. We hypothesized that ischemia-reperfusion creates permissive conditions for autoantibodies to interact with their antigenic targets and leads to enhanced renal damage and dysfunction. In 172 kidney transplant recipients, we found that pretransplant anti-LG3 antibodies were associated with an increased risk of delayed graft function (DGF). Pretransplant anti-LG3 antibodies are inversely associated with graft function at 1 year after transplantation in patients who experienced DGF, independent of rejection. Pretransplant anti-AT1R and antivimentin were not associated with DGF or its functional outcome. In a model of renal IRI in mice, passive transfer of anti-LG3 IgG led to enhanced dysfunction and microvascular injury compared with passive transfer with control IgG. Passive transfer of anti-LG3 antibodies also favored intrarenal microvascular complement activation, microvascular rarefaction and fibrosis after IRI. Our results suggest that anti-LG3 antibodies are novel aggravating factors for renal IRI. These results provide novel insights into the pathways that modulate the severity of renal injury at the time of transplantation and their impact on long-term outcomes.

No effects of pantoprazole on the pharmacokinetics of rosuvastatin in healthy subjects.

PURPOSE: Rosuvastatin disposition is modulated by the expression and activity of several membrane transporters including BCRP (ABCG2). The objective of our study was to investigate the effects of pantoprazole, a previously proposed BCRP inhibitor, on the disposition of rosuvastatin. METHODS: The impact of pantoprazole (40 mg ID for 2 days) on rosuvastatin pharmacokinetics was evaluated in healthy volunteers (n = 16) who received a single oral dose of rosuvastatin (10 mg) either alone or with pantoprazole. Rosuvastatin, N-desmethylrosuvastatin, and rosuvastatin lactone levels were quantified in plasma while rosuvastatin and N-desmethylrosuvastatin excretion were measured in urine. RESULTS: Ratios and 90 % standard confidence interval of geometric means for C max (1.03 [0.91-1.16]), AUC0-∞ (1.03 [0.89-1.19]) and renal clearance (0.96 [0.85-1.09]) were all within the pre-specified range of 0.8-1.25, indicating a lack of drug-drug interaction between pantoprazole and rosuvastatin. CONCLUSIONS: Concomitant administration of pantoprazole with rosuvastatin did not affect rosuvastatin plasma concentrations. The use of pantoprazole as a BCRP inhibitor should be revisited when characterizing BCRP-mediated transport in humans.

Interpreting the flock algorithm: a reply to Anderson & Barry (2015).

Anderson & Barry (Molecular Ecology Resources, 2015, 10, 1020-1030) compared a reprogrammed version of flock (Duchesne & Turgeon , Molecular Ecology Resources, 2009, 9, 1333-1344), flockture, to a particular model of structure (Pritchard , Genetics, 2000, 155, 945-959) that they propose is equivalent to flock, a non-MCMC, non-Bayesian algorithm. They conclude that structure performs better than flockture at clustering individuals from simulated populations with very low level of differentiation (FST c. 0.008) based on 15 microsatellites or 96 SNPs. We rather consider that both algorithms failed, with proportions of correct allocations lower than 50%. The authors also noted the slightly better performance of flockture with SNPs at intermediate FST values (c. 0.02-0.04) but did not comment. Finally, we disagree with the way the processing time of each program was compared. When compared on the basis of a run leading to a clustering solution, the main output of any clustering algorithm, flock, is, as users can readily experience, much faster. In all, we feel that flock performs at least as well as structure as a clustering algorithm. Moreover, flock has two major assets: high speed and clear, well validated, rules to estimate K, the number of populations. It thus provides a valuable addition to the set of tools at the disposal of the many researchers dealing with real empirical data sets.

The combined effects of pre- and post-copulatory processes are masking sexual conflict over mating rate in Gerris buenoi.

In polygynandrous animals, post-copulatory processes likely interfere with precopulatory sexual selection. In water striders, sexual conflict over mating rate and post-copulatory processes are well documented, but their combined effect on reproductive success has seldom been investigated. We combine genetic parentage analyses and behavioural observations conducted in a competitive reproductive environment to investigate how pre- and post-copulatory processes influence reproductive success in Gerris buenoi Kirkaldy. Precopulatory struggles had antagonistic effects on male and female reproductive success: efficiently gaining copulations was beneficial for males, whereas efficiently avoiding copulations was profitable for females. Also, high mating rates and an intermediate optimal resistance level of females supported the hypothesis of convenience polyandry. Contrary to formal predictions, high mating rates (i.e. the number of copulations) did not increase reproductive success in males or decrease reproductive success in females. Instead, the reproductive success of both sexes was higher when offspring were produced with several partners and when there were few unnecessary matings. Thus, male and female G. buenoi displayed different interests in reproduction, but post-copulatory processes were masking the effects of copulatory mating success on reproductive success. Given the high mating rates observed, sperm competition could easily counter the effect of mating rates, perhaps in interaction with cryptic female choice and/or fecundity selection. Our study presents a complex but realistic overview of sexual selection forces at work in a model organism for the study of sexual conflict, confirming that insights are gained from investigating all episodes in the reproduction cycle of polygynandrous animals.

Natural enemies associated with Sirex noctilio (Hymenoptera: Siricidae) and S. nigricornis in Ontario, Canada.

Sirex noctilio F. is an exotic woodwasp now found in eastern North America where it shares natural enemies with native woodwasps of Pinus spp. To study the extent to which native hymenopteran parasitoids and parasitic nematodes could affect woodwasp populations, 60 Pinus trees with symptoms of S. noctilio attack were felled in 2007 and 2008 in Ontario, Canada. Each tree bole was cut into 1-m sections that were placed in individual rearing tubes; emergence was monitored from May to November of the year of felling. Female S. noctilio were dissected to assess parasitism by the nematode Deladenus siricidicola Bedding. Two species of Siricidae emerged from these trees; S. noctilio, which accounted for most of the specimens collected, and S. nigricornis F. Of the three species of parasitoid that emerged, Ibalia leucospoides (Hochenwarth) was the most abundant, accounting for an overall hypothetical Siricidae parasitism rate of almost 20%. This parasitoid emerged over a similar time period as S. noctilio-between early July and early September. Except in trees >15 m in height, parasitism by I. leucospoides generally appeared uniform throughout the bole. Parasitism rates did not vary between the 2 yr, but did between sites in 1 yr. Parasitic nematodes were found in the haemocoel of about one third of S. noctilio females dissected but were never found sterilizing the eggs; none were found in S. noctilio emerging from P. resinosa. These findings suggest that I. leucospoides is currently the primary invertebrate natural enemy of S. noctilio in Ontario.

[Focal dermal hypoplasia (Goltz syndrome) in the neonate: report of a case presenting with cleft lip and palate]. / Syndrome de Goltz chez un nouveau-né avec fente labiopalatine.

Focal dermal hypoplasia (Goltz syndrome) is a rare congenital dysplasia of the mesoectodermal derived tissues. It is an X-linked inheritance syndrome caused by mutations in the PORCNgene mapped on Xp11.23. The condition is characterized by cutaneous lesions distributed in linear areas associated with diverse congenital deformities. Given the rarely described neonatal features, we report a case of Goltz syndrome in a black female newborn. This is the first case known in Burkina Faso. The cutaneous, hair, and nail lesions usually observed were present, characterized by their preponderance on the left side of the body with exclusive ipsilateral skeletal abnormalities (cleft lip and palate, agenesis of the metatarsals and toes of the foot, syndactyly, lobster claw, and absence of a rib). The limits in the management and the negative social and cultural perception of the deformities in the context of a developing country did not favor the child's survival.

A prospective pilot study investigating the musculoskeletal pain in postmenopausal breast cancer patients receiving aromatase inhibitor therapy.

BACKGROUND: Although arthralgia is a known adverse effect of aromatase inhibitor (ai) treatment in postmenopausal breast cancer patients, few studies have carried out a comprehensive evaluation of the nature, onset, and incidence of musculoskeletal (msk) pain in these patients. We therefore used a pilot study to identify conditions or markers predictive of pain. METHODS: For 24 weeks, we monitored 30 eligible postmenopausal women starting ai therapy. Pre-existing and incident msk conditions and pain were assessed clinically and with ultrasonography of the hands and wrists. In addition, patient questionnaires were used to assess pain before and during ai therapy. Biochemical markers were measured at baseline and at regular intervals after anastrozole therapy began. Gene profiling studies were carried out before and 48 hours after the initial ai administration. RESULTS: Over the 24-week study period, 20 participants (67%) showed no pain symptoms; 5 (17%) experienced low or moderate pain at baseline, which did not increase with ai treatment; and during therapy, 5 (17%) showed exacerbation of pain attributable to osteoarthritis of the hand and to finger flexor tenosynovitis. Although all 30 participants had some degree of msk conditions before anastrozole therapy started, the pre-existing conditions did not necessarily predispose the women to increased pain during anastrozole treatment. Higher levels of urinary N-telopeptides of type i collagen were associated with the groups presenting pain, suggesting a higher extent of pre-existing bone resorption, without significant evolution over the 24-week treatment period. Slightly higher levels of 1,25(OH)(2) vitamin D(3) were observed at baseline in patients with pain increase, but did not significantly change during treatment; however, average levels of 25(OH) vitamin D(3) increased, likely because of supplementation. Although biochemical markers did not discriminate efficiently between pain groups, a signature of 166 genes in peripheral blood mononuclear cells was identified that could stratify patients into the various groups observed in this pilot study. The gene signature was enriched in components of inflammatory signalling and chemokine expression, of antitumoural immunity pathways, and of metabolic response to hormones and xenobiotics, although no clinically significant association could be made in the present study, considering the small number of patients. Nevertheless, the observed trend suggests the feasibility of developing surrogate predictive markers of msk pain. Patient compliance was high in this study and was not affected by pain exacerbation. CONCLUSIONS: Baseline msk assessment showed pre-existing causes for pain in most of the study patients before initiation of the ai. Exacerbation of existing osteoarthritis pain and tenosynovial symptoms was the primary cause of pain increase. Musculoskeletal pain assessment at baseline and prompt treatment of pain symptoms may help to optimize adherence to ai therapy. The value of routinely assessing inflammatory markers such as C-reactive protein and erythrocyte sedimentation rate was not supported by our pilot study. Gene expression profiles in peripheral blood mononuclear cells may be further explored in larger-scale studies as stratification markers to identify patients at risk of developing arthralgia.

Sexual conflict in Gerris gillettei (Insecta: Hemiptera): intraspecific intersexual correlated morphology and experimental assessment of behaviour and fitness.

The contemporary dynamics of sexually antagonistic coevolution caused by sexual conflicts have seldom been investigated at the intraspecific level. We characterized natural populations of Gerris gillettei and documented significant intersexual correlations for morphological traits previously related to sexual conflict in water striders. These results strongly indicate that sexually antagonistic coevolution contributed to population differentiation and resulted in different balances of armaments between the sexes within natural populations of this species. No-choice mating experiments further revealed that both male and male-female relative arms levels influence copulation duration. However, there were no asymmetries in reproductive behaviour and fitness between sympatric and allopatric mating pairs, suggesting that differentiation by sexual conflict was not sufficient to influence the outcome of mating interactions. Altogether, these results question the relative importance of female connexival spines vs. genitalia traits in mediating pre- and post-copulatory conflict in Gerris.

Experimental evidence for the phenotypic impact of admixture between wild and biocontrol Asian ladybird (Harmonia axyridis) involved in the European invasion.

Hybridization can fuel evolutionary processes during biological invasions. The harlequin ladybird Harmonia axyridis has long been used as a biocontrol agent before the species became invasive worldwide. Previous analysis based on microsatellite data has shown that European invasive populations bear traces of admixture between an eastern North American source, which is at the origin of the worldwide invasion, and biocontrol strains used in Europe. In this study, we tested the hypothesis that this early admixture event may have fostered the European invasion by impacting on the phenotypes of wild European populations. Mean life history traits of experimental F(1) hybrids are compared with pure parental sources and wild European crosses. Our results reveal a biased impact whereby North American beetles benefitted from being admixed with European biocontrol strains. Resemblance between experimental hybrids and wild European invasive crosses further suggests a long-lasting effect of admixture that may still be at work and fostering invasiveness.

The pharmacokinetics of a long-acting OROS hydromorphone formulation.

IMPORTANCE OF THE FIELD: New formulations of opiods can provide round-the-clock pain relief to improve pain management and quality of life for patients with chronic pain. AREAS COVERED IN THIS REVIEW: Information and comments on the pharmacokinetic processes associated with a new once-daily formulation of the potent opiod hydromorphone. WHAT THE READER WILL GAIN: This review presents an overview of data from several small pharmacokinetic studies to gain a better perspective on the pharmacokinetic properties of a new long-acting formulation of hydromorphone. TAKE HOME MESSAGE: The development of advanced oral formulation that deliver analgesic drugs over an extended period provides new solutions to improve pain management and quality of life for patients with chronic pain.

Diversity and distribution of gynogenetic hybrids between Fundulus diaphanus and Fundulus heteroclitus in Porter's Lake (Nova Scotia) in relation to salinity.

Unisexual hybrids between Fundulus diaphanus and Fundulus heteroclitus were found in low proportions at intermediate salinity in Porter's Lake, Nova Scotia. One clone accounted for 72% of the hybrids, with most other hybrids being different at a single microsatellite allele. This clone thrives over a wide range of salinities, suggesting a general-purpose genotype.

FLOCK: a method for quick mapping of admixture without source samples.

Identifying and estimating individual and/or population admixture is a very common objective in evolution and conservation biology. There are many situations where samples from one or many of the putatively hybridizing entities are not available or easily identified. Here we describe FLOCK, a new method especially designed to provide spatial and/or temporal admixture maps in the absence of one or several source samples. FLOCK is a non-Bayesian method and therefore differs substantially from previous clustering algorithms. Its working principle is repeated re-allocation of all collected specimens (total sample) to the k subsamples, each re-allocation being more effective than the previous one in attracting genetically similar individuals. This snowball effect, more formally referred to as a positive feedback mechanism, makes FLOCK an efficient and quick sorting process. The usage of FLOCK is illustrated with two empirical situations which have been thoroughly analysed previously with other approaches. A number of simulations were run to better assess the power of the FLOCK algorithm. Performance comparisons were made between the FLOCK and Structure algorithms. When non-negligible numbers of pure genotypes were present, the two performed equally well. However, FLOCK proved significantly more powerful in the absence of pure genotypes. Moreover, FLOCK showed more potential for fast processing. Run times were shown to increase linearly with size of total sample and with size of k, the number of reference samples from which admixture mapping is performed.

Combination of phenotype assessments and CYP2C9-VKORC1 polymorphisms in the determination of warfarin dose requirements in heavily medicated patients.

The relative contribution of phenotypic measures and CYP2C9-vitamin K epoxide reductase complex subunit 1 (VKORC1) polymorphisms to warfarin dose requirements at day 14 was determined in 132 hospitalized, heavily medicated patients. Phenotypic measures were (1) the urinary losartan metabolic ratio before the first dose of warfarin, (2) the S:R-warfarin ratio at day 1, and (3) a dose-adjusted international normalized ratio (INR) at day 4. CYP2C9 and VKORC1 genotypes were determined by gene chip analysis. In multivariate analyses, the dose-adjusted INR at day 4 explained 31% of variability observed in warfarin doses at day 14, whereas genotypic measures (CYP2C9-VKORC1) contributed 6.5%. When S:R-warfarin ratio was used, genotypes contributed more significantly (23.5%). Finally, urinary losartan metabolic ratio was of low predictive value. The best models obtained explained 51% of intersubject variability in warfarin dose requirements. Thus, combination of a phenotypic measure to CYP2C9-VKORC1 genotypes represents a useful strategy to predict warfarin doses in patients receiving multiple drugs (11+/-4 drugs/day).

Identification of the cytochrome P450 enzymes involved in the metabolism of domperidone.

The objective was to identify the major cytochrome P450 enzyme(s) involved in the metabolism of domperidone. Experiments were performed using human liver microsomes (HLMs), recombinant human cytochrome P450 enzymes, cytochrome P450 chemical inhibitors and monoclonal antibodies directed against cytochrome P450 enzymes. Four metabolites were identified from incubations performed with HLMs and excellent correlations were observed between the formation of domperidone hydroxylated metabolites (M1, M3 and M4), N-desalkylated domperidone metabolite (M2) and enzymatic markers of CYP3A4/5 (r2 = 0.9427, 0.951, 0.9497 and 0.8304, respectively). Ketoconazole (1 microM) decreased the formation rate of M1, M2, M3 and M4 by 83, 78, 75 and 88%, respectively, whereas the effect of other inhibitors (quinidine, furafylline and sulfaphenazole) was minimal. Important decreases in the formation rate of M1 (68%), M2 (64%) and M3 (54%) were observed with anti-CYP3A4 antibodies. Formation of M1, M2 and M3 in HLMs exhibited Michaelis-Menten kinetics (Km: 166, 248 and 36 microM, respectively). Similar Km values were observed for M1, M2 and M3 when incubations were performed with recombinant human CYP3A4 (Km: 107, 273 and 34 microM, respectively). The data suggest that CYP3As are the major enzymes involved in the metabolism of domperidone.

Comparative efficacy of terbutaline sulphate delivered by Turbuhaler dry powder inhaler or pressurised metered dose inhaler with Nebuhaler spacer in children during an acute asthmatic episode.

AIMS: To compare the efficacy of terbutaline sulphate delivered via Turbuhaler with a pressurised metered dose inhaler (pMDI) connected to Nebuhaler spacer in a population of asthmatic children presenting to emergency departments because of an acute episode of asthma. METHODS: Randomised double blind, double dummy, parallel study of acute asthma in the emergency department. A total of 112 children (6-16 years), who had a diagnosis of asthma, a baseline FEV1 of 25-60% of predicted normal value (PNV), and the ability to perform spirometry were studied. Patients received two doses of 0.5 mg/10 kg (maximum 2.0 mg) of terbutaline sulphate at time 0 minutes and time 30 minutes. The two groups were also stratified into subgroups based on FEV1: 25-45% and 45.1-60% PNV. FEV1 before treatment and at two 15-minute intervals after each treatment was the main outcome measure. PIF, PEF, heart rate, SpO2, and tremor were also measured at these times. RESULTS: Both the Turbuhaler and pMDI+Nebuhaler groups showed significant increases from baseline to final value in their FEV1 results, 49% and 50% change from baseline to t = 60 min, respectively (p < 0.001) using last value carried forward. No significant difference was found between the two groups for these results. Subanalysis of the stratified groups revealed similar results. In addition, no significant difference was found in the group and subgroup comparisons for heart rate, SpO2, and tremor. CONCLUSION: Results show that Turbuhaler and pMDI+Nebuhaler are similar in terms of benefit and side effects in the treatment of acute moderate to severe asthma attacks in this study population.

Role of specific cytochrome P450 enzymes in the N-oxidation of the antiarrhythmic agent mexiletine.

1. Mexiletine is extensively metabolized in man by C- and N-oxidation and the aim of the present study was to characterize major cytochrome P450 enzyme(s) involved in the formation of N-hydroxymexiletine. 2. Incubations with genetically engineered microsomes indicated that the formation rate of N-hydroxymexiletine was highest in the presence of microsomes expressing high levels of either CYP1A2 or CYP2E1 and the formation of N-hydroxymexiletine by human liver microsomes was inhibited about 40% by antibodies directed against CYP1A1/1A2 or CYP2E1. Additional incubations demonstrated that formation of N-hydroxymexiletine was decreased 47 and 51% by furafylline, 40 microm and 120 microm, respectively, and decreased 55 and 67% by alpha-naphthoflavone, 1 microm and 3 microm, respectively (all p < 0.05 versus control). 3. The formation rate of N-hydroxymexiletine in human liver microsomes was highly correlated with CYP2B6 (RS-mexiletine, r = 0.7827; R-(-)-enantiomer, r = 0.7034; S-(+)-enantiomer, r = 0.7495), CYP2E1 (S-(+)-enantiomer, r = 0.7057) and CYP1A2 (RS-mexiletine, r = 0.5334; S-(+)-enantiomer, r = 0.6035). 4. In conclusion, we have demonstrated that CYP1A2 is a major human cytochrome P450 enzyme involved in the formation of N-hydroxymexiletine. However, other cytochrome P450 enzymes (CYP2E1 and CYP2B6) also appear to play a role in the N-oxidation of this drug.

Phylogeographic analysis of a recent radiation of Enallagma damselflies (Odonata: Coenagrionidae).

A phylogenetic hypothesis revealed two recent radiations among species of Enallagma damselflies, and extensive ecological work suggests that both adaptive and nonadaptive processes are involved in these radiations. We analysed the geographical pattern of genetic variability at 868 bp of mitochondrial DNA (mtDNA) among 283 individuals of 5 species displaying little ecological differentiation to identify the ancestral lineage, support their independent evolutionary trajectories and identify historical events and the underlying mechanism for one of these radiations. Nested clade analysis results clearly support a past event of range fragmentation in E. hageni. These Atlantic and Continental hageni races experienced distinct dispersal histories and still maintain nearly nonoverlapping ranges All four other species derive from the Continental hageni. Whereas three species endemic to the Atlantic coastal plain show little genetic variation, E. ebrium shared several haplotypes with the Continental hageni. Contrasting levels of genetic differentiation between E. hageni and E. ebrium in geographical areas associated with distinct events of E. hageni's recent history support the recent origin of this species. Altogether, our results are compatible with a process of radiation via divergence in mate recognition systems within the Continental hageni race following secondary contacts between putative refugial races.