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[Abstract] The clinical success of cisplatin, carboplatin, and oxaliplatin has sparked the interest of medicinal inorganic chemistry to synthesize and study compounds with non-platinum metal centers. Despite Ru(II)–polypyridyl complexes being widely studied and well established for their antitumor properties, there are not enough in vivo studies to establish the potentiality of this type of compound. Therefore, we report to the best of our knowledge the first in vivo study of Ru(II)–polypyridyl complexes against breast cancer with promising results. In order to conduct our study, we used MCF7 zebrafish xenografts and ruthenium complexes [Ru(bipy)2(C12H8N6-N,N)][CF3SO3]2Ru1 and [{Ru(bipy)2}2(μ-C12H8N6-N,N)][CF3SO3]4Ru2, which were recently developed by our group. Ru1 and Ru2 reduced the tumor size by an average of 30% without causing significant signs of lethality when administered at low doses of 1.25 mg·L−1. Moreover, the in vitro selectivity results were confirmed in vivo against MCF7 breast cancer cells. Surprisingly, this work suggests that both the mono- and the dinuclear Ru(II)–polypyridyl compounds have in vivo potential against breast cancer, since there were no significant differences between both treatments, highlighting Ru1 and Ru2 as promising chemotherapy agents in breast cancer therapy. Xunta de Galicia; ED431C 2018/39 Portugal. Fundação para a Ciência e a Tecnologia; PEst 2015-2020 Portugal. Fundação para a Ciência e a Tecnologia; UID/Multi/04349/2013 Portugal. Fundação para a Ciência e a Tecnologia; RECI/QEQ-QIN/0189/2012 Portugal. Fundação para a Ciência e a Tecnologia; UID/QUI/00100/2020 Portugal. Fundação para a Ciência e a Tecnologia; UIDP/04378/2020 Portugal. Fundação para a Ciência e a Tecnologia; UIDB/04378/2020 Portugal. Fundação para a Ciência e a Tecnologia; LA/P/0140/2020

Zebrafish has become a popular research model in the last years, and several diseases affecting zebrafish research facilities have been reported. However, only one case of naturally occurring viral infections was described for this species. In 2015, infectious spleen and kidney necrosis virus (ISKNV) was detected in zebrafish from a research facility in Spain. Affected fish showed lethargy, loss of appetite, abnormal swimming, distention of the coelomic cavity and, in the most severe cases, respiratory distress, pale gills and petechial haemorrhages at the base of fins. Cytomegaly was the most relevant histopathological finding in organs and tissues, sometimes associated to degenerative and necrotic changes. ISKNV belongs to the relatively newly defined genus Megalocytivirus, family Iridoviridae, comprising large, icosahedral cytoplasmic DNA viruses. This is the first case of naturally occurring Megalocytivirus infection in zebrafish research facilities, associated with morbidity. The virus has been identified based on both pathologic and genetic evidence, to better understand the pathogenesis of the infection in zebrafish and the phylogenetic relationship with other iridoviruses. Given the ability of megalocytiviruses to cross-species boundaries, it seems necessary to implement stringent biosecurity practices as these infections may invalidate experimental data and have major impact on laboratory and cultured fish.

The zebrafish, Danio rerio, has become recognized as a valuable model for infectious diseases. Here we evaluated the susceptibility of zebrafish to be infected with the mammalian vesicular stomatitis virus (VSV). Both zebrafish cells and embryos were highly susceptible to VSV infection. Mortalities exceeded 80% in infected embryos and were preceded by the invasion of the central nervous system by VSV. Live imaging of the infection with GFP-VSV as well as virus titration from infected fish confirmed the viral replication. Immunohistochemical analysis of embryonic fish provided evidence of viral antigens as well as of the apoptosis marker caspase-3 in the brain, eye, liver, pronephros, and skeletal muscle. So far, this is the first report describing the susceptibility of zebrafish to the mammalian virus VSV.

Ruthenium-based drugs exhibit interesting properties as potential anticancer pharmaceuticals. We herein present the synthesis and characterization of a new family of ruthenium complexes with formulas [{Ru(bipy)2}2(μ-L)][CF3SO3]4 (L = bptz, 1a) and [{Ru(bipy)2}2(μ-L)][CF3SO3]2 (L = arphos, 2a; dppb, 3a; dppf, 4a), which were synthesized from the Ru(II) precursor compound cis-Ru(bipy)2Cl2. The complexes were characterized by elemental analysis, mass spectrometry, 1H and 31P{1H} NMR, IR spectroscopy, and conductivity measurements. The molecular structures for three Ru(II) compounds were determined by single-crystal X-ray diffraction. The newly developed compounds interact with CT-DNA by intercalation, in particular, 2a, 3a, and 4a, which also seemed to induce some extent of DNA degradation. This effect seemed to be related with the formation of reactive oxygen species. The cytotoxic activity was evaluated against A2780, MCF7, and MDAMB231 human tumor cells. Compounds 2a and 4a were the most cytotoxic with activity compared to cisplatin (∼2 μM, 72 h) in the A2780 cisplatin sensitive cells. All the compounds induced A2780 cell death by apoptosis, however, to a lesser extent for compounds 4a and 2a. For these compounds, the mechanism of cell death in addition to apoptosis seemed to involve autophagy. In vivo toxicity was evaluated using the zebrafish embryo model. LC50 estimates varied from 5.397 (3a) to 39.404 (1a) mg/L. Considering the in vivo toxicity in zebrafish embryos and the in vitro cytotoxicity in cancer cells, compound 1a seems to be the safest having no effect on dechirionation and presenting a good antiproliferative activity against ovarian carcinoma cells.

Cellular senescence is considered a stress response imposing a stable cell cycle arrest to restrict the growth of damaged cells. More recently however, cellular senescence was identified during mouse embryo development at particular structures during specific periods of time. This programmed cell senescence has been proposed to serve developmental and morphogenetic functions and to potentially represent an evolutionary origin of senescence. Cellular senescence has also been described to take place during bird (chick and quail) and amphibian (xenopus and axoltl) development. Fish however, have been described to show a very narrow and restricted pattern of developmental cell senescence. Here we carried out a detailed characterization of senescence during zebrafish development and found it to be conserved and widespread. Apart from yolk and cloaca, previously described structures, we also identified senescence in the developing central nervous system, intestine, liver, pronephric ducts, and crystalline. Interestingly, senescence at these developing structures disappeared upon treatment with senolytic compound ABT-263, supporting their senescent identity and opening the possibility of studying the contribution of this process to development. In summary, our findings extend the description of developmentally-programmed cell senescence to lower vertebrates contributing to the notion of the relevance of this process for embryo development Funding at the laboratory of M.C. is provided by the Ministerio de Ciencia, Innovación y Universidades, Fondos Europeos de Desarrollo Regional (FEDER) (RTI2018-095818-B-100). Work in the laboratory of A.B.-I. was funded by grants from the Xunta de Galicia (2016-PG008) and the crowdfunding platform Precipita (FECYT; 2017-CP081). Funding at laboratory of L.S. is provided by Xunta de Galicia (ED431C2018/28) SI

Macrophages are pivotal cells of the innate immune system specialized in the phagocytosis of foreign elements. Nanoparticles intentionally designed to target macrophages and modulate their response are of especial interest in the case of chronic inflammatory diseases, cancer and for vaccine development. This work aimed to understand the role of size and shell composition of polymeric nanocapsules (NCs) in their interaction with macrophages, both in vitro and in vivo. A systematic study was performed using two different sizes of inulin and chitosan NCs, negatively and positively charged, respectively, small (≈ 70 nm) and medium (170–250 nm). The in vitro results showed that small NCs interacted more efficiently with macrophages than their larger counterparts. Inulin NCs were significantly less toxic than chitosan NCs. Finally, following in vivo administration (intravenous/intramuscular) to zebrafish, small NCs, regardless of their composition, disseminated considerably faster and further than their medium size counterparts. These results emphasize how small changes in the nanometric range can lead to a remarkably different interaction with the immune cells and biodistribution profile.

Cellular senescence is a stress response that limits the proliferation of damaged cells by establishing a permanent cell cycle arrest. Different stimuli can trigger senescence but excessive production or impaired clearance of these cells can lead to their accumulation during aging with deleterious effects. Despite this potential negative side of cell senescence, its physiological role as a pro‐regenerative and morphogenetic force has emerged recently after the identification of programmed cell senescence during embryogenesis and during wound healing and limb regeneration. Here, we explored the conservation of tissue injury‐induced senescence in a model of complex regeneration, the zebrafish. Fin amputation in adult fish led to the appearance of senescent cells at the site of damage, and their removal impaired tissue regeneration. Despite many conceptual similarities, this tissue repair response is different from developmental senescence. Our results lend support to the notion that cell senescence is a positive response promoting tissue repair and homeostasis., Tissue injury‐induced senescence is conserved in zebrafish. Fin amputation in adult fish lead to the appearance of senescent cells at the site of damage, and their removal impairs tissue regeneration. Tissue injury‐induced senescence is different from developmental senescence.

The zebrafish model can play a role in education because of its suitability for manipulation and attractiveness to students compared to traditional lecture-based instruction. Furthermore, zebrafish offer advantages over other model species. Seeing as fewer and fewer students are entering science degree programs, this project has been developed to encourage scientific vocations among secondary school students. To do so, an aquarium was given to 114 schools so that they could look after adult zebrafish, mate them, and visualize embryo development. For training on more sophisticated techniques, a virtual tool was developed to simulate a real genetics laboratory on a personal computer. Results based on teachers' feedback indicate that the students were fully dedicated to the project and achieved better understanding of genetic concepts and techniques. These results demonstrate the potential of alternative teaching methods for engaging students in science learning.

31 p.-6 fig.-14 fig. supl., The cell membrane regulates the exchange of molecules and information with the external environment. However, this control barrier hinders the delivery of exogenous bioactive molecules that can be applied to correct cellular malfunctions. Therefore, the traffic of macromolecules across the cell membrane represents a great challenge for the development of the next generation of therapies and diagnostic methods. Cell‐penetrating peptides are short peptide sequences capable of delivering a broad range of biomacromolecules across the cellular membrane. However, penetrating peptides still suffer from limitations, mainly related to their lack of specificity and potential toxicity. Glycosylation has emerged as a potential promising strategy for the biological improvement of synthetic materials. In this work we have developed a new convergent strategy for the synthesis of penetrating peptides functionalized with glycan residues by an oxime bond connection. The uptake efficiency and intracellular distribution of these glycopeptides have been systematically characterized by means of flow cytometry and confocal microscopy and in zebrafish animal models. The incorporation of these glycan residues into the peptide structure influenced the internalization efficiency and cellular toxicity of the resulting glycopeptide hybrids in the different cell lines tested. The results reported herein highlight the potential of the glycosylation of penetrating peptides to modulate their activity., We acknowledge funding from the Spanish Agencia Estatal de Investigación (AEI; CTQ2014‐59646‐R, SAF2017‐89890‐R, CTQ2016‐76263‐P, CTQ2015‐64597‐C2‐2P), the Xunta de Galicia (ED431G/09, ED431C 2017/25, and 2016‐AD031), and the ERDF. I.G. received a predoctoral fellowship from the Xunta de Galicia (ED481A‐2018/116). A.R. received a predoctoral fellowship from the Fundación Segundo Gil Dávila. J.G.‐V. and L.S. acknowledge financial support received from the Xunta de Galicia (Galicia, Spain) under the Grupos de Referencia Competitiva Programme: Project GRC2014/010. J.M. received a Ramón y Cajal (RYC‐2013‐13784), an ERC Starting Investigator Grant (DYNAP‐677786) and a Young Investigator Grant from the HFSP (RGY0066/2017).

Viral vector use is wide-spread in the field of gene therapy, with new clinical trials starting every year for different human pathologies and a growing number of agents being approved by regulatory agencies. However, preclinical testing is long and expensive, especially during the early stages of development. Nowadays, the model organism par excellence is the mouse (Mus musculus), and there are few investigations in which alternative models are used. Here, we assess the possibility of using zebrafish (Danio rerio) as an in vivo model for adenoviral vectors. We describe how E1/E3-deleted adenoviral vectors achieve efficient transduction when they are administered to zebrafish embryos via intracranial injection. In addition, helper-dependent (high-capacity) adenoviral vectors allow sustained transgene expression in this organism. Taking into account the wide repertoire of genetically modified zebrafish lines, the ethical aspects, and the affordability of this model, we conclude that zebrafish could be an efficient alternative for the early-stage preclinical evaluation of adenoviral vectors This research was funded by Xunta de Galicia, Local Government, grant number ED431C 2018/28, and Gobierno de Navarra, Local Government, grant numbers 0011-1383-2018-000011 PT038 and 0011-1383-2019-000006 PT013 (XANTOGEN) SI

This is the peer reviewed version of the following article: C. Penas, M. I. Sánchez, J. Guerra-Varela, L. Sanchez, M. E. Vázquez, J. L. Mascareñas, ChemBioChem 2016, 17, 37-41, which has been published in final form at https://doi.org/10.1002/cbic.201500455. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions We synthesized octa‐arginine conjugates of DNA‐binding agents (bisbenzamidine, acridine and Thiazole Orange) and demonstrated that their DNA binding and cell internalization can be inhibited by appending a (negatively charged) oligoglutamic tail through a photolabile linker. UV irradiation released the parent conjugates, thus restoring cell internalization and biological activity. Assays with zebrafish embryos demonstrates the potential of this prodrug strategy for controlling in vivo cytotoxicity We are thankful for the support given by the Spanish grants SAF2013–41943‐R, CTQ2012–31341 and CTQ2013–49317‐EXP, the Xunta de Galicia GRC2013–041, the ERDF and the European Research Council (Advanced Grant 340055). C.P. and M.I.S. thank the Spanish MCINN for their PhD fellowships SI

Mediterranean mussel (Mytilus galloprovincialis) is one of the main species of the European aquaculture, Galicia being (NW Spain) the main productive region (~ 200,000 t/year). The main problem for mussel production is the accumulation of toxins coming from microalgae blooms, which have even increased in recent years determining the banning of commercialization for long periods. We estimated the heritability and correlations (genetic and phenotypic) for toxin concentration after a toxic episode related to the main diarrheic toxin, okadaic acid, and after a depuration period at indoor facilities using a large amount of full-sib (170) and half-sib families (above 600 both via father and via mother). We took advantage of the experimental design to estimate the same parameters for other relevant traits for mussel production like growth-related traits and color. Heritabilities were moderate and significant after both the accumulation and detoxification periods (h2 ~ 0.35), suggesting the possibility of reducing toxin concentration through breeding programs in mussels. Growth-related traits also showed moderate heritabilities (~ 0.30), while color showed a very high heritability (~ 0.90), making them suitable for selection depending on producer and consumer demands. Interestingly, growth-related traits showed negative and significant genetic and phenotypic correlations with toxin concentration, while color showed positive ones, strongly suggesting that bigger and whitish mussels may accumulate less toxin. The results of our study support the viability of breeding programs to face the main problems of mussel industry, but this approach will determine a change in seed production, the wild seed being replaced or complemented with hatchery-produced seed.

Aim: To assess the capacity of a new drug delivery nanocapsule (NC) with a double shell of hyaluronic acid and protamine to overcome biological barriers using the zebrafish model. Materials & methods: NCs were prepared by the solvent displacement method, tagged with fluorescent makers and physicochemically characterized. Toxicity was evaluated according to the Fish Embryo Acute Toxicity test, and permeability was tested by exposing zebrafish, with and without chorion, to the fluorescent NCs. Results: Toxicity of NCs was very low as compared with that of a control nanoemulsion. Double-shell NCs were able to cross chorion and skin. Conclusion: Beyond the potential value of hyaluronic acid:protamine NCs for overcoming epithelial barriers, this works highlights the utility of zebrafish for fast screening of nanocarriers.

Ruthenium-based drugs exhibit interesting properties as potential anticancer pharmaceuticals. We herein present the synthesis and characterization of a new family of ruthenium complexes with formulas [{Ru(bipy)

Background: Zebrafish (Danio rerio) is a model organism that has emerged as a tool for cancer research, cancer being the second most common cause of death after cardiovascular disease for humans in the developed world. Zebrafish is a useful model for xenotransplantation of human cancer cells and toxicity studies of different chemotherapeutic compounds in vivo. Compared to the murine model, the zebrafish model is faster, can be screened using high-throughput methods and has a lower maintenance cost, making it possible and affordable to create personalized therapies. While several methods for cell proliferation determination based on image acquisition and quantification have been developed, some drawbacks still remain. In the xenotransplantation technique, quantification of cellular proliferation in vivo is critical to standardize the process for future preclinical applications of the model. Methods: This study improved the conditions of the xenotransplantation technique - quantification of cellular proliferation in vivo was performed through image processing with our ZFtool software and optimization of temperature in order to standardize the process for a future preclinical applications. ZFtool was developed to establish a base threshold that eliminates embryo auto-fluorescence and measures the area of marked cells (GFP) and the intensity of those cells to define a 'proliferation index'. Results: The analysis of tumor cell proliferation at different temperatures (34 °C and 36 °C) in comparison to in vitro cell proliferation provides of a better proliferation rate, achieved as expected at 36°, a maintenance temperature not demonstrated up to now. The mortality of the embryos remained between 5% and 15%. 5- Fluorouracil was tested for 2 days, dissolved in the incubation medium, in order to quantify the reduction of the tumor mass injected. In almost all of the embryos incubated at 36 °C and incubated with 5-Fluorouracil, there was a significant tumor cell reduction compared with the control group. This was not the case at 34 °C. Conclusions: Our results demonstrate that the proliferation of the injected cells is better at 36 °C and that this temperature is the most suitable for testing chemotherapeutic drugs like the 5-Fluorouracil. Fil: Cabezas-Sainz, Pablo. Universidad de Santiago de Compostela; España Fil: Guerra-Varela, Jorge. Universidad de Santiago de Compostela; España Fil: Carreira, María J.. Universidad de Santiago de Compostela; España Fil: Mariscal, Javier. Universidad de Santiago de Compostela; España Fil: Roel, María. Universidad de Santiago de Compostela; España Fil: Rubiolo, Juan Andrés. Universidad de Santiago de Compostela; España Fil: Sciara, Andres Angel. Universidad Nacional de Rosario; Argentina. Instituto de Biología Molecular y Celular de Rosario (IBR-CONICET) - Facultad de Bioquímica y Ciencias Farmacéuticas, ; Argentina Fil: Abal, Miguel. Universidad de Santiago de Compostela; España Fil: Botana, Luis M.. Universidad de Santiago de Compostela; España Fil: López, Rafael. Universidad de Santiago de Compostela; España Fil: Sánchez, Laura. Universidad de Santiago de Compostela; España

In the last few decades, the field of nanomedicine applied to cancer has revolutionized cancer treatment: several nanoformulations have already reached the market and are routinely being used in the clinical practice. In the case of genetic nanomedicines, i.e., designed to deliver gene therapies to cancer cells for therapeutic purposes, advances have been less impressive. This is because of the many barriers that limit the access of the therapeutic nucleic acids to their target site, and the lack of models that would allow for an improvement in the understanding of how nanocarriers can be tailored to overcome them. Zebrafish has important advantages as a model species for the study of anticancer therapies, and have a lot to offer regarding the rational development of efficient delivery of genetic nanomedicines, and hence increasing the chances of their successful translation. This review aims to provide an overview of the recent advances in the development of genetic anticancer nanomedicines, and of the zebrafish models that stand as promising tools to shed light on their mechanisms of action and overall potential in oncology This work was supported by the Xunta de Galicia, Spain (Competitive Reference Groups, GRC2014/010), the Carlos III Health Institute/FEDER (PI15/00828), and the Spanish Ministry of Education, Culture and Sport (FPU15/06595) SI

// Maria Roel 1 , Juan A. Rubiolo 1 , Jorge Guerra-Varela 2 , Siguara B. L. Silva 3, 4 , Olivier P. Thomas 3, 5 , Pablo Cabezas-Sainz 2 , Laura Sanchez 2 , Rafael Lopez 6 , Luis M. Botana 1 1 Department of Pharmacology, Universidade de Santiago de Compostela, Campus Lugo, 27002 Lugo, Spain 2 Department of Genetics, Universidade de Santiago de Compostela, Campus Lugo, 27002 Lugo, Spain 3 Geoazur, UMR Universite Nice Sophia Antipolis-CNRS-IRD-OCA, 06560 Valbonne, France 4 Laboratoire de Pharmacognosie, UMR CNRS 8076 BioCIS, LabEx LERMIT, Universite Paris-Sud, Faculte de Pharmacie, 92290 Châtenay-Malabry, France 5 School of Chemistry, Marine Biodiscovery, National University of Ireland Galway, SW4 Galway, Ireland 6 Translational Medical Oncology, Health Research Institute of Santiago (IDIS), Complexo Hospitalario Universitario de Santiago de Compostela (SERGAS), 15706 Santiago de Compostela, Spain Correspondence to: Luis M. Botana, email: luis.botana@usc.es Keywords: crambescidins, cell cycle inhibition, apoptosis, zebrafish xenograft model, cancer treatment Received: March 13, 2016 Accepted: September 27, 2016 Published: November 04, 2016 ABSTRACT The marine environment constitutes an extraordinary resource for the discovery of new therapeutic agents. In the present manuscript we studied the effect of 3 different sponge derived guanidine alkaloids, crambescidine-816, -830, and -800. We show that these compounds strongly inhibit tumor cell proliferation by down-regulating cyclin-dependent kinases 2/6 and cyclins D/A expression while up-regulating the cell cyclin-dependent kinase inhibitors -2A, -2D and -1A. We also show that these guanidine compounds disrupt tumor cell adhesion and cytoskeletal integrity promoting the activation of the intrinsic apoptotic signaling, resulting in loss of mitochondrial membrane potential and concomitant caspase-3 cleavage and activation. The crambescidin 816 anti-tumor effect was fnally assayed in a zebrafish xenotransplantation model confirming its potent antitumor activity against colorectal carcinoma in vivo . Considering these results crambescidins could represent promising natural anticancer agents and therapeutic tools.

Sixteen polymorphic microsatellite primers were developed for Pseudoplatystoma reticulatum and provided an efficient multiplex protocol to amplify these loci in four multiplex PCRs. Most microsatellites here obtained showed moderate to high polymorphism levels in P. reticulatum and P. corruscans, as well as interspecific allelic differences, and represents useful genetic markers for population studies in these fishes.

Two ecotypes of Nucella lapillus can occur allopatrically or sympatrically, according to different degrees of wave exposure and microhabitats, on rocky shores of NW Spain. We studied differences in shell size and shape in adults and families of shelled embryos captured in the wild, and molecular variation at 9 microsatellite loci and 4 mtDNA regions. Both adults and shelled embryos of the two ecotypes showed significant differences in size and shell-shape components (the most important being size of the shell aperture). Strong among-family variation suggests that shell-shape variation has an important genetic component, and the pattern of genetic differentiation was in agreement with traits being affected by divergent natural selection. No significant overall molecular differentiation was observed between the two ecotypes for mtDNA, although we found significant genetic structure at two microsatellite loci. Microsatellite differentiation between ecotypes at these two loci was larger in sympatry than in allopatry. These results and the comparison of tenacity suggest that the two forms are distinctly adapted to differences in shore level and degree of exposure to wave action.

The present paper integrates phylogenetic and population genetics analyses based on mitochondrial and nuclear molecular markers in silversides, genus Odontesthes, from a non-sampled area in the SW Atlantic Ocean to address species discrimination and to define Managements Units for sustainable conservation. All phylogenetic analyses based on the COI mitochondrial gene were consistent to support the monophyly of the genus Odontesthes and to include O. argentinensis, O. perugiae-humensis and some O. bonariensis haplotypes in a basal polytomy conforming a major derivative clade. Microsatellites data revealed somewhat higher genetic variability values in the O. argentinensis-perugia populations than in O. bonariensis and O. perugia-humensis taxa. Contrasting population genetics structuring emerged from mitochondrial and microsatellites analyses in these taxa. Whereas mitochondrial data supported two major groups (O. argentinensis-perugia-humensis vs. O. bonariensis-perugiae-humensis populations), microsatellite data detected three major genetic entities represented by O. bonariensis, O. perugiae-humensis and an admixture of populations belonging to O. argentinensis-perugiae respectively. Therefore, the star COI polytomy in the tree topology involving these taxa could be interpreted by several hypothetic scenarios such as the existence of shared ancestral polymorphisms, incomplete lineage sorting in a radiating speciation process and/or reticulation events. Present findings support that promiscuous and recent contact between incipient species sharing asymmetric gene flow exchanges, blurs taxa boundaries yielding complicated taxonomy and Management Units delimitation in silverside genus Odontesthes from SW Atlantic Ocean basins. This research received financial support from the project Fondo María Viñas_2009_1_2793 (FMV_2009_1_2793_Project) granted by the Agencia Nacional de Investigación e Inovación (ANII) of Uruguay SI

The dogwhelk Nucella lapillus is a predatory marine gastropod living on rocky shores in the North Atlantic. As with many other gastropod species, Nucella was affected by tributyltin (TBT) pollution during the 1970s and 1980s, and local populations underwent extinction. After a partial ban on TBT in the UK in 1987, vacant sites have been recolonized. Levels of genetic diversity and quantitative genetic variation in shell form were compared between recolonized sites and sites that showed continuous population at three localities across the British Isles. Overall, estimates of genetic diversity were only slightly lower in recolonized populations, suggesting that populations have recovered from previous impacts due to the relatively high levels of migration from non-impacted sites. Molecular and quantitative analyses are broadly concordant and a positive correlation was observed (although not statistically significant) between molecular and quantitative estimates of genetic diversity, indicating the potential usefulness of quantitative methods to complement molecular population genetics analyses.

Background Chromosome rearrangements are an important part of the speciation process in many taxa. The study of chromosome evolution in bivalves is hampered by the absence of clear chromosomal banding patterns and the similarity in both chromosome size and morphology. For this reason, obtaining good chromosome markers is essential for reliable karyotypic comparisons. To begin this task, the chromosomes of the mussels Brachidontes puniceus and B. rodriguezi were studied by means of fluorochrome staining and fluorescent in situ hybridization (FISH). Results Brachidontes puniceus and B. rodriguezi both have 2n = 32 chromosomes but differing karyotype composition. Vertebrate-type telomeric sequences appear at both ends of every single chromosome. B. puniceus presents a single terminal major rRNA gene cluster on a chromosome pair while B. rodriguezi shows two. Both mussels present two 5S rDNA and two core histone gene clusters intercalary located on the long arms of two chromosome pairs. Double and triple-FISH experiments demonstrated that one of the 5S rDNA and one of the major rDNA clusters appear on the same chromosome pair in B. rodriguezi but not in B. puniceus. On the other hand, the second 5S rDNA cluster is located in one of the chromosome pairs also bearing one of the core histone gene clusters in the two mussel species. Conclusion Knowledge of the chromosomal distribution of these sequences in the two species of Brachidontes is a first step in the understanding of the role of chromosome changes on bivalve evolution.