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A histological study was conducted in red mullet, Mullus barbatus, collected from two sites characterized by different anthropogenic impacts. The aim of the study was to assess sex-, size-, season- and site-related variation in gonadal macrophage aggregate (MA) size, number, and relative area. Gonadal MAs were most abundant in males than in females. The number of MA was significantly higher in males from the most impacted site in October, with larger individuals showing more MA than smaller ones. MAs were always found in ripe testes, whereas they occurred only in regressing ovaries. These preliminary findings suggest that the presence of ovarian MA in red mullet is most likely related to ovary regression after spawning, whereas the presence of testicular MA is not necessarily associated to gonad regression, and may vary with season, size, and water quality.

A histological study was conducted in red mullet, Mullus barbatus, collected from two sites characterized by different anthropogenic impacts. The aim of the study was to assess sex-, size-, season- and site-related variation in gonadal macrophage aggregate (MA) size, number, and relative area. Gonadal MAs were most abundant in males than in females. The number of MA was significantly higher in males from the most impacted site in October, with larger individuals showing more MA than smaller ones. MAs were always found in ripe testes, whereas they occurred only in regressing ovaries. These preliminary findings suggest that the presence of ovarian MA in red mullet is most likely related to ovary regression after spawning, whereas the presence of testicular MA is not necessarily associated to gonad regression, and may vary with season, size, and water quality.

The peptide hormone cholecistokinin (CCK) plays a key role in the central and peripheral nervous system. It is known to be involved in the digestive physiology and in the regulation of food intake. Moreover, the CCK expression has also been detected in the retina of different vertebrates, including fish, although its biological activity in this tissue remains to be elucidated. In literature no data are yet available about the CCK-immunoreactivity in the zebrafish retina during development. Therefore, the aim of the study was to investigate the distribution of sulfated cholecystokinin octapeptide (CCK8-S) as a well preserved form during evolution in the zebrafish retina from 3 days post hatching (dph) until adult stage, using immunohistochemistry in order to elucidate the potential role of this protein in the development and maintenance of normal retinal homeostasis. The cellular distribution of CCK in the retina was similar from 3 dph to 40 days post fertilization (dpf) when immunoreactivity was found in the photoreceptors layer, in the outer plexiform layer, in the inner plexiform layer and, to a lesser extent, in the ganglion cell layer (GCL). Immunohistochemical localization at 50 dpf as well as in the adult stage was observed in a subpopulation of amacrine cells in the proximal inner nuclear layer, in the inner plexiform layer, in displaced amacrine cells and in retinal ganglion cells in the GCL. Our results demonstrate for the first time the occurrence of CCK in the zebrafish retina from larval to adult stage with a different pattern of distribution, suggesting different roles of CCK during retinal cells maturation.

This study reports the occurrence of some endocrine disrupting chemicals in red mullet samples and sediments collected in two representative sites of the northern Sicilian coast (Italy). For this purpose, an improved method, using solid extraction and high-performance liquid chromatography analyses for the simultaneous determination of bisphenol A (BPA), 4-nonylphenol (4-NP) and 4-t-octylphenol (4-t-OP) in fish tissues and sediments, has been developed and validated. Method performance was demonstrated over the concentration range 0.1–200 ng/mL, with detection limits from 0.06 to 0.1 ng/mL. Recoveries ranged from 83.4% to 102.6%, with relative standard deviations of 7.7–14.0% for the entire procedure. Results showed that BPA, 4-t-OP and 4-NP were detected in all fish samples and sediments from two sampling sites, indicating that these chemicals have contaminated Mediterranean aquatic ecosystem and have accumulated in fish. The study provided more comprehensive fundamental data for risk assessment and contamination control of phenolic EDCs in aquatic environment.

A brief review of investigations carried out at IAMC-CNR Messina on the reproductive biology of farmed blackspot sea bream Pagellus bogaraveo, a new candidate species for aquaculture, is presented. Different aspects of sexuality, including hermaphroditism, gametogenesis, spermatozoal ultrastructure, maturity staging, seasonal development of gonads, and spawning in captivity, were studied. Protandrous hermaphroditism appeared to be the prevalent sexual pattern, although a high incidence of gonochorism occurred. Males were most abundant in smaller size classes and reached sexual maturity at 28.0 cm, while females predominated in the larger size classes and reached maturity at 29.5 cm. The reproductive season lasted from February until April, as in the natural environment. Spontaneous spawning in tank occurred occasionally, whereas approximately 50% of females produced eggs upon stripping. The remaining 50% failed to ovulate, or even to achieve oocyte maturation, indicating an impaired reproductive functionality. A high incidence of oocyte atresia was also recorded. Males produced a small quantity of highly viscous milt, containing only 3% motile spermatozoa. The spermatozoon of P.bogaraveo has an ovoid head, a midpiece containing only one mitochondrion, and a 50.64 ± 4.82 ?m long flagellum, inserted perpendicularly and eccentrically into the nucleus.

The present review reports the results of two studies carried out at the Institute for the Coastal Marine Environment of Messina (Italy) on the suitability of plant-derived raw materials as dietary components for an innovative fish species for aquaculture, black spot sea bream Pagellus bogaraveo. The research focused on the effects of the use of plant (protein/lipid) dietary sources on growth performance, body composition and gastro-intestinal enzyme patterns in nutrient digestion of wild juveniles of P. bogaraveo. In particular, the effect of a total replacement of Fish oil by Echium and linseed oils and a partial substitution of fish meal with a graded level of Rice Protein Concentrate, will be discussed.

The physiological effects of short-term starvation on some haematological, biochemical and non-specific immune response parameters together with the histological structure of the skin, were investigated in the European eel, Anguilla anguilla. Blood haemoglobin and haematocrit, serum glucose and cortisol, hemolysins, haemagglutinins, and lysozyme in the plasma, kidney and epidermal extract, were measured in fish after 31, 42 and 58 days of starvation, and compared to those of fed fish. Starvation did not affect haemoglobin and haematocrit values, while an increase in glucose and cortisol levels was found in starved eels by day 42. Haemolytic and haemagglutinating activities decreased in starved eels. On the other hand, starvation caused an increase in the lysozyme content in the epidermal extracts, while no significant variations were observed in kidney and plasma. On the whole, no major changes in metabolic, haematological and non-specific immune parameters were observed when short-term (less than 2 months) starvation was applied to the European eel, suggesting an adaptive response to starvation, rather than a typical alarm-stress response, allowing this species to withstand food deprivation.

Scanning and transmission electron microscopy were used to investigate the fine structure of sperm of the Mediterranean amberjack Seriola dumerilii. Each spermatozoon has an ovoid head which lacks an acrosome, a short, irregularly-shaped midpiece and a long flagellar tail. The midpiece houses eight spherical mitochondria, which are separated from the axoneme by the cytoplasmic canal. The centrioles are arranged approximately at right angles to each other. The proximal centriole lies inside, and the distal centriole outside, the nuclear fossa. The flagellum is inserted eccentrically into the head and is tangential to the nucleus, so that the spermatozoon is asymmetrical. It contains the conventional 9 + 2 axoneme, shows intratubular differentiations in the A microtubules of doublets 1, 2, 5 and 6, and possesses one pair of lateral fins. On the basis of its ultrastructural organization, the amberjack sperm resembles type II sperm as defined previously, except for the presence of the proximal centriole inside the nuclear fossa. This could result from a partial rotation of the nucleus during spermiogenesis.

(2001). Preliminary investigation of the digestive enzymes in Pagellus erythrinus (Linneo 1758) larvae. Marine and Freshwater Behaviour and Physiology: Vol. 34, No. 4, pp. 265-268.

A study of the reproductive biology of the amberjack, Seriola dumerilii, held in captivity was carried out, describing oogenesis as well as the different stages of the ovarian cycle. Seven stages of oocyte development, as well as oogonia, were distinguished. Cortical alveoli were hardly detectable within the oocyte, as they were small, sparse and contained few mucopolysaccharides. It is suggested that their role in the fertilization process might be less important than in other teleost species. Fish aged 3 and 4 years were found to be sexually immature, with ovaries containing only previtellogenic oocytes. Vitellogenesis started in December in fish aged 4 + years. Late-developing ovaries showing deposition of yolk protein granules were found at the end of the 5th year of life (May) in specimens measuring 80.0 6 3.5 cm standard length. This should be regarded as the minimum size at which sexual maturity is reached in S. dumerilii. Final vitellogenesis and oocyte maturation were, however, inhibited in captivity, and extensive follicular atresia took place as the natural spawning season approached. It is suggested that insufficient gonadotrophic stimulation because of confinement stress may be the cause of failed maturation and spawning in this species under culture conditions.

A histological study was made of the reproductive cycle of captive sharpsnout seabream Diplodus puntazzo during the 3rd year of life. All specimens contained both ovarian and testicular tissue. Ovarian tissue prevailed in 64 out of 120 fish sampled throughout one year. The ovary underwent a long resting phase from October to August, during which time it contained oogonia, primary growth phase (PGP) and occasional yolk vesicle stage oocytes. Intense ovarian development took place in September leading to the formation of advanced vitellogenic oocytes. Final maturation and spawning were not achieved in any of the fish. The testicular area in the predominantly female gonads contained only spermatogonia and was found to be histologically normal in 76% of individuals, but severely regressed in the remaining fish. The testicular portion was found to be dominant in 56 out of 120 fish sampled throughout one year. These underwent a resting phase from November to July, a maturing/pre-spawning phase in August, and a spawning phase with spontaneous sperm release in September to October, as in the natural environment. The ovarian portion in the functional male gonads contained only oogonia and oocytes which did not develop beyond the perinucleolus stage at any point during the reproductive cycle. Apart from the pre-spawning/spawning phase, during which the gonadosomatic index (GSI) was comparable in the two groups, the GSI values were approximately ten-fold higher in those fish in which the ovary predominated than in those in which the testis predominated.

The reproductive features of the giant red shrimp,Aristaeomorpha foliacea, were investigated in the southern Tyrrhenian sea by experimental trawl sampling. The annual length-frequency distribution showed a multimodal trend in females, ranging between 16 and 67 mm carapace length (CL), and a unimodal trend in males (18–45 mm CL). Mature males occurred in different proportions all year round, while females displayed seasonal maturity (June—September), with a peak in July. Six oocyte developmental stages were identified, the most advanced of which (Pv, postvitellogenic) had never been described before in this species. Ovary development followed a group-synchronous pattern, with the yolked oocyte stock clearly separated from the reservoir of unyolked oocytes, suggesting thatA. foliaceais a total spawner, with determinate fecundity. Based upon histological findings, a revision of macroscopic maturity staging employed in Mediterranean bottom trawl surveys (MEDITS) is proposed.

Histological events associated with the process of sex inversion are described in the protandrous teleost Diplodus sargus reared in captivity. This species possesses typical sparid ovotestes in which the testis and ovary occur in separate zones. After spawning, the testicular tissue within the bisexual gonad undergoes regressive changes such as spermatogonial degeneration, deposition of yellowish pigment and proliferation of connective cells and fibres. At the same time ovarian tissue begins to develop. Transitional individuals were 26–7% of examined fish, while the other 73–3% were functional males.

The ontogeny of the alimentary tract and its associated structures (liver, pancreas, gall bladder) was compared between sharpsnout sea bream Diplodus puntazzo and Pagellus erythrinus and some differences were highlighted. The first goblet cells appeared earlier in sharpsnout seabream (5-7 DAH) and later in common pandora (9-11 DAH), as well as taste buds which appeared at 11 DAH in sharpsnout seabream and at 17 DAH in common pandora. On the contrary, stomach segmentation in a cardiac, fundic and pyloric portions was more precocious in common pandora than in sharpsnout seabream, although gastric glands were formed almost simultaneously. Early caecal development was much more precocious in sharpsnout seabream (10 DAH) than in common pandora (22 DAH). At 3 DAH in sharpsnout seabream and later (10 DAH) in common pandora, primordial mucosal folding of intestine began, which appeared well developed in the anterior portion, by 22 DAH in common pandora and by 28 DAH in sharpsnout seabream. The functional meanings of these aspects were discussed comparatively.

A short overview on the regional distribution of the gastro-intestinal peptide hormone cholecystokin (CCK) in fish is presented. In particular, the results of molecular and immunological studies on seabreams, Diplodus puntazzo and Diplodus sargus, are reported, which, by demonstrating CCK in the hindgut, open new questions regarding the functional role of this hormone in that part of the intestine. The putative involvement of hindgut CCK in the feedback control of digestive processes was tested by measuring CCK gene and protein expression in fed and fasted fish. The results of this study led to hypothesize different roles for the two CCK isoforms in D. sargus, one of which related to regulation of digestive processes from pyloric caeca through hindgut. On the other hand, a functional role alternative to regulation of digestive processes may be inferred for the other isoform.

The appearance and regional distribution of cholecystokinin-immuno reactive cells (CCK-IR) in the developing gut of larval Diplodus puntazzo were studied by means of immunohistochemistry, with the aim of understanding the role of this peptide hormone in the acquisition of digestive capacity. Immunohistochemical reaction showed CCK-IR cells from 10 days after hatching (DAH), near the pyloric sphincter and past the first bend in the midgut, as well as in the hindgut. At 25 DAH CCK-IR cells were scattered throughout the midgut, as well as in the hindgut. Since gastric glands appeared at 30 DAH, CCK-IR cells were most abundant in the anterior midgut, near and including the pyloric caeca, and just afore the ileo-rectal sphincter in the posterior midgut, as well as in the hindgut. In older larvae (39 DAH), CCK-IR cells were mainly distributed in the anterior midgut, including the pyloric caeca, as well as in the hindgut. No CCK-IR cells were detected in the foregut at any stage. The distribution pattern of CCK-IR cells differed from other species which also possess a rotated gut as D. puntazzo. In fact, although cells were abundant in regions where the ingested food is retained, so that they can be stimulated to modulating the release of digestive enzymes, a large number of cells occurred also in the hindgut.

The influence of photoperiod prolongation on gonadal maturation was investigated in the hermaphroditic fish Sparus aurata (L.), the common gilthead bream. Fish born in captivity under controlled conditions were kept in tanks under different photoperiod regimes from January 1986 to January 1987 and gonadal samples were taken for histological examination throughout the experiment. All fish developed into functional males. Gonadal maturation was delayed, though not completely inhibited, in fish kept under prolonged photoperiod compared with the control fish kept under natural photoperiod. Testicular regression and/or ovarian recrudescence were never observed. It is concluded that a seasonally‐changing light regime is not a necessary cue of the spermatogenic process in S. aurata. A constant 16L: 8D photoperiod, while postponing the breeding season by about 2 months, may cause some loss of spawning synchrony between members of the same brood‐stock.

The aim of this study was to evaluate the performances and the flesh quality of Pagellus bogaraveo fed with diets containing rice protein concentrate [RPC, 70% crude protein (CP) and 10% ether extract]. Three isoproteic and isoenergetic (CP 47%, 22 MJ/kg DM) diets were formulated with an increasing level of RPC: 0%, 20% and 35%. The fish (mean weight 75 g) from the Messina Straits were randomly distributed in 12 tanks (3 diets · 4 replications, 10 fish/tank). The daily ratio (1.5% of the fish biomass) was updated every 15 days. Biomass gain showed an opposite trend to the RPC diet inclusion. No differences appeared in the somatic indexes. Differences appeared between fatty acid profiles of the dorsal muscle. Fatty acid of series n-6 increased and fatty acid of series n-3 decreased in fillets of fish fed with increasing levels of RPC. The inclusion of RPC in the diets, as a partial replacement of fish meal (20%), is possible without affecting the growth performance and fillet quality.

The ontogenesis of the alimentary tract and its associated structures (liver, pancreas, gall bladder) was studied histologically in common pandora from hatching (0 DAH, days after hatching) until day 50 (50 DAH). Larvae were obtained by natural spawning from a broodstock adapted to captivity. They were stocked in 1500 l tanks supplied with Isochrysis galbana and Tetraselmis suecica from hatching until the Artemia feeding stage, at a temperature of 18.5–20 °C. Larvae were fed Selco-enriched Brachionus plicatilis from day 3, Artemia nauplii from day 28 and formulated feed from day 35. At hatching, the digestive tract was a histologically undifferentiated straight tube lying dorsally to the yolk sac. At first feeding (3–4 DAH) both the mouth and anus had opened and the digestive tract was differentiated into four portions: buccopharynx, oesophagus, incipient stomach and intestine. The pancreas, liver and gall bladder were also differentiated at this stage. Within 2 days after the commencement of exogenous feeding, the anterior intestinal epithelium showed large vacuoles indicating the capacity for absorption of lipids, whereas supranuclear ninhydrin-Schiff (NS) positive inclusions indicating protein absorption were observed in the posterior intestinal epithelium. Both the bile and main pancreatic ducts had opened in the anterior intestine, just after the pyloric sphincter, at this stage. Intestinal coiling was apparent since 4 DAH, while mucosal folding began at 10 DAH. Scattered PAS-positive mucous cells occurred in the oral cavity and the intestine, while they were largely diffused in the oesophagus. Gastric glands and pyloric caeca appeared at 28 DAH, indicating the transition from larval to juvenile stage and the acquisition of an adult mode of digestion.

The ontogenesis of the alimentary tract and its associated structures (liver, pancreas, gall bladder) was studied in common pandora Pagellus eythrinus L., a promising species for diversification in Mediterranean aquaculture. Mass production of pandora has been limited so far by high larval and juvenile mortalities, which appear to be related to nutritional deficiencies. The development of the larval digestive system was studied histologically from hatching (0 DAH) until day 50 (50 DAH) in reared specimens, obtained by natural spawning from a broodstock adapted to captivity. At first feeding (3–4 DAH) both the mouth and anus had opened and the digestive tract was differentiated in four portions: buccopharynx, oesophagus, incipient stomach and intestine. The pancreas, liver and gall bladder were also differentiated at this stage. Soon after the commencement of exogenous feeding (5–6 DAH), the anterior intestinal epithelium showed large vacuoles indicating the capacity for absorption of lipids, whereas acidophilic supranuclear inclusions indicating protein absorption were observed in the posterior intestinal epithelium. Both the bile and main pancreatic ducts had opened in the anterior intestine, just after the pyloric sphincter, at this stage. Intestinal coiling was apparent since 4 DAH, while mucosal folding began at 10 DAH. Scattered mucous cells occurred in the oral cavity and the intestine, while they were largely diffused in the oesophagus. Gastric glands and pyloric caeca were firstly observed at 28 DAH and appeared well developed by 41 DAH, indicating the transition from larval to juvenile stage and the acquisition of an adult mode of digestion.

Gonadal development, maturation, spawning and hermaphroditism were investigated in captive blackspot sea bream, Pagellus bogaraveo Brünnich, 1768, during the second, third and fourth years of life. The gonads of 224 fish were examined macroscopically and histologically. Undifferentiated gonads were found in fish smaller than 22.0 cm. Adult fish showed four gonadal differentiations: ovotestes with functional testis and quiescent ovary (Mf), ovotestes with functional ovary and regressed testis (mF), ovotestes with both ovary and testis at a resting stage (mf), and ovaries with no male tissue (F). The overall incidence of gonochoric females F was 41%. Functional males Mf were more frequent in age classes 1+ and 2+, whereas functional females mF predominated in the 3+ age class, above 25.0 cm TL. Histological examination revealed testicular degeneration and atrophy in functional females mF. On the basis of both histological data as well as size and age frequency distribution, it is suggested that the reproductive strategy of P. bogaraveo in captivity is characterized by protandrous hermaphroditism, with a high incidence of female gonochorism. Spawning occurred in March-April, at a size of 28.0 cm and age 3 in males and at 29.5 cm and age 4 in females. The gonosomatic index (GSI) remained constantly low (

BACKGROUND: The peptide hormone cholecystokinin (CCK), secreted by the midgut, plays a key role in digestive physiology of vertebrates including teleosts, by stimulating pancreatic secretion, gut motility, and gallbladder contraction, as well as by delaying gastric emptying. Moreover, CCK is involved in the regulation of food intake and satiation. Secretion of CCK by the hindgut is controversial, and its biological activity remains to be elucidated. The present paper addresses the regional distribution of intestinal CCK in the white sea bream, Diplodus sargus, as well as the possible involvement of hindgut CCK in digestive processes. METHODOLOGY/PRINCIPAL FINDINGS: Full-lengths mRNAs encoding two CCK isoforms (CCK-1 and CCK-2) were sequenced and phylogenetically analyzed. CCK gene and protein expression levels in the different gut segments were measured 3 h and 72 h after feeding, by quantitative real-time RT-PCR and Western blot, respectively. Moreover, endocrine CCK cells were immunoistochemically detected. Fasting induced a significant decrease in CCK-2 in all intestinal segments, including the hindgut. On the other hand, no significant difference was induced by fasting on hindgut CCK-1. CONCLUSIONS/SIGNIFICANCE: The results demonstrated two CCK isoforms in the hindgut of D.sargus, one of which (CCK-2) may be involved in the feedback control of uncompleted digestive processes. On the other hand, a functional role alternative to regulation of digestive processes may be inferred for D.sargus CCK-1, since its expression was unaffected by feeding or fasting.

The blackspot seabream Pagellus bogaraveo(Brunnich, 1768) has been regarded as a possible alternative to traditionally cultured Mediterranean species such as seabream and seabass, due to its high market value and good adaptation to captivity. Broodstock establishment and management represent the first step towards reliable production of eggs and fry, which is required to develop aquaculture of this new species. Two different broodstocks were tested for gonadal maturation and spawning, one constituting of wild fish caught as juveniles and reared in tanks until sexual maturity (4 years), and one assembled from wild adult fish caught during or just before the reproductive season. All fish were maintained under the same rearing conditions and fed the same diet. Gonadal stripping and biopsies were performed weekly to monitor maturation in both males and females. Ovarian samples were staged for maturity on the basis of follicular diameter and migration of germinal vesicle. Sperm samples were tested for density (number of spermatozoa ml−1) and motility. The fish reared in captivity reached ovarian maturity during the breeding season of the wild stock. Eggs were obtained by stripping from both farmed and wild specimens, but appeared degenerated as a result of being retained too long in the ovarian cavity due to the absence of spontaneous spawning. Spermiation was prolonged in the farmed fish, but appeared to be blocked in the wild breeders after first sampling. However, the sperm was very viscous and the motile spermatozoa did not exceed 10%.