Ultrastructural observations on the lung of Pelophylax kl. esculentus tadpoles during development

The anatomical organization of the lung in Pelophylax kl. esculentus tadpoles has been described in early and middle-staged tadpoles (Gosner stages 32 and 39). The lung's ontogenic development has been studied using light microscopy and both scanning and transmission electron microscopy. In the early-staged tadpoles, the lung is well vascularized, but it is provided with only two internal septa types and devoid of ciliated epithelium and lamellar bodies. The complete differentiation process is putatively reached in the middle-staged tadpoles (GS 39), and numerous septa of first, second, and third-order deeply protruding into the lung lumen could be recognized. The histological and ultrastructural features at this stage correspond to that described in the adults' lungs, but mucous secreting goblet cells could not be detected. Pneumocytes, with numerous apical microvilli, surround the growing network of capillaries and show a cytoplasm rich in dense bodies and few mature lamellar bodies. On the epithelial surface, a thin layer of mucous covering the underlying epithelium could also be seen. Neuroepithelial bodies, supposed to be involved in chemoreception, are organized in clusters on the first order septa and surrounded by cytoplasmatic processes originating by neighboring cells. The lung arrangement of P. kl. esculentus is compared with that of other Anurans in order to further elucidate the putative role of the lung in gas exchanges. The present studies reduce the information gap existing in the literature regarding lung morphology and development in amphibian larval stages, also contributing to the discussion on the onset of airbreathing respiration in tadpoles

First Insights on the Karyotype Diversification of the Endemic Malagasy Leaf-Toed Geckos (Squamata: Gekkonidae: Uroplatus)

We provide here the first karyotype description of eight Uroplatus species and a characterization of their chromosomal diversity. We performed a molecular taxonomic assessment of several Uroplatus samples using the mitochondrial 12S marker and a comparative cytogenetic analysis with standard karyotyping, silver staining (Ag-NOR) and sequential C-banding + Giemsa, +Chromomycin A3 (CMA3), +4',6-diamidino-2-phenylindole (DAPI). We found chromosomal variability in terms of chromosome number (2n = 34-38), heterochromatin composition and number and localization of loci or Nucleolar Organizer Regions (NORs) (alternatively on the 2nd, 6th, 10th or 16th pair). Chromosome morphology is almost constant, with karyotypes composed of acrocentric chromosomes, gradually decreasing in length. C-banding evidenced a general low content of heterochromatin, mostly localized on pericentromeric and telomeric regions. Centromeric bands varied among the species studied, resulting in CMA3 positive and DAPI negative or positive to both fluorochromes. We also provide evidence of a first putative heteromorphic sex chromosome system in the genus. In fact, in U. alluaudi the 10th pair was highly heteromorphic, with a metacentric, largely heterochromatic W chromosome, which was much bigger than the Z. We propose an evolutionary scenario of chromosome reduction from 2n = 38 to 2n = 34, by means of translocations of microchromosomes on larger chromosomes (often involving the NOR-bearing microchromosomes). Adding our data to those available from the literature, we show that similar processes characterized the evolutionary radiation of a larger gecko clade. Finally, we hypothesize that sex chromosome diversification occurred independently in different genera

Chromosome Diversity and Evolution of the Endemic Malagasy Velvet Geckos of the Genus Blaesodactylus (Reptilia, Gekkonidae)

: We performed a molecular and phylogenetic analysis and a comparative cytogenetic study with standard karyotyping, silver staining (Ag-NOR) and sequential C-banding + Giemsa, + fluorochromes on several Blaesodactylus samples. The phylogenetic inference retrieved two main clades, the first comprises B. victori, B. microtuberculatus and B. boivini, while the second includes B. sakalava, B. antongilensis and B. ambonihazo. The available samples of B. sakalava form two different clades (here named B. sakalava clade A and clade B), which probably deserve a taxonomic re-evaluation. We found a karyological variability in Blaesodactylus in terms of chromosome number (2n = 40-42), morphology, location of NORs, and heterochromatin distribution pattern. Blaesodactylus antongilensis and B. sakalava clade A and B showed a karyotype of 2n = 40 mostly telocentric chromosomes. Pairs 1 and 6 were metacentric in B. sakalava clade A and B, while pair 1 was composed of subtelocentric/submetacentric elements in B. antongilensis. In contrast, B. boivini displayed a karyotype with 2n = 42 only telocentric chromosomes. NORs were on the first chromosome pair in B. boivini, and on the second pair in B. antongilensis. Adding our data to those available from the literature on evolutionarily related species, we highlight that the chromosome diversification in the genus probably proceeded towards a progressive reduction in the chromosome number and the formation of metacentric elements

Evolutionary and Genomic Diversity of True Polyploidy in Tetrapods

: True polyploid organisms have more than two chromosome sets in their somatic and germline cells. Polyploidy is a major evolutionary force and has played a significant role in the early genomic evolution of plants, different invertebrate taxa, chordates, and teleosts. However, the contribution of polyploidy to the generation of new genomic, ecological, and species diversity in tetrapods has traditionally been underestimated. Indeed, polyploidy represents an important pathway of genomic evolution, occurring in most higher-taxa tetrapods and displaying a variety of different forms, genomic configurations, and biological implications. Herein, we report and discuss the available information on the different origins and evolutionary and ecological significance of true polyploidy in tetrapods. Among the main tetrapod lineages, modern amphibians have an unparalleled diversity of polyploids and, until recently, they were considered to be the only vertebrates with closely related diploid and polyploid bisexual species or populations. In reptiles, polyploidy was thought to be restricted to squamates and associated with parthenogenesis. In birds and mammals, true polyploidy has generally been considered absent (non-tolerated). These views are being changed due to an accumulation of new data, and the impact as well as the different evolutionary and ecological implications of polyploidy in tetrapods, deserve a broader evaluation

Tissues injury and pathological changes in Hyla intermedia juveniles after chronic larval exposure to tebuconazole.

Abstract Tebuconazole (TBZ), an azole pesticide, is one of the most frequently detected fungicides in surface water. Despite its harmful effects, mainly related to endocrine disturbance, the consequences of TBZ exposure in amphibians remain poorly understood. Here, we investigated the adverse and delayed effects of TBZ chronic exposure on a native anuran species, often inhabiting cultivated areas, the Italian tree frog (Hyla intermedia). To disclose the multiple mechanisms of action through which TBZ exerts its toxicity we exposed tadpoles over the whole larval period to two sublethal TBZ concentrations (5 and 50 μg/L), and we evaluated histological alterations in three target organs highly susceptible to xenobiotics: liver, kidney, and gonads. We also assessed morphometric and gravimetric parameters: snout-vent length (SVL), body mass (BM), liver somatic index (LSI), and gonad-mesonephros complex index (GMCI) and determined sex ratio, gonadal development, and differentiation. Our results show that TBZ induces irreversible effects on multiple target organs in H. intermedia, exerting its harmful effects through several pathological pathways, including a massive inflammatory response. Moreover, TBZ markedly affects sexual differentiation also by inducing the appearance of sexually undetermined individuals and a general delay of germ cell maturation. Given the paucity of data on the effects of TBZ in amphibians, our results will contribute to a better understanding of the environmental risk posed by this fungicide to the most endangered group of vertebrates

Chromosome analysis on Central and Southern Italy population of the common toad, Bufo bufo (Amphibia, Anura)

Amphibians constitute a very good model to explore the historical aspects of species distributions due to their low dispersal capacity and low individual vagility. Bufonidae are one of the most speciose family of Anura, including taxa, such as Bufo bufo, widespread in Eurasian regions. We performed a karyological study with standard and sequential C-banding + fluorochromes (Chromomycin A3 (CMA) and Diamidinophenylindole (DAPI) on several tadpoles from different populations of Central and Southern Italy. All the examined tadpoles exhibited the standard Bufokaryotype of 2n = 22 biarmed chromosomes, with the first six pairs larger than the other five (7 - 11) pairs and NOR associated heterochromatin distal on the long arms of the 6thchromosome pair, that was also the only chromosome CMA-positive region. C-banding evidenced centromeric heterochromatin, DAPI positive, on all the chromosomes in all the studied populations from Central Italy. The Southern Italy populations differed in additional paracentromeric C-bands on the short arms of chromosomes 1, 3 and 5. These results support the partition of Central populations of B. bufo from the Southern ones, as evidenced also from molecular phylogenetic studie

Comparative cytogenetics on Zamenis lineatus and Elaphe quatuorlineata (Serpentes: Colubridae)

Because of their peculiar genomic and chromosomal characteristics, reptiles are extraordinary model organisms to study karyotype and sex chromosome evolution, but despite the growing interest in their evolutionary cytogenetics, only a small fraction of species have a known karyotype. We performed a comparative cytogenetic analysis on Elaphe quatuorlineata and Zamenis lineatus, using classic and molecular techniques. We provide the karyotype of these two species and an assessment of their chromosomal features. Chromosome analysis was performed with standard karyotyping, C-banding, sequential C-banding + CMA3 + DAPI and Ag-NOR staining. On E. quatuorlineata, we also performed CMA3-methyl green staining and Fluorescence in situ Hybridization mapping NOR loci (NOR-FISH). Elaphe quatuorlineata and Z. lineatus show a very similar karyotype of 2n = 36, with 8 macro- and 10 microchromosome pairs, but differ in the morphology of the pair 8, which resulted submetacentric in the former and metacentric in the latter species. By comparing our data to those available from the literature on congeneric species, we analysed the occurrence of primitive and derivate chromosomal characters and provide cytotaxonomic insights, which further support the species status of Z. lineatus. In both species, the 4th pair was identified as the sex chromosome pair (ZZ/ZW) and NORs were localized on a microchromosome pair. We finally highlight in both genera Elaphe and Zamenis different stages of heterochromatinization of the W chromosome, in agreement with the progressive diversification model of sex chromosome as already shown in different reptile taxa

Sightings and successful reproduction of allochthonous reptiles in Calabria

This paper reports information about the presence of three allochthonous reptiles species in Calabria: Testudo marginata, Trachemys scripta elegans and Chamaeleo chamaeleon. The first one was found in three sites located in the Catena Costiera Massif and in the Crati Valley (Northern Calabria). The slider turtle was found in seven different sites throughout all the region. It massively colonised the Angitola artificial lake: here, this turtle lives in natural conditions and its reproduction was confirmed by the presence of nests, eggs and hatchlings. C. chamaeleon is present in sandy coastal habitats near Palmi and Gioia Tauro (Southern Calabria). From a conservationistic point of view, serious damages to autochtonous species could be caused by the spreading of T. scripta elegans: this species has already determined the local extinction of Angitola’s Emys orbicularis populations

Morphological and Functional Alterations in Zebrafish (Danio rerio) Liver after Exposure to Two Ecologically Relevant Concentrations of Lead

Lead (Pb) is a non-essential, highly toxic, and persistent element widely recognized as one of the most concerning pollutants. It is listed on the Priority List of Hazardous Substances. Widespread environmental contamination from Pb is a serious issue for human health and wildlife. In fish, Pb mainly accumulates in the liver, which is a key component for metal detoxification and excretion processes. In this study, we investigated, for the first time, the morphological and functional injuries induced in zebrafish (Danio rerio) liver by two very low and environmentally relevant concentrations of Pb (2.5 and 5 μg/L) after 48, 96, and 192 h of exposure. We observed significant histological alterations in all the exposed samples, and it was demonstrated that the extent of injuries increased with dose and exposure time. The most common modifications observed were congestion of blood vessels and sinusoids, cytoplasmic vacuolizations, parenchyma dyschromia, and macrophage proliferation. Pb administration also resulted in a significant increase in lipid content and the upregulation of key genes that are involved in metal detoxification (mtf1) and the defensive response against oxidative stress (sod1 and cat). We show that even very low doses of Pb can disrupt liver morphology and function.Fil: Macirella, Rachele. Università della Calabria; ItaliaFil: Curcio, Vittoria. Università della Calabria; ItaliaFil: Ahmed, Abdalmoiz I. M.. Università della Calabria; ItaliaFil: Talarico, Federica. Università della Calabria; ItaliaFil: Sesti, Settimio. Università della Calabria; ItaliaFil: Paravani, Enrique Valentin. Universidad Nacional de Entre Ríos. Facultad de Ingeniería; ArgentinaFil: Odetti, Lucia Magdalena. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Cátedra de Toxicología y Bioquímica Legal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Mezzasalma, Marcello. Università della Calabria; ItaliaFil: Brunelli, Elvira. Università della Calabria; Itali