Aspects of reproductive biology of the Mediterranean amberjack (Seriola dumerilii Risso) during the spawning period

Wild Seriola dumerilii were collected in the South Mediterranean Sea during the 1990, 1991 and 1992 spawning seasons. Macroscopic and cytological characteristics of ovary and testis were analysed. Depending on the presence and the number of oocytes at different stages, a five point maturity scale was proposed for ovarian maturity. Nine maturity stages of the oocytes are described. Oocyte size-frequency distribution has shown a group synchronous ovarian development type. The testes have a typical lobular-type structure. Depending on lobules and sperm duct development and on the abundance of germ cells, testis maturity was classified in four stages. Maturity rate according to age and size was also determined. 100% of 1-year-old fish show immature gonads. The proportion of females with mature ovaries was 0, 12.5, 84.6 and 100% at the age of 2, 3, 4 and 5 years respectively. The proportion of males with mature testes was 14.3, 40, 80 and 100 at the same age. The median standard length at which 50% of the fish attained maturity is 109 and 113 cm SL in males and females respectively.     

Circadian self-feeding rhythms in greater amberjack Seriola dumerili (Risso)

In the present study, 10 greater amberjack Seriola dumerili held individually were given free access to self-feeder, and trigger actuations were continuously monitored. Most (80%) of greater amberjack developed stable self-feeding activity within 1 week. The fish was a rigidly diurnal feeder under a 12L:12D cycle, with a feeding peak of 1–3 h occurring just after the onset of lights. Under constant conditions, all 10 individual fish exhibited free-running self-feeding rhythms, which persisted for 43 days without a sign of damping out. As far as is known, this seems to be the longest record of free-running behavioural rhythms in fishes. Except one fish free-ran at a period length (τ) of 25·2 h, τ of free-running self-feeding rhythms in the other nine fish was shorter than 24 h, ranging from 20·0 h to 21·5 h. In addition, phase transients of feeding activity were observed in response to the advancement of lights on. It is concluded that feeding activity in greater amberjack is mediated by endogenous circadian oscillators.     

Numerical taxonomy of Vibrionaceae isolated from cultured amberjack (Seriola dumerili) and surrounding water

A numerical taxonomic study was performed on 148 isolates of Gram-negative, heterotrophic, facultative anaerobic bacteria isolated from amberjack (Seriola dumerili) and its surrounding culture water. The study included 30 type and reference strains belonging to genera Vibrio, Listonella, and Photobacterium. The strains were characterized by 109 morphological, biochemical, physiological, and nutritional tests. Cluster analysis of similarity matrices obtained with S(SM) and S(J) coefficients was carried out. UPGMA (unweighted pair group mathematical average) analysis defined 11 phena at S(SM) values > or = 86%. Nine phena were identified as Vibrio alginolyticus, V. fischeri, V. harveyi, V. carchariae, V. mediterranei, V. splendidus, V. furnissii, V. parahaemolyticus, and Photobacterium damselae subsp. damselae. The two latter comprised strains isolated from diseased fish.     

Isolation and characterization of microsatellite loci from yellowtail Seriola dumerilii (Perciformes: Carangidae)

The Mediterranean yellowtail Seriola dumerilii (Risso, 1810) is a semi‐pelagic fish economically important to fisheries and aquaculture. We developed nine microsatellite primer pairs for this species. Microsatellites were isolated from a partial genomic library enriched for an AC motif. The developed loci proved to be polymorphic showing three to 11 alleles per locus. They can be employed for population studies as well as for parentage assessment. Cross‐species amplification was successfully tested in Seriola fasciata and Seriola rivoliana.     

First report on antibody response of Seriola dumerilii (Risso 1810) challenged with Listonella anguillarum

The greater amberjack, Seriola dumerilii (Risso 1810) is a semi-pelagic fish and a worldwide species; it is considered a promising candidate for the diversification of Mediterranean aquaculture. In this paper an experimental injection with Listonella (Vibrio) anguillarum was performed to study the immune response of S. dumerilii. Antibody titres to L. anguillarum O1 were determined with indirect-ELISA at different times over a period of 42 days. Results showed that the antibody levels against L. anguillarum were significantly higher in the challenged fish compared to the control. They started developing since the 5th day reaching the highest peak on day 20 after injection, indicating a fast response of the immune system. The observed antibody titre was very high versus L. anguillarum if compared to other fish species.     

The evolution of sperm ultrastructure among Boidae (Serpentes)

We investigate the evolution of sperm ultrastructure of three species of Boidae (Epicrates cenchria, Boa constrictor amarali, and Corallus hortulanus). Spermatozoa of these species are filiform consisting of a head region, containing the nucleus and acrosome complex, a midpiece, and a tail region subdivided into principal piece and endpiece. Multilaminar membranes and extracellular microtubules were observed next to the plasma membrane of the spermatozoa. The following differences were observed among the species: ridge on acrosome surface in Boa constrictor amarali (absent in Epicrates cenchria and Corallus hortulanus), stopper-like perforatorium base plate in Boa constrictor amarali and Epicrates cenchria (absent in Corallus hortulanus), rounded mitochondria in transverse section in Epicrates cenchria and Corallus hortulanus (irregular in Boa constrictor amarali). We mapped sperm characters onto two phylogenies based on morphological (Kluge in Misc Publ Mus Zool Univ Michigan 178:1–58, 1991) and molecular (Austin in Copeia 2:341–352, 2000) data, using a number of squamate species as outgroups. We identified 31 unambiguous character transformations in the morphological phylogeny and 30 in the molecular phylogeny, but only 13 and 12 transformations, respectively, are possible synapomorphies. We identified novel sperm synapomorphies, which were common between the morphological and molecular phylogenies: absence of perforatorium base plate and mitochondria arranged as sinuous tubes in oblique section (Serpentes), acrosome vesicle not subdivided and fibers 3 and 8 at the anteriormost region of principal piece (Boidae), and absence of an electron dense structure inside the proximal centriole (Elapidae + Colubridae). Our results suggest greater agreement between sperm ultrastructure and gross anatomical characters. In addition, we found no tendency for more homoplasies in the sperm head than in the flagellum, as recorded in previous studies.     

Growth rate of tank-reared Mediterranean amberjack, Seriola dumerili (Risso 1810) fed on three different diets

Mediterranean amberjack Seriola dumerili (Risso 1810) has aquaculture potential. The growth rate and food conversion ratio of S. dumerili on three different types of food (A: 100% frozen sardines; B: 50% frozen sardines and 50% pellet; C: 100% pellet) were measured, and analysed with respect to temperature, condition index and chemical composition of the fish fillet. Wild S. dumerili, average body weight 248 g and average total length 26.9 cm were caught in August and September 1994 in the South Adriatic Sea near Dubrovnik, Croatia and kept in three tanks (n=15 in each tank; duration of experiment, 226 days). The fish that were fed on diet A [initial weight, 252 ± 71 g; total length (TL), 24.3 ± 2.6 cm] started feeding immediately; however, fish assigned to diet C began to feed entirely on the pelleted diet 1 month after the start of the experiment. The mortality of fish fed on diet A was negligible, the registered growth rate was 313 ± 74 g (124.2%), specific growth rate was 0.32% day^?1 and the food conversion rate was 6.7. The fish fed on diet B (initial weight, 246 ± 74 g; TL, 28.2 ± 2.5 cm) started to feed on day 3 and achieved a total growth rate of about 98% (final weight gain, 241 ± 69 g) and specific growth rate of 0.24% day^?1 (feed conversion rate of 9.00 and mortality 13%). The fish fed on diet C (initial weight, 246 ± 74 g; TL, 28.2 ± 2.5 cm) started to feed on the pellets after 1 month and had a growth rate of 87% (weight gain 214 ± 85 g), a specific growth rate of 0.24% day^?1 and a food conversion rate of 10.6 with considerable mortality (27%). In all three diet groups the fish grew with considerable variation in food consumption and growth rate, depending on seasonal temperature variations of the ambient sea water supplied to the rearing tanks. Chemical analysis showed that the protein level (amount) in the fish meat exceeded 20% in all three fish fillet samples.     

Girdles as the main infection site for Paradeontacylix kampachi (Sanguinicolidae) in the greater amberjack Seriola dumerili

In this study, we provide new information about the habitats selected by the blood fluke Paradeontacylix kampachi in the greater amberjack Seriola dumerili based on an exhaustive anatomical examination. From May to October 1998, 21 fish of the 0+ age class were collected from tanks of the Spanish Institute of Oceanography in Puerto de Mazarrón, Spain, for parasitological analysis. Individuals of P. kampachi were found in 17 of the 21 fish analysed (mean intensity +/- SD: 13.6 +/- 16.6; median: 6). Worms occurred in the girdles, cephalic kidney, sinus venosus, kidney and branchial arteries. A Friedman test with a post-hoc contrast revealed a significantly higher number of worms in the girdles when compared with the other sites, suggesting this may be the main habitat for P. kampachi. This location had never been reported as a habitat for any species of Paradeontacylix, probably because it had not been examined before. Girdles should be routinely examined to estimate the actual intensity of infection and to maximize the likelihood of finding this species of digenean.     

Using molecular markers for pedigree reconstruction of the greater amberjack (Seriola dumerili) in the absence of parental information

Ensuring appropriate levels of genetic diversity in captive populations is essential to avoid inbreeding and loss of rare alleles by genetic drift. Pedigree reconstruction and parentage analysis in the absence of parental genotypes can be a challenging task that relies in the assignment of sibship relationships among the offspring. Here, we used eight highly variable microsatellite markers and three different assignment methods to reconstruct the most likely genotypes of a parental group of wild Seriola dumerili fish based on the genotypes of six cohorts of their offspring, to assess their relative contributions to the offspring. We found that a combination of the four most variable microsatellites was enough to identify the number of parents and their contribution to the offspring, suggesting that the variability of the markers can be more critical than the number of markers. Estimated effective population sizes were lower than the number of breeders and variable among years. The results suggest unequal parental contribution that should be accounted for breeding programs in the future.     

Age, growth and mortality of the Mediterranean amberjack Seriola dumerili (Risso 1810) from the south-eastern Adriatic Sea

The age, growth and mortality of the Mediterranean amberjack Seriola dumerili (Risso 1810) were determined in 298 specimens collected in the south-eastern Adriatic Sea (Donji Molunat Bay) from 17 May to 26 June 1997. The total length ranged from 32.0 to 160.0 cm and weight from 0.5 to 46.5 kg. Ten age classes, ranging from 1° to 10° years were defined by scale readings. Mean total length and weight-at-age data were used to estimate the growth parameters of the von Bertalanffy equation: L _∞=174.6 cm, K =0.190, t ₀=–0.314; W _∞=79.01 kg, K =0.139 and t ₀=–0.746. The length–weight relationship was estimated at: W =0.000123 ×  L ^2.847. The overall sex ratio was 1 : 1.06 in favour of males. Total ( Z ) and natural ( M ) mortality were found to be 0.41 years⁻¹ and 0.30 years^–1, respectively. The exploitation ratio ( E =0.27) indicates that the fishing pressure on the Mediterranean amberjack was low in the investigated region.     

PCR primers for trinucleotide and tetranucleotide microsatellites in greater amberjack,Seriola dumerili

Eighteen nuclear‐encoded microsatellites from a genomic DNA library of greater amberjack, Seriola dumerili, were isolated and characterized. The microsatellites include 13 perfect (five tetranucleotide and eight trinucleotide) and five imperfect (three tetranucleotide, one trinucleotide and one combination dinucleotide/trinucleotide) repeat motifs. The number of alleles at the 18 microsatellites among a sample of 29 fish ranged from two to 20; gene diversity (expected heterozygosity) ranged from 0.068 to 0.950, whereas observed heterozygosity ranged from 0.069 to 0.966. Following Bonferroni correction, genotypes at all 18 microsatellites fit expectations of Hardy–Weinberg equilibrium, and all pairwise comparisons of microsatellites did not deviate significantly from genotypic equilibrium. Greater amberjack support commercial and recreational fisheries along both the Atlantic and the Gulf coasts of the USA and represent a species with potential for worldwide aquaculture. The microsatellites developed will be useful for population genetic studies of ‘wild’ populations and breeding studies of domesticated populations.     

The sperm ultrastructure of Merluccius merluccius (Teleostei,Gadiformes): phylogenetic considerations

The anacrosomal aquasperm of the gadiform Merluccius merluccius is ultrastructurally similar to the advanced type II spermatozoa (perciform‐type sperm) typically found in most Perciformes. The perciform‐type spermatozoon is characterized by the lateral insertion of the flagellum and the location of the centrioles outside the nuclear fossa. Apart from these characteristics, the spermatozoon of M. merluccius is remarkable because of the mutually parallel arrangement of the centrioles, a rare feature among fishes, which is considered an apomorphic condition for animal sperm cells. Within the superorder Paracanthopterygii, which contains a large diversity of sperm patterns resulting from a high number of apomorphies, a perciform‐type sperm is present only in the order Gadiformes. The significance of the presence of perciform‐type spermatozoa in the three investigated gadiform families is discussed in a phylogenetic context.     

Complete genome sequence and characterization of virulence genes in Lancefield group C Streptococcus dysgalactiae isolated from farmed amberjack (Seriola dumerili)

Lancefield group C Streptococcus dysgalactiae causes infections in farmed fish. Here, the genome of S. dysgalactiae strain kdys0611, isolated from farmed amberjack (Seriola dumerili) was sequenced. The complete genome sequence of kdys0611 consists of a single chromosome and five plasmids. The chromosome is 2,142,780 bp long and has a GC content of 40%. It possesses 2061 coding sequences and 67 tRNA and 6 rRNA operons. One clustered regularly interspaced short palindromic repeat, 125 insertion sequences, and four predicted prophage elements were identified. Phylogenetic analysis based on 126 core genes suggested that the kdys0611 strain is more closely related to S. dysgalactiae subsp. dysgalactiae than to S. dysgalactiae subsp. equisimilis. The genome of kdys0611 harbors 87 genes with sequence similarity to putative virulence‐associated genes identified in other bacteria, of which 57 exhibit amino acid identity (>52%) to genes of the S. dysgalactiae subsp. equisimilis GGS124 human clinical isolate. Four putative virulence genes, emm5 (FGCSD_0256), spg_2 (FGCSD_1961), skc (FGCSD_1012), and cna (FGCSD_0159), in kdys0611 did not show significant homology with any deposited S. dysgalactiae genes. The chromosomal sequence of kdys0611 has been deposited in GenBank under Accession No. AP018726. This is the first report of the complete genome sequence of S. dysgalactiae isolated from fish.     

Spermatogenesis and sperm ultrastructure in the polychaete genusOphryotrocha (Dorvilleidae)

The details of spermatogenesis and spermiogenesis are described forOphryotrocha puerilis. The ultrastructure of mature sperm is shown forO. puerilis, O. hartmanni, O. gracilis, O. diadema, O. labronica, andO. notoglandulata. Clusters of sixteen cells each are proliferated by two stem cells in each setigerous segment ofO. puerilis representing the very early stages of both oogenesis and spermatogenesis. In each spermatocyte-I cluster, the cells are interconnected by cytoplasmic bridges. Early, clusters are enveloped by peritoneal sheath cells. These transient gonad walls break down prior to meiosis. The meiotic processes may start in the clusters with the cells still interconnected, or during breakdown of the original cluster, giving rise to smaller subclusters of both spermatocytes I and spermatocytes II with various numbers of cells. Finally, spermatid tetrads are present. As spermiogenesis progresses, the tetrads disintegrate. Golgi vesicles in both spermatocytes and spermatids contain electron-dense material, presumably preacrosomal. The acrosome is formed by such vesicles. In the six species studied here, the acrosomes appear to be of a similar overall structure but are of different shape. Centrioles are usually located beneath the acrosome. The distal centriole forms the basal body of a flagellum-like cytoplasmic process. The microtubules of these flagellar equivalents do not show a normal ciliar arrangement. The flagellar equivalent appears to be non-motile. InO. hartmanni and inO. notoglandulata, a flagellar equivalent is missing. Microtubules originating from the proximal end of the distal centriole stretch to the nuclear envelope. This feature appears to be especially conspicuous inO. puerilis and inO. labronica. InO. labronica and inO. notoglandulata, bundles of microtubules paralleling the cell perimeter appear to stabilise the sperm. Various numbers of mitochondria are either randomly distributed around the nucleus or accumulate on one side, often directly under the acrosome. Parts of the present paper were presented at the 2^nd International Polychaete Conference, Copenhagen 1986 and at the 3^rd International Polychaete Conference, Long Beach, Ca. 1989.     

Basal chromodorid sperm ultrastructure (Nudibranchia, Gastropoda, Mollusca)

The relationship between three genera considered basal in the Chromodorididae (Cadlina, Tyrinna, Cadlinella) has not yet been resolved by traditional morphological means. Here we examined the sperm ultrastructure of Tyrinna nobilis, Tyrinna evelinae, Cadlina flavomaculata and Cadlina cf. nigrobranchiata, with the expectation of finding phylogenetically informative characters. No Tyrinna or Cadlina species showed sperm similarities to Cadlinella. Both Cadlina species and Tyrinna nobilis (but not T. evelinae) exhibited coarse striations in the acrosomal pedestal. The putative fibers that occurred between the coarse striations of the pedestal are condensed into a layer in Cadlina and Tyrinna, but not in other species that also have coarse striations (Gymnodoris), and may constitute evidence for a close relationship. Tyrinna evelinae possessed fine acrosomal striations, which was shared with other Chromodorididae, Actinocyclidae and the cryptobranchs Rostanga and Aphelodoris. We also examined the sperm ultrastructure of ‘Chromodoris’ ambiguus, an animal which has shown molecular affinities to species of Cadlina, and not Chromodoris. The sperm of ‘C.’ ambiguus did not exhibit the typical Cadlina characteristics, but also showed important differences to other investigated Chromodoris species.     

Drug resistance and pulsed-field gel electrophoresis patterns of Lactococcus garvieae isolates from cultured Seriola (yellowtail, amberjack and kingfish) in Japan

AIMS: To investigate the existing antimicrobial susceptibility and genetic characteristics of Lactococcus garvieae isolates from cultured Seriola in Japan. METHODS AND RESULTS: Minimum inhibitory concentrations (MICs) of 14 antimicrobial agents for 170 isolates were determined using the agar dilution method. Seventy-five isolates (44.1%) were simultaneously resistant to erythromycin (EM) (MIC>or=2 microg ml-1), lincomycin (LCM) (MIC>or=128 microg ml-1) and oxytetracycline (OTC) (MIC>or=4 microg ml-1). Resistance to EM was grouped as intermediate- and high-level resistant by MIC values. All resistant isolates possessed ermB and tet(S) genes. The number of different bands between pulsed-field gel electrophoresis patterns of 25 isolates and two ATCC strains (isolated in 1974), determined using two enzymes (ApaI and SmaI), did not exceed 3. CONCLUSIONS: The present resistance pattern observed with ermB and tet(S) is similar to that observed in previous reports. Moreover, the genetic characteristics of L. garvieae isolates from a wide area in Japan in 2002 and ATCC strains were closely related. SIGNIFICANCE AND IMPACT OF THE STUDY: This study suggests that EM-, LCM- and OTC-resistant isolates have been present for 15 years and that L. garvieae strains with same origin have spread among Seriola spp. in Japan since 1974.     

Vaccine efficacy of Mycobacterium bovis BCG against Mycobacterium sp. infection in amberjack Seriola dumerili

Mycobacteriosis, caused by the intracellular parasitism Mycobacterium sp., causes economic damages to aquaculture production in Japan, particularly in seriola fish production. Antibiotics are not effective against Mycobacterium sp. and so a potent vaccine is needed. We previously reported that BCG vaccine (Mycobacterium bovis BCG) induces adaptive immunity against Mycobacterium sp. in Japanese flounder, Paralichthys olivaceus. In a phylogenetic tree, the genes for a major antigen, the Ag85 complex, in Mycobacterium sp. TUMSAT-Msp001 are closely related to homologues in Mycobacterium ulcerans. M. bovis BCG was detected until 7 days post-injection at the injection site (muscle) and 28 days post-vaccination in spleen. Cumulative mortality of amberjack, Seriola dumerili vaccinated intramuscularly (i.m.) and intraperitoneally (i.p.) with M. bovis BCG was 32.3% and 59.5% respectively, at 24 days post-infection of Mycobacterium sp., compared to 97.8% in PBS-injected fish. The bacterial counts of Mycobacterium sp. in spleen of both i.m.-and i.p.-vaccinated fish (6.2 × 10³ and 1.3 × 10⁴ CFU/mg tissue, respectively) at 20 days post-infection were significantly lower (P < 0.01) than those of PBS-injected fish (8.0 × 10⁶ CFU/mg). Furthermore, Immersion challenge with Mycobacterium sp. TUMSAT Msp-001 showed 50% RPS value in BCG i.m.-vaccinated fish at the end of the experiment. These results support our previous study using Japanese flounder and suggest that BCG vaccine is also effective against Mycobacterium sp. infection in amberjack.     

The digestive tract of the amberjack Seriola dumerili, Risso: a light and scanning electron microscope study

The histology of the digestive tract of the amberjack ( Seriola dumerili , Risso) was studied using light and scanning electron microscopy. The anterior oesophagus mucosa displays primary and secondary folds lined with a stratified squamous epithelium with fingerprint-like microridges which is substituted, on the top of the oesogaster folds, by a simple columnar epithelium with short microvilli. Only primary folds are present in the stomach. The anterior portion is rich in simple tubular glands, whereas the oesogaster and the pyloric region are devoid of them. Pyloric caeca and anterior and middle intestine mucosa display the same pattern of folding. The dominant cell type is the enterocyte, which exhibits larger and thinner microvilli in the caeca than in the intestine. The columnar epithelium of the rectum is replaced, in the anal sphincter, by a stratified flattened epithelium. Goblet cells are numerous throughout the whole length of the tract with the exception of the initial part of the oesophagus, the oesogaster, the stomach and the anal sphincter. Mucosubstances have been shown to vary in the different regions of the gut: acid mucines are found in the oesophagus, pyloric stomach, caeca, intestine and rectum, whereas neutral mucosubstances dominate in the anterior portion of the stomach. The muscularis is well developed throughout the length of the tract: two layers of striated muscle at the oesophageal level; two layers of smooth muscle in the stomach wall and three at the intestinal level.     

New findings on sperm ultrastructure of Xenos vesparum (Rossi) (Strepsiptera,Insecta)

The systematic position of insect order Strepsiptera is still under debate. It was, therefore, thought of interest to examine the ultrastructure of a strepsipteran in a search for synapomorphies shared with Coleoptera, Diptera, or any other insect order. The fine structure of spermatozoa and the spermatid from Xenos vesparum (Rossi) was re-examined using scanning and transmission electron microscopy and a fixation technique that permits the visualization of the macromolecular organization of the organelles. The spermatozoon was shown to possess several traits that are characteristics of insects in general, such as a 9 + 9 + 2 axoneme, two mitochondrial derivatives containing a crystalline material and two 'zipper lines' present along the sperm tail. Seventeen protofilaments occurred along most of the accessory tubules, which reduced to 16 posteriorly. An acrosome is absent. The neck region contains a prominent centriolar adjunct, which gives rise to two accessory bodies which adhere to the mitochondrial derivatives, and to slender strands of the so-called intertubular material found between the accessory tubules. Of interest is the finding that the glycocalyx consists of prominent filamentous strands, similar to those found in siphonapterans, mecopterans and basal dipterans.