Spatiotemporal evolution of COVID-19 in Portugal's Mainland with self-organizing maps.

BACKGROUND: Self-Organizing Maps (SOM) are an unsupervised learning clustering and dimensionality reduction algorithm capable of mapping an initial complex high-dimensional data set into a low-dimensional domain, such as a two-dimensional grid of neurons. In the reduced space, the original complex patterns and their interactions can be better visualized, interpreted and understood. METHODS: We use SOM to simultaneously couple the spatial and temporal domains of the COVID-19 evolution in the 278 municipalities of mainland Portugal during the first year of the pandemic. Temporal 14-days cumulative incidence time series along with socio-economic and demographic indicators per municipality were analyzed with SOM to identify regions of the country with similar behavior and infer the possible common origins of the incidence evolution. RESULTS: The results show how neighbor municipalities tend to share a similar behavior of the disease, revealing the strong spatiotemporal relationship of the COVID-19 spreading beyond the administrative borders of each municipality. Additionally, we demonstrate how local socio-economic and demographic characteristics evolved as determinants of COVID-19 transmission, during the 1st wave school density per municipality was more relevant, where during 2nd wave jobs in the secondary sector and the deprivation score were more relevant. CONCLUSIONS: The results show that SOM can be an effective tool to analysing the spatiotemporal behavior of COVID-19 and synthetize the history of the disease in mainland Portugal during the period in analysis. While SOM have been applied to diverse scientific fields, the application of SOM to study the spatiotemporal evolution of COVID-19 is still limited. This work illustrates how SOM can be used to describe the spatiotemporal behavior of epidemic events. While the example shown herein uses 14-days cumulative incidence curves, the same analysis can be performed using other relevant data such as mortality data, vaccination rates or even infection rates of other disease of infectious nature.

Oxidative Stress and Antioxidant Defense in the Brain of Bat Species with Different Feeding Habits.

Assessing the levels of oxidative stress markers and antioxidant enzymes in the brain is crucial in evaluating its antioxidant capacity and understanding the influence of various dietary patterns on brain well-being. This study aimed to investigate the antioxidant status and oxidative damage in the brain of bat species with different feeding habits to gain insights into their protective mechanisms against oxidative stress and their interspecific variation. The levels of oxidative damage markers and the activities of antioxidants were measured in the brain of four bat species with different feeding habits, namely insectivorous, frugivorous, nectarivorous, and hematophagous. Insectivorous bats showed higher levels of SOD and fumarase compared to the other groups, while hematophagous bats showed lower levels of these enzymes. On the other hand, the activities of glutathione peroxidase and glutathione S-transferase were higher in hematophagous bats and lower in insectivorous bats. The carbonyl groups and malondialdehyde levels were lower in frugivores, while they were similar in the other feeding guilds. Nitrite and nitrate levels were higher in the hematophagous group and relatively lower in all other groups. The GSSG/GSH ratio was higher in the hematophagous group and lower in frugivores. Overall, our results indicate that the levels of oxidative stress markers and the activities of antioxidant enzymes in the brain vary significantly among bat species with different feeding habitats. The findings suggest that the antioxidant status of the brain is influenced by diet and feeding habits.

Oxidative Stress and Antioxidant Defense in the Heart, Liver, and Kidney of Bat Species with Different Feeding Habits.

The aim of this study was to compare the oxidative metabolism of four neotropical bat species with different feeding habits and investigate the relationship between their feeding habits and oxidative status. In terms of oxidative damage, our findings revealed major differences among the four bat species. In particular, hematophagous bats had lower levels of oxidative damage in the heart but higher levels in the liver. Nectarivorous bats had lower levels of carbonyl groups in the kidneys compared to insectivorous and hematophagous bats. The activity of various antioxidant and non-antioxidant enzymes in the heart, liver, and kidney also showed significant differences among the bat species. H2O2 consumption was lower in the heart of hematophagous bats, while insectivorous bats exhibited the highest enzymatic activity in the kidney. SOD activity was lower in the heart of hematophagous bats and lower in nectarivorous bats in the liver. Fumarase activity was higher in the heart of frugivorous/insectivorous and lower in nectarivorous/hematophagous bats. GPx activity was higher in the heart of nectarivorous/insectivorous and higher in the kidney of insectivorous bats. GST activity was higher in the heart of nectarivorous and lower in hematophagous bats. The correlation analysis between oxidative markers and enzymatic/non-enzymatic antioxidants in the heart, liver, and kidney exhibited distinct patterns of correlations due to variations in antioxidant defense mechanisms and oxidative stress responses in different organs. The observed differences in oxidative damage, antioxidant enzyme activities, and correlations between oxidative markers and antioxidants highlight the adaptability and complexity of the antioxidant defense systems in these bats. Each organ appears to have specific demands and adaptations to cope with oxidative stress based on its physiological functions and exposure to dietary components. Our results have major significance for the conservation and management of bats, which are threatened species despite being crucial components of ecosystems. Our study's implications go beyond bat biology and offer valuable insights into comparative oxidative physiology.

How aerial insectivore bats of different sizes respond to nightly temperature shifts.

Small, volant and nocturnal, bats face strong challenges to avoid heat loss. Among aerial insectivores, body mass varies by two orders of magnitude between the smallest and the largest species. At low temperatures, physiological constraints should be harsher for smaller bats, as they lose more heat through their body surface than larger species. So, temperature variations should lead to distinct behavioural responses by bats of different body masses. Also, because they feed on arthropods, dependent on ambient temperature, aerial insectivores should halt feeding at low temperatures. Using ultrasound detectors and temperature and humidity sensors, we investigated how aerial insectivores of the coldest region in austral Brazil respond to nightly temperature variations and compared those responses between guilds of distinct body masses. We predict that smaller bats reduce their activity faster than larger bats, but that foraging should reduce simultaneously in the two guilds, as they depend on ectothermic prey. Bat activity reduced significantly below 12 °C. Larger bats maintained their activity at temperatures where the activity of smaller bats had already halted. However, larger bats foraged mostly during the first half of the night, at higher temperatures than those chosen by smaller bats to forage. We associate these differential responses to the thermal convection process, which may increase prey availability at higher altitudes, where larger molossids are known to forage. Smaller species, mostly edge-space hunters, probably take advantage of less variable prey availability during the night, resulting in a more regular behavioural pattern of navigation and foraging.

Motion Sensors for Knee Angle Recognition in Muscle Rehabilitation Solutions.

The progressive loss of functional capacity due to aging is a serious problem that can compromise human locomotion capacity, requiring the help of an assistant and reducing independence. The NanoStim project aims to develop a system capable of performing treatment with electrostimulation at the patient's home, reducing the number of consultations. The knee angle is one of the essential attributes in this context, helping understand the patient's movement during the treatment session. This article presents a wearable system that recognizes the knee angle through IMU sensors. The hardware chosen for the wearables are low cost, including an ESP32 microcontroller and an MPU-6050 sensor. However, this hardware impairs signal accuracy in the multitasking environment expected in rehabilitation treatment. Three optimization filters with algorithmic complexity O(1) were tested to improve the signal's noise. The complementary filter obtained the best result, presenting an average error of 0.6 degrees and an improvement of 77% in MSE. Furthermore, an interface in the mobile app was developed to respond immediately to the recognized movement. The systems were tested with volunteers in a real environment and could successfully measure the movement performed. In the future, it is planned to use the recognized angle with the electromyography sensor.

Geostatistical COVID-19 infection risk maps for Portugal.

The rapid spread of the SARS-CoV-2 epidemic has simultaneous time and space dynamics. This behaviour results from a complex combination of factors, including social ones, which lead to significant differences in the evolution of the spatiotemporal pattern between and within countries. Usually, spatial smoothing techniques are used to map health outcomes, and rarely uncertainty of the spatial predictions are assessed. As an alternative, we propose to apply direct block sequential simulation to model the spatial distribution of the COVID-19 infection risk in mainland Portugal. Given the daily number of infection data provided by the Portuguese Directorate-General for Health, the daily updates of infection rates are calculated by municipality and used as experimental data in the geostatistical simulation. The model considers the uncertainty/error associated with the size of each municipality's population. The calculation of daily updates of the infection risk maps results from the median model of one ensemble of 100 geostatistical realizations of daily updates of the infection risk. The ensemble of geostatistical realizations is also used to calculate the associated spatial uncertainty of the spatial prediction using the interquartile distance. The risk maps are updated daily and show the regions with greater risks of infection and the critical dynamics related to its development over time.

Eating down the food chain: generalism is not an evolutionary dead end for herbivores.

The role of trophic specialisation in taxonomic diversification remains unclear. Plant specialists diversify faster than omnivores and animalivores, but at shorter macroevolutionary scales this pattern sometimes reverses. Here, we estimate the effect of diet diversification on speciation rates in noctilionoid bats, controlling for tree shape, rate heterogeneity and macroevolutionary regimes. We hypothesise that niche subdivision among herbivores positively relates to speciation rates, differing between macroevolutionary regimes. We found the rate at which new herbivorous lineages originate decreases as rates of diet evolution increase. Herbivores experience higher speciation rates, but generalist herbivores and predominantly herbivorous omnivores speciate faster than specialised herbivores, omnivores and animalivores. Generalised herbivory is not a dead end. We show that analysing ecological traits and diversification requires accounting for macroevolutionary regimes and within- and between-clade variation in evolutionary rates. Our approach overcomes the high false-positive rates of other methods and illuminates the roles of herbivory and specialisation in speciation.

Updated distribution maps for neotropical bats in the superfamily Noctilionoidea.

The IUCN provides a spatial database for many species, including terrestrial mammals. This database includes shapefiles with taxonomic information and the extent of occurrence for each species, and has been used in hundreds of studies in ecology, biogeography and conservation. Here we provide updated distribution maps that comprise the extent of occurrence of the neotropical bat species in the superfamily Noctilionoidea (Mammalia: Chiroptera) after a thorough research of new records published between January 2008 and March 2018. The main motivation for this update was the inclusion of spatial and climatic variables in explaining the ecological and taxonomic diversity of noctilionoid bats. The core of the superfamily (246 species distributed in five families out of 250 species) occurs in the Neotropics and shows ecological diversity unparalleled among mammals. This clade also shows the only evolutionary shift towards higher speciation rates within the order Chiroptera. Updating the range maps for these bats resulted in the modification of maps of 94 species, and the creation of new maps for 37 species missing from the IUCN database. From the 94 modified maps, 55 species increased their latitudinal range and 38 increased their longitudinal range. These modifications did not change the overall extent of occurrence of the clade. Altogether, modified and new maps represent 53% of Noctilionoidea. No copyright or proprietary restrictions are associated with the use of this data set other than citation of this Data Paper.

Disentangling natural and anthropogenic sources of atmospheric sulfur in an industrial region using biomonitors.

Despite reductions in atmospheric sulfur (S) concentrations due to abatement policies in some countries, modeling the dispersion of this pollutant and disentangling anthropogenic sources from natural ones is still of great concern. Lichens have been used as biomonitors of the impacts of S for over 40 years, but their potential as source-tracers of specific sources, including natural ones, remains unexplored. In fact, few attempts have been made to try to distinguish and spatially model different sources of S using lichens. We have measured S concentrations and isotopic values in lichens within an industrial coastal region where different sources of S, natural and anthropogenic, interplay. We detected a prevailing influence of natural sea-originated S that mixed with anthropogenic sources of S. We were then able to disentangle the sources of S, by removing the ocean influence on S isotopic values, enabling us to model the impact of different anthropogenic sources on S deposition and highlighting the potential use of lichens to evaluate the weight of different types of anthropogenic sources.

Host specificity and metamorphosis of the glochidium of the freshwater mussel Unio tumidiformis (Bivalvia: Unionidae).

The glochidium larvae of freshwater mussels of the family Unionidae need to find suitable hosts to attach themselves and metamorphose into free-living juveniles. The specificity of the host-parasite relationship was investigated for the Iberian Unio tumidiformis Castro, 1885 by means of experimental infections and also by analyzing naturally infected fish. The process of encapsulation of glochidia was studied using scanning electron microscopy. Unio tumidiformis has proven to be an unusual host-specific unionid mussel, apparently parasitizing only fish of the genus Squalius Bonaparte, 1837. Successful encapsulation or complete metamorphosis was observed in five fish taxa: S. aradensis (Coelho, Bogutskaya, Rodrigues et Collares-Pereira), S. caroliterti (Doadrio), S. pyrenaicus (Günther), S. torgalensis (Coelho, Bogutskaya, Rodrigues et Collares-Pereira) and S. alburnoides (Steindachner) complex (only for the nuclear hybrids with at least one copy of the S. pyrenaicus genome). Complete metamorphose was achieved in 6 to 14 days at mean temperatures ranging from 21.8 to 26.1 degrees C. The current study provides support for cell migration being the main force of cyst formation and shows the influence of potential host's genome in response to the infection process to determine the success of the metamorphosis.

Distribution and Utilization of Vitamin E in Different Organs of Wild Bats from Different Food Groups.

In this work, we examined the levels of vitamin E in the heart, liver, and kidneys of four species of adult male bats with distinct feeding habits. Our results indicate consistent vitamin E levels in the heart across all four bat species, suggesting the presence of regulatory mechanisms. Additionally, the liver displayed notably higher vitamin E levels in nectarivorous and frugivorous bats, while hematophagous bats exhibited lower levels, indicating a link between dietary intake and liver vitamin E levels. Furthermore, correlation analysis provided additional insights into the relationships between vitamin E and key antioxidant parameters in the livers of bats. On the other hand, no correlation was observed between vitamin E and key antioxidant parameters in the heart. Intriguingly, vitamin E was not detected in the kidneys, likely due to physiological factors and the prioritization of vitamin E mobilization in the heart, where it serves critical physiological functions. This unexpected absence of vitamin E in bat kidneys highlights the unique metabolic demands and prioritization of vitamin mobilization in wild animals like bats, compared to conventional animal models. These findings provide insight into the intricate distribution and utilization of vitamin E in bats, emphasizing the influence of dietary intake and metabolic adaptations on vitamin E levels in different organs.

Understanding spatiotemporal patterns of COVID-19 incidence in Portugal: A functional data analysis from August 2020 to March 2022.

During the SARS-CoV-2 pandemic, governments and public health authorities collected massive amounts of data on daily confirmed positive cases and incidence rates. These data sets provide relevant information to develop a scientific understanding of the pandemic's spatiotemporal dynamics. At the same time, there is a lack of comprehensive approaches to describe and classify patterns underlying the dynamics of COVID-19 incidence across regions over time. This seriously constrains the potential benefits for public health authorities to understand spatiotemporal patterns of disease incidence that would allow for better risk communication strategies and improved assessment of mitigation policies efficacy. Within this context, we propose an exploratory statistical tool that combines functional data analysis with unsupervised learning algorithms to extract meaningful information about the main spatiotemporal patterns underlying COVID-19 incidence on mainland Portugal. We focus on the timeframe spanning from August 2020 to March 2022, considering data at the municipality level. First, we describe the temporal evolution of confirmed daily COVID-19 cases by municipality as a function of time, and outline the main temporal patterns of variability using a functional principal component analysis. Then, municipalities are classified according to their spatiotemporal similarities through hierarchical clustering adapted to spatially correlated functional data. Our findings reveal disparities in disease dynamics between northern and coastal municipalities versus those in the southern and hinterland. We also distinguish effects occurring during the 2020-2021 period from those in the 2021-2022 autumn-winter seasons. The results provide proof-of-concept that the proposed approach can be used to detect the main spatiotemporal patterns of disease incidence. The novel approach expands and enhances existing exploratory tools for spatiotemporal analysis of public health data.

The role of CD20+ T cells: Insights in human peripheral blood.

CD20+ T cells constitute a small subset of T cells. These are found among CD4+, CD8+, CD4+CD8+, CD4-CD8- T, and TCRγδ+ T cells, and have been poorly characterized. The aim of this study was to characterize peripheral blood (PB) CD20+ T cells and compare them to their PB CD20- T cell counterparts. PB from 17 healthy individuals was collected. The distribution of CD20+ T cells among maturation-associated T cells compartments (naïve, central memory, transitional memory, effector memory, and effector T cells), their polarization, activation status, and expression of immune-regulatory proteins were evaluated by flow cytometry. Their function was also assessed, by measuring IFN-γ, TNF-α, and IL-17 production. Compared with CD20- T cells, CD20+ T cells represent a higher proportion of transitional memory cells. Furthermore, CD20+ T cells display a proinflammatory phenotype, characterized by the expansion of Th1, Th1/17, and Tc1 cell subsets , associated to a high expression of activation (CD25) and exhaustion (PD-1) markers. In addition, the simultaneous production of the proinflammatory cytokines IFN-γ, TNF-α, and IL-17 was also detected in CD4+CD20+ T cells. Our results show that CD20+ T cells are phenotypically and functionally different from CD20- T cells, suggesting that these cells are a distinct subset of T cells.

Overview of Ethnobotanical-Pharmacological Studies Carried Out on Medicinal Plants from the Serra da Estrela Natural Park: Focus on Their Antidiabetic Potential.

The Serra da Estrela Natural Park (NPSE) in Portugal stands out as a well-preserved region abundant in medicinal plants, particularly known for their pharmaceutical applications in diabetes prevention and treatment. This comprehensive review explores these plants' botanical diversity, traditional uses, pharmacological applications, and chemical composition. The NPSE boast a rich diversity with 138 medicinal plants across 55 families identified as traditionally and pharmacologically used against diabetes globally. Notably, the Asteraceae and Lamiaceae families are prevalent in antidiabetic applications. In vitro studies have revealed their significant inhibition of carbohydrate-metabolizing enzymes, and certain plant co-products regulate genes involved in carbohydrate metabolism and insulin secretion. In vivo trials have demonstrated antidiabetic effects, including glycaemia regulation, insulin secretion, antioxidant activity, and lipid profile modulation. Medicinal plants in NPSE exhibit various activities beyond antidiabetic, such as antioxidant, anti-inflammatory, antibacterial, anti-cancer, and more. Chemical analyses have identified over fifty compounds like phenolic acids, flavonoids, terpenoids, and polysaccharides responsible for their efficacy against diabetes. These findings underscore the potential of NPSE medicinal plants as antidiabetic candidates, urging further research to develop effective plant-based antidiabetic drugs, beverages, and supplements.

Comparative Analysis of the Gut Microbiota of Bat Species with Different Feeding Habits.

Bats are a diverse and ecologically important group of mammals that exhibit remarkable diversity in their feeding habits. These diverse feeding habits are thought to be reflected in the composition and function of their gut microbiota, which plays important roles in nutrient acquisition, immune function, and overall health. Despite the rich biodiversity of bat species in South America, there is a lack of microbiome studies focusing on bats from this region. Such studies could offer major insights into conservation efforts and the preservation of biodiversity in South America. In this work, we aimed to compare the gut microbiota of four bat species with different feeding habits from Southern Brazil, including nectarivorous, frugivorous, insectivorous, and hematophagous bats. Our findings demonstrate that feeding habits can have a significant impact on the diversity and composition of bat gut microbiotas, with each species exhibiting unique metabolic potentials related to their dietary niches. In addition, the identification of potentially pathogenic bacteria suggests that the carriage of microbial pathogens by bats may vary, depending on feeding habits and host-specific factors. These findings provide novel insights into the relationship between bat feeding habits and gut microbiota composition, highlighting the need to promote diverse habitats and food sources to support these ecologically important species.

Chemogenetic manipulation of astrocyte activity at the synapse- a gateway to manage brain disease.

Astrocytes are the major glial cell type in the central nervous system (CNS). Initially regarded as supportive cells, it is now recognized that this highly heterogeneous cell population is an indispensable modulator of brain development and function. Astrocytes secrete neuroactive molecules that regulate synapse formation and maturation. They also express hundreds of G protein-coupled receptors (GPCRs) that, once activated by neurotransmitters, trigger intracellular signalling pathways that can trigger the release of gliotransmitters which, in turn, modulate synaptic transmission and neuroplasticity. Considering this, it is not surprising that astrocytic dysfunction, leading to synaptic impairment, is consistently described as a factor in brain diseases, whether they emerge early or late in life due to genetic or environmental factors. Here, we provide an overview of the literature showing that activation of genetically engineered GPCRs, known as Designer Receptors Exclusively Activated by Designer Drugs (DREADDs), to specifically modulate astrocyte activity partially mimics endogenous signalling pathways in astrocytes and improves neuronal function and behavior in normal animals and disease models. Therefore, we propose that expressing these genetically engineered GPCRs in astrocytes could be a promising strategy to explore (new) signalling pathways which can be used to manage brain disorders. The precise molecular, functional and behavioral effects of this type of manipulation, however, differ depending on the DREADD receptor used, targeted brain region and timing of the intervention, between healthy and disease conditions. This is likely a reflection of regional and disease/disease progression-associated astrocyte heterogeneity. Therefore, a thorough investigation of the effects of such astrocyte manipulation(s) must be conducted considering the specific cellular and molecular environment characteristic of each disease and disease stage before this has therapeutic applicability.

Network and parasitological analyses reveal latitudinal gradient in bats-ectoparasitic fly interactions across the Neotropic.

Ecological interactions between parasites and their hosts play a fundamental role in evolutionary processes. Selection pressures are exerted on parasites and their hosts, usually resulting in high levels of specificity. Such is the case of ectoparasitic bat-flies, but how large-scale spatial gradients affect the dynamics of their interactions with their bat hosts is still unknown. In the present study, we investigated interaction patterns between bats and their ectoparasitic flies (Streblidae and Nycteribiidae), both presenting their peak of diversity in the Neotropical region, along a latitudinal gradient. Using network analyses and parasitic indices, grounded on the latitudinal diversity gradient pattern, we evaluated how spatial gradients affect species interactions and parasitic indices at the biogeopraphic scale, with increasing species richness in interaction networks closer to the tropics, leading to increases in network modularity, size, and specialization, and to a decrease in nesting and connectivity. We conducted a literature review, focusing on studies done in the Neotropical region, and own data. We obtained a bat richness of 97 species parasitized by 128 species of ectoparasitic flies, distributed into 57 interaction networks between latitudes 29° S and 19° N in the Neotropic. Network metrics and parasitic indices varied along the latitudinal gradient, with changes in the richness of bats and their ectoparasitic flies and in the structure of their interactions; network specialization, modularity, and connectance increase with latitude, while network size decreases with latitude. Regions closer to the equator had higher parasite loads. Our results show that interaction network metrics present a latitudinal gradient and that such interactions, when observed at a local scale, hide variations that only become perceptible at larger scales. In this way, ectoparasites such as bat flies are not only influenced by the ecology and biology of their hosts, but also by other environmental factors acting directly on their distribution and survival.

Neotropical mustelids: fecal metabolome diversity and its potential for taxonomic discrimination.

Chemical profiles of non-invasive biological material, such as feces, have great potential to study elusive animals or those with low population densities. Here, we use a metabolomic approach to evaluate Neotropical mustelids as a biological model to describe the diversity of the metabolites present in fecal samples, as well as to evaluate the potential of chemical profiles for taxonomic discrimination. We collected fecal samples from captive individuals of 5 species of mustelids occurring in Brazil and analyzed them by liquid chromatography coupled to high-resolution mass spectrometry. Over 200 compounds have been annotated; "bile acids, alcohols and derivatives" was the most expressive class in the metabolome of all the species. We successfully discriminated 3 taxonomic groups: 1-tayra (Eira barbara); 2-otters (Lontra longicaudis and Pteronura brasiliensis; 1); and 3-grisons (Galictis vittata and Galictis cuja). Several compounds seemed to be associated with food intake and the digestive process, while others were found for the first time in Neotropical mustelids. We concluded that mustelids show high metabolome diversity and that species-specific identification through metabolomic profiles is possible, thus contributing to the development and implementation of additional non-invasive approaches in the study of mustelids.

Chromosomes of European cyprinid fishes: comparative cytogenetics and chromosomal characteristics of ribosomal DNAs in nine Iberian chondrostomine species (Leuciscinae).

Karyotypes and chromosomal features of both minor and major ribosomal RNA genes (rDNA) were investigated in nine Iberian chondrostomine species by fluorescent in situ hybridization (FISH) with 5S and 45S rDNA probes. All species presented invariably diploid values of 2n = 50 and the characteristic leuciscin karyotype pattern with 6-7 metacentric (m), 15-16 submetacentric (sm) and 3-4 subtelo- to acrocentric (st/a) chromosome pairs. The largest chromosome pair of the set was st/a as typical of Leuciscinae and no heteromorphic chromosomes could be unequivocally associated to sex determination. Achondrostoma occidentale and Pseudochondrostoma willkommii were cytogenetically characterized for the first time while Achondrostoma arcasii and Iberochondrostoma lemmingii were revisited regarding previous karyotype descriptions. Remarkable variability in number and location was observed for both molecular chromosome markers, especially within Achondrostoma and Iberochondrostoma genera. Clusters of 5S rDNA were mostly terminally associated to st/a chromosomes varying from four to eight positive signals, whilst NOR sites directly detected by the 45S rDNA probe were identified in sm chromosomes varying from three to six independent clusters. Frequent population bottlenecks in Mediterranean-type semiarid habitats were hypothesized to explain not only such extensive polymorphism which seems unique among leuciscin cyprinids but also the increased probability of fixation of rDNA translocation events.