This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.A C C E P T E D M A N U S C R I P T Skeletal anomalies in reared dusky grouper juveniles ACCEPTED MANUSCRIPTThe first attempts to reproduce dusky grouper (Epinephelus marginatus, Lowe 1834) under controlled conditions started in 1995, but the egg and larvae quality was very low. Mass production is still encountering many difficulties, mainly concentrated in the larval period when very high mortality rates are observed, confirming what has been observed in the rearing of other grouper species. The main bottlenecks have been identified as the difficulty to properly nourish the larvae, stress shock syndrome, and the high deformation rates.We analysed 633 dusky grouper larvae and juveniles (0.2-7.2 cm total length, TL), sampled during two larval rearing cycles carried out in 2001 and 2002 in Italy. The specimens at different development stages were stained in toto for bone and cartilage and examined for skeletal anomalies during dusky grouper ontogenesis. The incidence of anomalies in groupers hatched from the same egg batch but reared using two different methods (green waters and semi-intensive rearing) and three stocking densities (8, 16 and 28 larvae/l) was compared, with a view to providing tools for identifying the most appropriate larval rearing method in order to at least limit the onset of skeletal anomalies.Our results suggest that during development no particular skeletal anomaly patterns (or fate) can be clearly identified as a high variability was observed in malformation typologies and the regions affected.No significant differences in the morphological quality between groupers reared using semi-intensive (LV02 lot) and green water (GW02-01 lot) methodologies were observed, whilst groupers reared at the highest stocking density (28 larvae/l) showed the highest frequency of deformed individuals (75.8%), the highest malformation charge (average of 5.5 anomalies per deformed individual), the largest range of anomaly typologies (38), and the highest incidence of individuals with at least one severe anomaly (30.9%). Whilst in green waters no evident effects of larvae density were observed on survival rates, the survival rate in large volume reared individuals (17.5%) was considerably higher with respect to those reared in green waters (0.2%) at 7-8 larvae/litre. This indicates that the semi-intensive methodology should be considered more effective in enhancing the survival rate of dusky grouper larvae.

This paper reports on a part of PRISMA project, funded by the Italian Ministry of Education, University and Research (M.U.R.S.T.), involving the biomonitoring of fish populations in two different Adriatic Sea sites by using some integrated descriptors at organismic level (skeletal anomalies and meristic count variation). Given the scarcity of up-to-date data on biology, physical and chemical oceanography, and environmental geophysics, as well on the degree of pollution of this Sea, the goal of the project was to contribute at enhancing the knowledge on environmental biology of Adriatic Sea. In particular, this part of the project analyzed different samplings of some mullets species for the presence of skeletal malformations under the hypothesis that stressed environment should induce alteration in skeletal development pattern in fish, as indicated by many authors. So, the investigation on mullets skeletal structures was utilized to monitor two different Adriatic Sea sites, where exhaustive information (chemical analysis of water and sediment, i.e.) on environmental conditions lack.We observed a total of 2169 individuals, 79% of which were fry and the remainder sub-adults, collected during the seasonal migration in freshwater (spring and autumn 1997) by purse-seine. Chelon labrosus accounted for 18% of the samples, Liza aurata 24%, Liza ramada 22% and Mugil cephalus 36%. Fish were sampled in two Adriatic sites, Sacca di Goro (northern Italy, Po River delta) and Lake Lesina (southern Italy). Other samplings from Tyrrhenian Sea (Fiumicino, Tiber river mouth) and from an aquaculture facility (Grosseto) were used in order to obtain additional information on skeletal plasticity of Mugilidae. Observations included meristic counts and skeletal anomalies and some multi-parametric analyses were performed on the data set. The results showed significant differences among the type and rate of skeletal anomalies observed in the fish samples: as far as Adriatic samples are concerned, higher malformation charge was observed in the Sacca di Goro individuals than in Lesina ones. Further, L. aurata showed a non-specific skeletal pattern and the lowest frequency of deformed individual (8.6% of total

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