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      Environmental Conditioning of Skeletal Anomalies Typology and Frequency in Gilthead Seabream (Sparus aurata L., 1758) Juveniles

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          Abstract

          In this paper, 981 reared juveniles of gilthead seabream ( Sparus aurata) were analysed, 721 of which were from a commercial hatchery located in Northern Italy (Venice, Italy) and 260 from the Hellenic Center for Marine Research (Crete, Greece). These individuals were from 4 different egg batches, for a total of 10 different lots. Each egg batch was split into two lots after hatching, and reared with two different methodologies: intensive and semi-intensive. All fish were subjected to processing for skeletal anomaly and meristic count analysis. The aims involved: (1) quantitatively and qualitatively analyzing whether differences in skeletal elements arise between siblings and, if so, what they are; (2) investigating if any skeletal bone tissue/ossification is specifically affected by changing environmental rearing conditions; and (3) contributing to the identification of the best practices for gilthead seabream larval rearing in order to lower the deformity rates, without selections. The results obtained in this study highlighted that: i) in all the semi-intensive lots, the bones having intramembranous ossification showed a consistently lower incidence of anomalies; ii) the same clear pattern was not observed in the skeletal elements whose ossification process requires a cartilaginous precursor. It is thus possible to ameliorate the morphological quality (by reducing the incidence of severe skeletal anomalies and the variability in meristic counts of dermal bones) of reared seabream juveniles by lowering the stocking densities (maximum 16 larvae/L) and increasing the volume of the hatchery rearing tanks (minimum 40 m 3). Feeding larvae with a wide variety of live (wild) preys seems further to improve juvenile skeletal quality. Additionally, analysis of the morphological quality of juveniles reared under two different semi-intensive conditions, Mesocosm and Large Volumes, highlighted a somewhat greater capacity of Large Volumes to significantly augment the gap with siblings reared in intensive (conventional) modality.

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          Enzyme clearing of alcian blue stained whole small vertebrates for demonstration of cartilage.

          Preparation of small vertebrates cleared after alcian blue staining of cartilage is facilitated by trypsin digestion. Specimens are fixed in formation, washed, skinned, and eviscerated. After staining in a solution of alcian blue in acetic acid-alcohol for 24-48 hours, they are transferred to water through graded alcohols. Excess alcian blue is removed over a period of up to three weeks by changes every 2-3 days of 1% trypsin in approximately one-third-saturated sodium borate. Bony tissues may be stained after this in a solution of alizarin red S in 0.5% KOH. Specimens are bleached if necessary and dehydrated through graded KOH-glycerine mixtures for storage in glycerine. Since alcohol treatment in addition to formalin fixation does not affect results with this method, it should be useful to researchers who want to study the cartilage or cartilaginous skeletons in museum specimens, which are routinely fixed in formalin and stored in alcohol.
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            Developmental morphology of the axial skeleton of the zebrafish, Danio rerio (Ostariophysi: Cyprinidae).

            Before our rapidly increasing knowledge of gene interactions can be connected with the morphologic defects in mutant zebrafish, the normal course of skeletal development must be understood. Here, we describe the developmental morphology of the axial skeleton of zebrafish and review it in relation to the morphology of related species. The relative sequence of ossification in the skeleton is described. Two separate centers of development were found in the axial skeleton (Weberian apparatus and caudal fin) in contrast to tetrapods, which have a single anterior center. Slight variation was found in the overall relative timing of development. The extensive ichthyological literature on teleost anatomy and recent genetic data form the basis for our review and interpretation of homologies of various elements of the axial skeleton. Because homology forms the basis for all evolutionary comparisons, these data are critical for integration in evo-devo studies. Copyright 2003 Wiley-Liss, Inc.
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              Formation of dermal skeletal and dental tissues in fish: a comparative and evolutionary approach.

              Osteichthyan and chondrichthyan fish present an astonishing diversity of skeletal and dental tissues that are often difficult to classify into the standard textbook categories of bone, cartilage, dentine and enamel. To address the question of how the tissues of the dermal skeleton evolved from the ancestral situation and gave rise to the diversity actually encountered, we review previous data on the development of a number of dermal skeletal elements (odontodes, teeth and dermal denticles, cranial dermal bones, postcranial dermal plates and scutes, elasmoid and ganoid scales, and fin rays). A comparison of developmental stages at the tissue level usually allows us to identify skeletogenic cell populations as either odontogenic or osteogenic on the basis of the place of formation of their dermal papillae and of the way of deposition of their tissues. Our studies support the evolutionary affinities (1) between odontodes, teeth and denticles, (2) between the ganoid scales of polypterids and the elasmoid scales of teleosts, and (3) to a lesser degree between the different bony elements. There is now ample evidence to ascertain that the tissues of the elasmoid scale are derived from dental and not from bony tissues. This review demonstrates the advantage that can be taken from developmental studies, at the tissue level, to infer evolutionary relationships within the dermal skeleton in chondrichthyans and osteichthyans.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, USA )
                1932-6203
                2013
                7 February 2013
                : 8
                : 2
                : e55736
                Affiliations
                [1 ]Department of Biology, University of Rome “Tor Vergata”, Rome, Italy
                [2 ]Department of Biology, University of Patras, Patras, Greece
                The Hebrew University, Israel
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Conceived and designed the experiments: SC CB. Performed the experiments: AS PM SC. Analyzed the data: LP CB MS VR EP MS. Contributed reagents/materials/analysis tools: CB AS MS SC. Wrote the paper: LP CB PM SC. Experimental design: SC CB. Larvae and rearing juveniles: AS PM. Observation on skeletal anomalies and meristic counts: LP CB EP VR. Data mining: MS. Interpretation of data: CB SC.

                Article
                PONE-D-12-21465
                10.1371/journal.pone.0055736
                3567126
                e9321f6f-1d15-46ea-8985-5508f29ba376
                Copyright @ 2013

                This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 21 June 2012
                : 30 December 2012
                Page count
                Pages: 22
                Funding
                This study was funded by the Italian Ministry for Agriculture, Food and Forestry Policy (Law 41/82) and by the Commission of the European Communities (specific RTD programme “Specific Support to Policies”, SSP-2005-44483 “SEACASE – Sustainable extensive and semi-intensive coastal aquaculture in Southern Europe”). It does not necessarily reflect the Italian Ministry for Agriculture, Food and Forestry Policy and the European Commission views. In no way anticipates the Commissions future policy in this area. This work benefits also from participation in LARVANET COST action FA0801. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Agriculture
                Aquaculture
                Fish Farming
                Biology
                Anatomy and Physiology
                Musculoskeletal System
                Biomechanics
                Bone
                Developmental Biology
                Evolutionary Developmental Biology
                Ecology
                Industrial Ecology
                Zoology
                Ichthyology

                Uncategorized
                Uncategorized

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