Receptor activator for NF-κB ligand in acute myeloid leukemia: expression, function, and modulation of NK cell immunosurveillance.

The TNF family member receptor activator for NF-κB ligand (RANKL) and its receptors RANK and osteoprotegerin are key regulators of bone remodeling but also influence cellular functions of tumor and immune effector cells. In this work, we studied the involvement of RANK-RANKL interaction in NK cell-mediated immunosurveillance of acute myeloid leukemia (AML). Substantial levels of RANKL were found to be expressed on leukemia cells in 53 of 78 (68%) investigated patients. Signaling via RANKL into the leukemia cells stimulated their metabolic activity and induced the release of cytokines involved in AML pathophysiology. In addition, the immunomodulatory factors released by AML cells upon RANKL signaling impaired the anti-leukemia reactivity of NK cells and induced RANK expression, and NK cells of AML patients displayed significantly upregulated RANK expression compared with healthy controls. Treatment of AML cells with the clinically available RANKL Ab Denosumab resulted in enhanced NK cell anti-leukemia reactivity. This was due to both blockade of the release of NK-inhibitory factors by AML cells and prevention of RANK signaling into NK cells. The latter was found to directly impair NK anti-leukemia reactivity with a more pronounced effect on IFN-γ production compared with cytotoxicity. Together, our data unravel a previously unknown function of the RANK-RANKL molecule system in AML pathophysiology as well as NK cell function and suggest that neutralization of RANKL with therapeutic Abs may serve to reinforce NK cell reactivity in leukemia patients.

First imaging results of an intraindividual comparison of (11)C-acetate and (18)F-fluorocholine PET/CT in patients with prostate cancer at early biochemical first or second relapse after prostatectomy or radiotherapy.

PURPOSE: (18)F-Fluorocholine (FCH) and (11)C-acetate (ACE) PET are widely used for detection of recurrent prostate cancer (PC). We present the first results of a comparative, prospective PET/CT study of both tracers evaluated in the same patients presenting with recurrence and low PSA to compare the diagnostic information provided by the two tracers. METHODS: The study group comprised 23 patients studied for a rising PSA level after radical prostatectomy (RP, 7 patients, PSA ≤ 3 ng/ml), curative radiotherapy (RT, 7 patients, PSA ≤ 5 ng/ml) or RP and salvage RT (9 patients, PSA ≤ 5 ng/ml). Both FCH and ACE PET/CT scans were performed in a random sequence a median of 4 days (range 0 to 11 days) apart. FCH PET/CT was started at injection (307 ± 16 MBq) with a 10-min dynamic acquisition of the prostate bed, followed by a whole-body PET scan and late (45 min) imaging of the pelvis. ACE PET/CT was performed as a double whole-body PET scan starting 5 and 22 min after injection (994 ± 72 MBq), and a late view (45 min) of the prostate bed. PET/CT scans were blindly reviewed by two independent pairs of two experienced nuclear medicine physicians, discordant subgroup results being discussed to reach a consensus for positive, negative end equivocal results. RESULTS: PET results were concordant in 88 out of 92 local, regional and distant findings (Cohen's kappa 0.929). In particular, results were concordant in all patients concerning local status, bone metastases and distant findings. Lymph-node results were concordant in 19 patients and different in 4 patients. On a per-patient basis results were concordant in 22 of 23 patients (14 positive, 5 negative and 3 equivocal). In only one patient was ACE PET/CT positive for nodal metastases while FCH PET/CT was overall negative; interestingly, the ACE-positive and FCH-negative lymph nodes became positive in a second FCH PET/CT scan performed a few months later. CONCLUSION: Overall, ACE and FCH PET/CT showed excellent concordance, on both a per-lesion and a per-patient basis, suggesting that both tracers perform equally for recurrent prostate cancer staging.

What can we learn from international comparisons of costs by DRG ?

OBJECTIVES: The objective of this study was to compare costs data by diagnosis related group (DRG) between Belgium and Switzerland. Our hypotheses were that differences between countries can probably be explained by methodological differences in cost calculations, by differences in medical practices and by differences in cost structures within the two countries. METHODS: Classifications of DRG used in the two countries differ (AP-DRGs version 1.7 in Switzerland and APR-DRGs version 15.0 in Belgium). The first step of this study was to transform Belgian summaries into Swiss AP-DRGs. Belgian and Swiss data were calculated with a clinical costing methodology (full costing). Belgian and Swiss costs were converted into US$ PPP (purchasing power parity) in order to neutralize differences in purchasing power between countries. RESULTS: The results of this study showed higher costs in Switzerland despite standardization of cost data according to PPP. The difference is not explained by the case-mix index because this was similar for inliers between the two countries. The length of stay (LOS) was also quite similar for inliers between the two countries. The case-mix index was, however, higher for high outliers in Belgium, as reflected in a higher LOS for these patients. Higher costs in Switzerland are thus probably explained mainly by the higher number of agency staff by service in this country or because of differences in medical practices. CONCLUSIONS: It is possible to make international comparisons but only if there is standardization of the case-mix between countries and only if comparable accountancy methodologies are used. Harmonization of DRGs groups, nomenclature and accountancy is thus required.

Atmospheric deposition and migration of artificial radionuclides in Alpine soils (Val Piora, Switzerland) compared to the distribution of selected major and trace elements.

Artificial radionuclides ((137)Cs, (90)Sr, Pu, and (241)Am) are present in soils because of Nuclear Weapon Tests and accidents in nuclear facilities. Their distribution in soil depth varies according to soil characteristics, their own chemical properties, and their deposition history. For this project, we studied the atmospheric deposition of (137)Cs, (90)Sr, Pu, (241)Am, (210)Pb, and stable Pb. We compared the distribution of these elements in soil profiles from different soil types from an alpine Valley (Val Piora, Switzerland) with the distribution of selected major and trace elements in the same soils. Our goals were to explain the distribution of the radioisotopes as a function of soil parameters and to identify stable elements with analogous behaviors. We found that Pu and (241)Am are relatively immobile and accumulate in the topsoil. In all soils, (90)Sr is more mobile and shows some accumulations at depth into Fe-Al rich horizons. This behavior is also observed for Cu and Zn, indicating that these elements may be used as chemical analogues for the migration of (90)Sr into the soil.

Roles of astrocytic sodium dynamics in the neurometabolic coupling : a study by cellular imaging

RESUME LARGE PUBLIC Le système nerveux central est principalement composé de deux types de cellules :les neurones et les cellules gliales. Ces dernières, bien que l'emportant en nombre sur les neurones, ont longtemps été considérées comme des cellules sans intérêts par les neuroscientifiques. Hors, les connaissances modernes à leurs sujets indiquent qu'elles participent à la plupart des tâches physiologiques du cerveau. Plus particulièrement, elles prennent part aux processus énergétiques cérébraux. Ceux-ci, en plus d'être vitaux, sont particulièrement intrigants puisque le cerveau représente seulement 2 % de la masse corporelle mais consomme environ 25 % du glucose (substrat énergétique) corporel. Les astrocytes, un type de cellules gliales, jouent un rôle primordial dans cette formidable utilisation de glucose par le cerveau. En effet, l'activité neuronale (transmission de l'influx nerveux) est accompagnée d'une augmentation de la capture de glucose, issu de la circulation sanguine, par les astrocytes. Ce phénomène est appelé le «couplage neurométabolique » entre neurones et astrocytes. L'ion sodium fait partie des mécanismes cellulaires entrant en fonction lors de ces processus. Ainsi, dans le cadre de cette thèse, les aspects dynamiques de la régulation du sodium astrocytaire et leurs implications dans le couplage neurométabolique ont été étudiés par des techniques d'imagerie cellulaires. Ces études ont démontré que les mitochondries, machineries cellulaires convertissant l'énergie contenue dans le glucose, participent à la régulation du sodium astrocytaire. De plus, ce travail de thèse a permis de découvrir que les astrocytes sont capables de se transmettre, sous forme de vagues de sodium se propageant de cellules en cellules, un message donnant l'ordre d'accroître leur consommation d'énergie. Cette voie de signalisation leur permettrait de fournir de l'énergie aux neurones suite à leur activation. RESUME Le glutamate libéré dans la fente synaptique pendant l'activité neuronale, est éliminé par les astrocytes environnants. Le glutamate est co-transporté avec des ions sodiques, induisant une augmentation intracellulaire de sodium (Na+i) dans les astrocytes. Cette élévation de Na+i déclenche une cascade de mécanismes moléculaires qui aboutissent à la production de substrats énergétiques pouvant être utilisés par les neurones. Durant cette thèse, la mesure simultanée du sodium mitochondrial (Na+mit) et cytosolique par des techniques d'imagerie utilisant des sondes fluorescentes spécifiques, a indiqué que les variations de Na+i induites par le transport du glutamate sont transmises aux mitochondries. De plus, les voies d'entrée et de sortie du sodium mitochondrial ont été identifiées. L'échangeur de Na+ et de Ca2+ mitochondrial semble jouer un rôle primordial dans l'influx de Na+mit, alors que l'efflux de Na+mit est pris en charge par l'échangeur de Na+ et de H+ mitochondrial. L'étude du Na+mit a nécessité l'utilisation d'un système de photoactivation. Les sources de lumière ultraviolette (UV) classiques utilisées à cet effet (lasers, lampes à flash) ayant plusieurs désavantages, une alternative efficace et peu coûteuse a été développée. Il s'agit d'un système compact utilisant une diode électroluminescente (LED) à haute puissance et de longueur d'onde de 365nm. En plus de leurs rôles dans le couplage neurométabolique, les astrocytes participent à la signalisation multicellulaire en transmettant des vagues intercellulaires de calcium. Ce travail de thèse démontre également que des vagues intercellulaires de sodium peuvent être évoquées en parallèle à ces vagues calciques. Le glutamate, suite à sa libération par un mécanisme dépendent du calcium, est réabsorbé par les transporteurs au glutamate. Ce mécanisme a pour conséquence la génération de vagues sodiques se propageant de cellules en cellules. De plus, ces vagues sodiques sont corrélées spatialement avec une consommation accrue de glucose par les astrocytes. En conclusion, ce travail de thèse a permis de montrer que le signal sodique astrocytaire, déclenché en réponse au glutamate, se propage à la fois de façon intracellulaire aux mitochondries et de façon intercellulaire. Ces résultats suggèrent que les astrocytes fonctionnent comme un réseau de cellules nécessaire au couplage énergétique concerté entre neurones et astrocytes et que le sodium est un élément clé dans les mécanismes de signalisations cellulaires sous-jacents. SUMMARY Glutamate, released in the synaptic cleft during neuronal activity, is removed by surrounding astrocytes. Glutamate is taken-up with Na+ ions by specific transporters, inducing an intracellular Na+ (Na+i) elevation in astrocytes which triggers a cascade of molecular mechanisms that provides metabolic substrates to neurons. Thus, astrocytic Na+i homeostasis represents a key component of the so-called neurometabolic coupling. In this context, the first part of this thesis work was aimed at investigating whether cytosolic Na+ changes are transmitted to mitochondria, which could therefore influence their function and contribute to the overall intracellular Na+ regulation. Simultaneous monitoring of both mitochondrial Na+ (Na+mit) and cytosolic Na+ changes with fluorescent dyes revealed that glutamate-evoked cytosolic Na+ elevations are indeed transmitted to mitochondria. The mitochondrial Na+/Ca2+ exchangers have a prominent role in the regulation of Na+mit influx pathway, and Na+mit extrusion appears to be mediated by Na+/H+ exchangers. To demonstrate the implication of Na+/Ca2+ exchangers, this study has required the technical development of an UV-flash photolysis system. Because light sources for flash photolysis have to be powerful and in the near UV range, the use of UV lasers or flash lamps is usually required. As an alternative to these UV sources that have several drawbaks, we developped a compact, efficient and lowcost flash photolysis system which employs a high power 365nm light emitting diode. In addition to their role in neurometabolic coupling, astrocytes participate in multicellular signaling by transmitting intercellular Ca2+ waves. The third part of this thesis show that intercellular Na+ waves can be evoked in parallel to Ca2+ waves. Glutamate released by a Ca2+ wave-dependent mechanism is taken up by glutamate transporters, resulting in a regenerative propagation of cytosolic Na+ increases. Na+ waves in turn lead to a spatially correlated increase in glucose uptake. In conclusion, the present thesis demonstrates that glutamate-induced Na+ changes occurring in the cytosol of astrocytes propagate to both the mitochondrial matrix and the astrocytic network. These results furthermore support the view that astrocytic Na+ is a signal coupled to the brain energy metabolism.

Long-term and long-range migration of radioactive fallout in a Karst system.

Mountainous areas are often covered by little evolved soils from which deposited radionuclides can potentially leak into the vadose zone. In the Swiss Jura mountains, we observed unusual isotopic ratios of nuclear weapon test (NWTs) fallout with an apparent loss of NWTs plutonium relative to &supl;³⁷Cs of Chernobyl origin in thinner soils. Here, we studied the karstic watershed of a vauclusian spring to determine the residence times of plutonium, ²⁴&supl;Am, and ⁹⁰Sr deposited by global fallout and their respective mobility in carbonaceous soils. The results show that ⁹⁰Sr is washed most efficiently from the watershed with a residence time of several hundred years. The estimated plutonium residence time is more than 10 times higher (in the range of 5000-10,000 years), and the ²⁴&supl;Am residence time is double that of plutonium. The spring water ²⁴&supl;Am/²³⁹+²⁴⁰Pu isotopic ratio is lower (0.12 - 0.28) than found in watershed soils (0.382 ± 0.077). Similar differences are found in aquatic mosses (²⁴&supl;Am/²³⁹+²⁴⁰Pu isotopic ratio 0.05-0.12), which are permanently submerged in spring waters. In contrast to plutonium, ⁹⁰Sr is leached from these mosses with 0.5M HCl, demonstrating that strontium is probably associated with calcium carbonate precipitations on the mosses. The higher plutonium to americium isotopic ratio found in the samples of spring water and mosses at the outlet of the karst shows that plutonium mobility is enhanced.

An overview of the metabolic differences between Bradyrhizobium japonicum 110 bacteria and differentiated bacteroids from soybean (Glycine max) root nodules: an in vitro 13C- and 31P-nuclear magnetic resonance spectroscopy study.

Bradyrhizobium japonicum is a symbiotic nitrogen-fixing soil bacteria that induce root nodules formation in legume soybean (Glycine max.). Using (13)C- and (31)P-nuclear magnetic resonance (NMR) spectroscopy, we have analysed the metabolite profiles of cultivated B. japonicum cells and bacteroids isolated from soybean nodules. Our results revealed some quantitative and qualitative differences between the metabolite profiles of bacteroids and their vegetative state. This includes in bacteroids a huge accumulation of soluble carbohydrates such as trehalose, glutamate, myo-inositol and homospermidine as well as Pi, nucleotide pools and intermediates of the primary carbon metabolism. Using this novel approach, these data show that most of the compounds detected in bacteroids reflect the metabolic adaptation of rhizobia to the surrounding microenvironment with its host plant cells.

The role of interval nodes in sentinel lymph node mapping and dissection for melanoma patients.

In sentinel node (SN) biopsy, an interval SN is defined as a lymph node or group of lymph nodes located between the primary melanoma and an anatomically well-defined lymph node group directly draining the skin. As shown in previous reports, these interval SNs seem to be at the same metastatic risk as are SNs in the usual, classic areas. This study aimed to review the incidence, lymphatic anatomy, and metastatic risk of interval SNs. METHODS: SN biopsy was performed at a tertiary center by a single surgical team on a cohort of 402 consecutive patients with primary melanoma. The triple technique of localization was used-that is, lymphoscintigraphy, blue dye, and gamma-probe. Otolaryngologic melanoma and mucosal melanoma were excluded from this analysis. SNs were examined by serial sectioning and immunohistochemistry. All patients with metastatic SNs were recommended to undergo a radical selective lymph node dissection. RESULTS: The primary locations of the melanomas included the trunk (188), an upper limb (67), or a lower limb (147). Overall, 97 (24.1%) of the 402 SNs were metastatic. Interval SNs were observed in 18 patients, in all but 2 of whom classic SNs were also found. The location of the primary was truncal in 11 (61%) of the 18, upper limb in 5, and lower limb in 2. One patient with a dorsal melanoma had drainage exclusively in a cervicoscapular area that was shown on removal to contain not lymph node tissue but only a blue lymph channel without tumor cells. Apart from the interval SN, 13 patients had 1 classic SN area and 3 patients 2 classic SN areas. Of the 18 patients, 2 had at least 1 metastatic interval SN and 2 had a classic SN that was metastatic; overall, 4 (22.2%) of 18 patients were node-positive. CONCLUSION: We found that 2 of 18 interval SNs were metastatic: This study showed that preoperative lymphoscintigraphy must review all known lymphatic areas in order to exclude an interval SN.

Study of the impact of tissue density heterogeneities on 3-dimensional abdominal dosimetry: comparison between dose kernel convolution and direct monte carlo methods.

Dose kernel convolution (DK) methods have been proposed to speed up absorbed dose calculations in molecular radionuclide therapy. Our aim was to evaluate the impact of tissue density heterogeneities (TDH) on dosimetry when using a DK method and to propose a simple density-correction method. METHODS: This study has been conducted on 3 clinical cases: case 1, non-Hodgkin lymphoma treated with (131)I-tositumomab; case 2, a neuroendocrine tumor treatment simulated with (177)Lu-peptides; and case 3, hepatocellular carcinoma treated with (90)Y-microspheres. Absorbed dose calculations were performed using a direct Monte Carlo approach accounting for TDH (3D-RD), and a DK approach (VoxelDose, or VD). For each individual voxel, the VD absorbed dose, D(VD), calculated assuming uniform density, was corrected for density, giving D(VDd). The average 3D-RD absorbed dose values, D(3DRD), were compared with D(VD) and D(VDd), using the relative difference Δ(VD/3DRD). At the voxel level, density-binned Δ(VD/3DRD) and Δ(VDd/3DRD) were plotted against ρ and fitted with a linear regression. RESULTS: The D(VD) calculations showed a good agreement with D(3DRD). Δ(VD/3DRD) was less than 3.5%, except for the tumor of case 1 (5.9%) and the renal cortex of case 2 (5.6%). At the voxel level, the Δ(VD/3DRD) range was 0%-14% for cases 1 and 2, and -3% to 7% for case 3. All 3 cases showed a linear relationship between voxel bin-averaged Δ(VD/3DRD) and density, ρ: case 1 (Δ = -0.56ρ + 0.62, R(2) = 0.93), case 2 (Δ = -0.91ρ + 0.96, R(2) = 0.99), and case 3 (Δ = -0.69ρ + 0.72, R(2) = 0.91). The density correction improved the agreement of the DK method with the Monte Carlo approach (Δ(VDd/3DRD) < 1.1%), but with a lesser extent for the tumor of case 1 (3.1%). At the voxel level, the Δ(VDd/3DRD) range decreased for the 3 clinical cases (case 1, -1% to 4%; case 2, -0.5% to 1.5%, and -1.5% to 2%). No more linear regression existed for cases 2 and 3, contrary to case 1 (Δ = 0.41ρ - 0.38, R(2) = 0.88) although the slope in case 1 was less pronounced. CONCLUSION: This study shows a small influence of TDH in the abdominal region for 3 representative clinical cases. A simple density-correction method was proposed and improved the comparison in the absorbed dose calculations when using our voxel S value implementation.

Context-dependent dual role of SKI8 homologs in mRNA synthesis and turnover.

Eukaryotic mRNA transcription and turnover is controlled by an enzymatic machinery that includes RNA polymerase II and the 3' to 5' exosome. The activity of these protein complexes is modulated by additional factors, such as the nuclear RNA polymerase II-associated factor 1 (Paf1c) and the cytoplasmic Superkiller (SKI) complex, respectively. Their components are conserved across uni- as well as multi-cellular organisms, including yeast, Arabidopsis, and humans. Among them, SKI8 displays multiple facets on top of its cytoplasmic role in the SKI complex. For instance, nuclear yeast ScSKI8 has an additional function in meiotic recombination, whereas nuclear human hSKI8 (unlike ScSKI8) associates with Paf1c. The Arabidopsis SKI8 homolog VERNALIZATION INDEPENDENT 3 (VIP3) has been found in Paf1c as well; however, whether it also has a role in the SKI complex remains obscure so far. We found that transgenic VIP3-GFP, which complements a novel vip3 mutant allele, localizes to both nucleus and cytoplasm. Consistently, biochemical analyses suggest that VIP3-GFP associates with the SKI complex. A role of VIP3 in the turnover of nuclear encoded mRNAs is supported by random-primed RNA sequencing of wild-type and vip3 seedlings, which indicates mRNA stabilization in vip3. Another SKI subunit homolog mutant, ski2, displays a dwarf phenotype similar to vip3. However, unlike vip3, it displays neither early flowering nor flower development phenotypes, suggesting that the latter reflect VIP3's role in Paf1c. Surprisingly then, transgenic ScSKI8 rescued all aspects of the vip3 phenotype, suggesting that the dual role of SKI8 depends on species-specific cellular context.

The interplay of spatial and climatic landscapes in the genetic distribution of a South American parrot

AimOur aim was to understand the interplay of heterogeneous climatic and spatial landscapes in shaping the distribution of nuclear microsatellite variation in burrowing parrots, Cyanoliseus patagonus. Given the marked phenotypic differences between populations of burrowing parrots we hypothesized an important role of geographical as well climatic heterogeneity in the population structure of this species. LocationSouthern South America. MethodsWe applied a landscape genetics approach to investigate the explicit patterns of genetic spatial autocorrelation based on both geography and climate using spatial principal component analysis (sPCA). This necessitated a novel statistical estimation of the species climatic landscape, considering temperature- and precipitation-based variables separately to evaluate their weight in shaping the distribution of genetic variation in our model system. ResultsGeographical and climatic heterogeneity successfully explained molecular variance in burrowing parrots. sPCA divided the species distribution into two main areas, Patagonia and the pre-Andes, which were connected by an area of geographical and climatic transition. Moreover, sPCA revealed cryptic and conservation-relevant genetic structure: the pre-Andean populations and the transition localities were each divided into two groups, each management units for conservation. Main conclusionssPCA, a method originally developed for spatial genetics, allowed us to unravel the genetic structure related to spatial and climatic landscapes and to visualize these patterns in landscape space. These novel climatic inferences underscore the importance of our modified sPCA approach in revealing how climatic variables can drive cryptic patterns of genetic structure, making the approach potentially useful in the study of any species distributed over a climatically heterogeneous landscape.

Probabilistic age classification with Bayesian networks: a study on the ossification status of the medial clavicular epiphysis

In the past few decades, the rise of criminal, civil and asylum cases involving young people lacking valid identification documents has generated an increase in the demand of age estimation. The chronological age or the probability that an individual is older or younger than a given age threshold are generally estimated by means of some statistical methods based on observations performed on specific physical attributes. Among these statistical methods, those developed in the Bayesian framework allow users to provide coherent and transparent assignments which fulfill forensic and medico-legal purposes. The application of the Bayesian approach is facilitated by using probabilistic graphical tools, such as Bayesian networks. The aim of this work is to test the performances of the Bayesian network for age estimation recently presented in scientific literature in classifying individuals as older or younger than 18 years of age. For these exploratory analyses, a sample related to the ossification status of the medial clavicular epiphysis available in scientific literature was used. Results obtained in the classification are promising: in the criminal context, the Bayesian network achieved, on the average, a rate of correct classifications of approximatively 97%, whilst in the civil context, the rate is, on the average, close to the 88%. These results encourage the continuation of the development and the testing of the method in order to support its practical application in casework.

Distribution and population genetic variation of cryptic species of the Alpine mayfly Baetis alpinus (Ephemeroptera: Baetidae) in the Central Alps.

BACKGROUND: Many species contain evolutionarily distinct groups that are genetically highly differentiated but morphologically difficult to distinguish (i.e., cryptic species). The presence of cryptic species poses significant challenges for the accurate assessment of biodiversity and, if unrecognized, may lead to erroneous inferences in many fields of biological research and conservation. RESULTS: We tested for cryptic genetic variation within the broadly distributed alpine mayfly Baetis alpinus across several major European drainages in the central Alps. Bayesian clustering and multivariate analyses of nuclear microsatellite loci, combined with phylogenetic analyses of mitochondrial DNA, were used to assess population genetic structure and diversity. We identified two genetically highly differentiated lineages (A and B) that had no obvious differences in regional distribution patterns, and occurred in local sympatry. Furthermore, the two lineages differed in relative abundance, overall levels of genetic diversity as well as patterns of population structure: lineage A was abundant, widely distributed and had a higher level of genetic variation, whereas lineage B was less abundant, more prevalent in spring-fed tributaries than glacier-fed streams and restricted to high elevations. Subsequent morphological analyses revealed that traits previously acknowledged as intraspecific variation of B. alpinus in fact segregated these two lineages. CONCLUSIONS: Taken together, our findings indicate that even common and apparently ecologically well-studied species may consist of reproductively isolated units, with distinct evolutionary histories and likely different ecology and evolutionary potential. These findings emphasize the need to investigate hidden diversity even in well-known species to allow for appropriate assessment of biological diversity and conservation measures.