The histopathology of fin rot disease in winter flounder from the New York Bight.

The histopathology of fin rot disease in winter flounder, Pseudopleuronectes americanus, from the New York Bight is described. Fin rot in winter flounder was characterized by progressive loss of portions of the anal and dorsal fins. Microscopic findings included epidermal hyperplasia accompanied by dermal fibrosis, hyperemia, and hemorrhage. Bacteria were not observed in situ and a pronounced leukocytic inflammatory response was not noted.     

Differential amplification of antifreeze protein genes in the pleuronectinae

Summary The organization of antifreeze protein (AFP) genes in the yellowtail flounder was investigated by Southern blotting and the characterization of clones from a genomic library. This flounder, like the closely related winter flounder, has a set of 10–12 linked but irregularly spaced AFP genes. However, it lacks the tandemly amplified set of 20 such genes that are present in the winter flounder. DNA sequence analysis of a tandemly repeated gene from winter flounder showed that it can code for one of the two most abundant AFP components in the serum. Consistent with this higher AFP gene dosage, the peak serum AFP level in midwinter was 9 mg/ml in the winter flounder and only 4 mg/ml in the yellowtail flounder. A recent amplification of the AFP gene in the winter flounder lineage might be responsible for the higher serum AFP levels in this fish. This increase in gene dosage might have helped the winter flounder colonize the ice-laden, shallow-water niche that it currently occupies along the east coast of North America. Genomic Southern blotting of two other righteye flounders, the smooth flounder and the American plaice, illustrates another example of a differential amplification of AFP genes that correlates with a species' exposure to ice.     

Factors affecting the post‐release survival of cultured juvenile Pseudopleuronectes americanus

Laboratory experiments were performed with cultured and wild juvenile winter flounder Pseudopleuronectes americanus to evaluate differences in behaviour and adaptation affecting post‐release vulnerability to predation. Studies revealed that the cryptic abilities of cultured winter flounder increased over time. Sediment‐naïve, cultured fish required a minimum of 2 days to improve their burying skills and at least 90 days for colour adaptation to match the sediment. Cultured winter flounder selected sediments consisting of small grains and colours matching their own pigment. Cultured winter flounder, regardless of their colour, were significantly more vulnerable to predation by birds. Additionally, cultured fish reacted differently than wild winter flounder when exposed to cues from a potential predator.     

The genetic stock structure of larval and juvenile winter flounder larvae in Connecticut waters of eastern Long Island Sound and estimations of larval entrainment

Variability at six microsatellite loci was examined among 536 winter flounder Pseudopleuronectes americanus larvae collected from three locations in eastern Long Island Sound shown to be nursery areas for the species. Substantial genetic differences were seen among the putative source populations and thus appeared to be geographically based. These differences were used to characterize the most likely sources of winter flounder larvae entrained at the Millstone Power Station as well as recruitment to juvenile winter flounder collected in the Niantic River. Samples were classified to the most likely geographical source population both by a conditional maximum likelihood method and by a multi‐layer feed‐forward neural net trained on the differences in microsatellite allele frequencies. The classification of samples by both methods is compared and discussed in the context of winter flounder management.     

Predatory impact of the green crab (Carcinus maenas Linnaeus) on post-settlement winter flounder (Pseudopleuronectes americanus Walbaum) as revealed by immunological dietary analysis

Predation on flatfish during the early juvenile stage is an important factor regulating year-class strength and recruitment. In this study, immunological dietary analysis was performed on green crabs (Carcinus maenas) collected from the Niantic River, Connecticut, in an effort to evaluate the predatory impact of this species on post-settlement winter flounder (Pseudopleuronectes americanus). Through the use of species-specific antiserum, winter flounder proteins were identified in 4.8% of the green crab stomachs analyzed (n = 313, size range = 14-74 mm carapace width, CW), revealing that crabs ≥ 29 mm CW are predators of post-settlement winter flounder in natural populations. The most significant factor underlying the predator-prey interaction was the relative size relationship between species, such that the incidence of winter flounder remains in the stomach contents of green crabs was positively correlated with predator-to-prey size ratio. Results from dietary analysis were incorporated into a deterministic model to estimate the average daily instantaneous mortality and cumulative mortality of winter flounder owing to green crab predation. Accordingly, green crabs may account for 0.4% to 7.7% (mean = 2.2%) of the daily mortality of winter flounder and consume 1.1% to 32.3% (mean = 10.2%) of the flounder year-class. Model simulations further indicate that variations in green crab abundance and size-structure account for the greatest variability in winter flounder mortality. Relative to other macro-crustacean predators, however, predation by green crabs has a minimal effect on winter flounder survival, due in large part to the low densities of these crabs in temperate estuaries.     

Isolation, characterization and seasonal variations in the concentration of N-epsilon-(gamma-glutamyl)-lysine isodipeptide in the blood plasma of the winter flounder (Pseudopleuronectes americanus).

A ninhydrin-positive compound was isolated from the plasma of the winter flounder (Pseudopleuronectes americanus) and identified as the isodipeptide N-epsilon-(gamma-glutamyl)-lysine. Wide seasonal variations in plasma concentration of this compound are found in the male flounder, whereas it is present only in trace amounts in the plasma of the female flounder throughout the year. In the male flounder the plasma concentration of N-epsilon-(gamma-glutamyl)-lysine rises from January to a peak in May and June. It is during these latter months that the flounder spawn. After spawning, the plasma concentration of the isodipeptide decreases to low basal values in October. These changes in plasma concentration of N-epsilon-(gamma-glutamyl)-lysine may be related to changes in testes weight, spermatogenesis and spawning activity of the winter flounder.     

Feeding behavior of newly settled winter flounder (Pseudopleuronectes americanus) on calanoid copepods

Field and laboratory investigations were conducted to examine feeding by newly settled winter flounder (Pseudopleuronectes americanus) on two co-occurring calanoid copepods, Eurytemora affinis and Acartia hudsonica. During the spring, these prey are present when winter flounder initiate their demersal lifestyle in estuaries of the northeastern United States. Epibenthic zooplankton were collected concurrently with winter flounder in the Navesink River estuary, NJ, in May 1998 and 1999. Although both calanoid species were in the estuary during the 2-year survey, E. affinis was consumed nearly to the exclusion of A. hudsonica by newly settled winter flounder. Annually, E. affinis and A. hudsonica had similar size distributions in field collections, indicating that species choice was not size selective. However, when preying on E. affinis, winter flounder preferred the larger sized organisms. In single species laboratory experiments, E. affinis and A. hudsonica were consumed equally by newly settled winter flounder (19-23 mm TL), but there were more strikes made toward E. affinis. Despite the lower catch efficiency, E. affinis was selected over A. hudsonica when the prey species were offered together in equal numbers. The selection for E. affinis over A. hudsonica by newly settled winter flounder may be the result of behavioral and/or morphological differences in the prey species.     

Size-related shifts in the habitat associations of young-of-the-year winter flounder (Pseudopleuronectes americanus): field observations and laboratory experiments with sediments and prey

Field surveys and laboratory studies were used to determine the role of substrata in habitat selection by young-of-the year winter flounder. A synoptic field survey of winter flounder and sediments in the Navesink River-Sandy Hook Bay estuarine system in New Jersey demonstrated that winter flounder distribution was related to sediment grain size. Analysis using a generalized additive model indicated that the probability of capturing 10-49 mm SL winter flounder was high on sediments with a mean grain diameter of /=40 mm SL) preferred coarse-grained sediments. Burying ability increased with size and all flounders avoided sediments that prevented burial. Subsequent laboratory experiments revealed that the presence of live prey (Mya arenaria) can over-ride sediment choice by winter flounder (50-68 mm SL) indicating the complexity of interrelated factors in habitat choice.     

A natural variant of type I antifreeze protein with four ice-binding repeats is a particularly potent antifreeze.

A 4.3-kDa variant of Type I antifreeze protein (AFP9) was purified from winter flounder serum by size exclusion chromatography and reversed-phase HPLC. By the criteria of mass, amino acid composition, and N-terminal sequences of tryptic peptides, this variant is the posttranslationally modified product of the previously characterized AFP gene 21a. It has 52 amino acids and contains four 11-amino acid repeats, one more than the major serum AFP components. The larger protein is completely alpha-helical at 0 degree C, with a melting temperature of 18 degrees C. It is considerably more active as an antifreeze than the three-repeat winter flounder AFP and the four-repeat yellowtail flounder AFP, both on a molar and a mg/mL basis. Several structural features of the four-repeat winter flounder AFP, including its larger size, additional ice-binding residues, and differences in ice-binding motifs might contribute to its greater activity. Its abundance in flounder serum, together with its potency as an antifreeze, suggest that AFP9 makes a significant contribution to the overall freezing point depression of the host.     

Predator-prey relations between age-1+ summer flounder (Paralichthys dentatus, Linnaeus) and age-0 winter flounder (Pseudopleuronectes americanus, Walbaum): predator diets, prey selection, and effects of sediments and macrophytes

Laboratory experiments and weekly trammel net surveys in the Navesink River, New Jersey (USA) were used to examine the predator-prey interaction between age-1+ summer flounder (Paralichthys dentatus) and age-0 winter flounder (Pseudopleuronectes americanus). Winter flounder (24-67 mm TL) were the dominant piscine prey of summer flounder (n=95, 252-648 mm TL) collected in trammel nets. We observed a temporal shift in summer flounder diets from sand shrimp (Crangon septemspinosa) and winter flounder, dominant during June and early July, to blue crabs (Callinectes sapidus) and other fishes (primarily Atlantic silversides, Menidia menidia and Atlantic menhaden, Brevortia tyrannus) later in the summer. Variations in prey selection appeared to be related to changes in the spatial distribution of predators and spatio-temporal variation in prey availability. In laboratory experiments, summer flounder (271-345 mm total length, TL) preferred demersal winter flounder to a pelagic fish (Atlantic silversides) and a benthic invertebrate (sand shrimp) prey, and the vulnerability of winter flounder increased with increasing prey body size from 20 to 90 mm TL. Experiments testing habitat effects showed that mortality of winter flounder in three different size classes (20-29, 40-49, 60-69 mm TL) was not influenced by sediment grain sizes permitting differential burial of the prey. However, vegetation enhanced survival, with fish suffering lower mortality in eelgrass (Zostera marina, 15+/-0.04%) than in sea lettuce (Ulva lactuca, 38+/-0.04%) or bare sand (70+/-0.07%) when the macrophytes were planted to produce similar leaf surface areas (5000 cm(2) m(-2)). Prey vulnerability appeared to be related to the role of vision in the predator's attack strategy and prey activity levels.     

Effect of sediment,salinity, and velocity on the behavior of juvenile winter flounder (<Emphasis Type="Italic">Pseudopleuronectes americanus</Emphasis>)

Winter flounder (Pseudopleuronectes americanus) is a benthic flatfish that is economically important for recreational and commercial fishing in North America. In the last twenty years, the species has undergone a drastic decline, mainly due to anthropic influence. The goal of this study was to gain knowledge on habitat preferences and behavior of juvenile winter flounder to improve the management of natural stocks and optimize release sites of juveniles produced for stock enhancement. Three abiotic factors (sediment, current, and salinity) potentially influencing the distribution of flatfish species were tested in a recircurlating flume with juvenile winter flounder. Time budgets of observed behaviors including swimming, orientation, and burying capacity were analyzed. Sediment texture was the only factor that significantly influenced the burying behavior of winter flounder juveniles; shear velocity, salinity, and sediment had no effect on the orientation of juveniles.     

Comparison of routine oxygen consumption rates of three species of pleuronectids at three temperatures

The oxygen consumption rates of three species of pleuronectids, the yellowtail flounder, Pleuronectes ferrugineus (Storer), the winter flounder, Pleuronectes americanus (Walbaum), and the American plaice, Hippoglossoides platessoides (Fabricius), were examined under simulated, land-based, aquaculture conditions. Routine oxygen consumption (ROC) rates for groups of each species were measured simultaneously using single-pass, flow-through respirometry. This study was conducted over three seasons at temperatures from 2°C to 14°C. An analysis of variance identified a significant interaction between temperature and species on the oxygen consumption rates of these flounder. The analysis indicated that at each temperature, ROC rates were significantly different among the three species (P < 0.05). A subsequent test of each species'ROC rate across the three temperatures indicated that both the yellowtail flounder and the winter flounder had significantly different ROC rates at each temperature experiment (P < 0.05). The ROC of yellowtail and winter flounder responded similarly to changes in experimental temperatures.     

Distribution and ontogeny of digestive enzymes in larval yellowtail and winter flounder

The distribution of digestive enzymes was studied using histoenzymological methods in yellowtail and winter flounder larvae reared on three different diets: live food, weaned at day 15, or starved. Alkaline phosphatase, dipeptidyl peptidase IV, aminopeptidase M and esterase were present at 3 days post-hatch and became differentially distributed coinciding with morphological development. For larvae fed a live diet, activity of these enzymes was present in the intestine of both species and rectum of yellowtail flounder. Alkaline phosphatase was also present in the post-oesophageal swelling (stomach anlage) of winter flounder. Activity of all enzymes was absent in starved winter flounder larvae and a decrease in aminopeptidase M and esterase activity occurred in starved yellowtail flounder larvae. Acid phosphatase was not identified in either species. The eVect of weaning on enzymatic activity was not evaluated fully as the larvae did not survive long enough after the introduction of the artificial diet to complete experimentation.     

A comparison of the effects of temperature on melanophore in vitro responses to noradrenalin in two teleost species adapted to cold ocean and tropical conditions

Melanosome-aggregating responses to noradrenalin in winter flounder, Pseudopleuronectes americanm Walbaum, manifest different in vitro temperature characteristics compared with such activity in sailfln molly, Poecilia velifera Regan. Winter flounder off St John's are adapted to cold ocean temperatures between − 1° C and + 15° C associated with the Labrador current, whilst sailfin molly is a tropical species adapted to temperatures up to about 30° C. The experimental data indicate that melanophores from these two species are physiologically adapted to function in contrasting thermal environments. Consequently, the optimum temperature for melanosome aggregation in winter flounder is approximately 20° C, and that of the tropical species approximately 40° C. Also, flounder melanophore In vitro responses are relatively uninfluenced by low temperatures, which drastically inhibit sailfin molly melanophore activity.     

Structural variations in the alanine-rich antifreeze proteins of the pleuronectinae

The sequence and activity of antifreeze proteins from two right eye flounder species were compared to assess the influence of structural variations on antifreeze capacity. The cDNA encoding the major serum antifreeze protein in the yellowtail flounder (Limanda ferruginea) was cloned from liver tissue. Its DNA sequence shows that the precursor to the antifreeze is a 97-residue preproportion. Edman degradation identified the N-terminus of the 48-amino-acid mature serum antifreeze protein and confirmed the sequence of the first 36 residues. A comparison with the previously determined winter flounder antifreeze protein and mRNA sequences shows strong homology through the 5' and 3' untranslated regions and in the peptide region. The mature protein section has the greatest sequence variation. Specifically, the yellowtail antifreeze protein, in contrast to that of the winter flounder, contains a fourth 11-amino-acid repeat and lacks several of the hydrophilic residues that have been postulated to aid in the binding of the protein to ice crystals. Intramolecular salt bridges are present in the antifreeze proteins from both species but in different registries with respect to the 11-amino-acid repeats. On a mass basis the yellowtail flounder antifreeze, though longer than that of the winter flounder, is only 80% as effective at depressing the freezing temperature of aqueous solutions. This lower activity might be due to the reduced number of hydrophilic ice-binding residues per molecule.     

A New PCR-Based Method Shows That Blue Crabs (Callinectes sapidus (Rathbun)) Consume Winter Flounder (Pseudopleuronectes americanus (Walbaum))

Winter flounder (Pseudopleuronectes americanus) once supported robust commercial and recreational fisheries in the New York (USA) region, but since the 1990s populations have been in decline. Available data show that settlement of young-of-the-year winter flounder has not declined as sharply as adult abundance, suggesting that juveniles are experiencing higher mortality following settlement. The recent increase of blue crab (Callinectes sapidus) abundance in the New York region raises the possibility that new sources of predation may be contributing to juvenile winter flounder mortality. To investigate this possibility we developed and validated a method to specifically detect winter flounder mitochondrial control region DNA sequences in the gut contents of blue crabs. A survey of 55 crabs collected from Shinnecock Bay (along the south shore of Long Island, New York) in July, August, and September of 2011 showed that 12 of 42 blue crabs (28.6%) from which PCR-amplifiable DNA was recovered had consumed winter flounder in the wild, empirically supporting the trophic link between these species that has been widely speculated to exist. This technique overcomes difficulties with visual identification of the often unrecognizable gut contents of decapod crustaceans, and modifications of this approach offer valuable tools to more broadly address their feeding habits on a wide variety of species.     

Annual variation in lymphocystis infection frequency in flounder, Platichthys flesus (L.)

The infection frequency of the virus induced lymphocystis disease has been registered in flounder from the Oslofjord, Norway, during a 4 year period. The disease has a clear regular seasonal variation, and also an annual variation. The percentage infection in the summer was 1–10%, and in the winter up to 57%. The possibility that the high pollution level of the inner Oslofjord could be the reason for the high frequency of lymphocystis disease, is discussed.     

Melanophore comparisons in different forms of ambicoloration in the flatfish Pseudopleuronectes americanus and Reinhardtius hippoglossoides

The melanophores associated with contrasting manifestations of ambicoloration in two species, winter flounder ( Pseudopleuronectes americanus ) and Greenland halibut ( Reinhardtius hippoglossoides ), from the Pleuronectidae are compared microscopically. Winter flounder is a benthic inshore species, whilst Greenland halibut is a deep water species considered to spend the greater part of its life away from the sea floor, although the relatively thick blind-side epidermis associated with a benthic life occurs in both species. Various forms of abnormal ambicoloration occur in some specimens of winter flounder. Melanophores from the flounder blind side are morphologically similar to those from the ocular side in the different forms of ambicoloration, although there are differences in melanophore densities. In Greenland halibut, ambicoloration is a normal characteristic which is considered to be secondarily acquired. The melanophores of the ocular side of Greenland halibut are similar to the melanophores of other Pleuronectidae, whereas those of the blind side are very different in morphology and size.     

Localization of vitellogenin-related protein in the developing oocytes of winter flounder (Pleuronectes americanus) by protein A-gold immunocytochemistry

Vitellogenin-related proteins were localized within the oocytes of a marine teleost, the winter flounder, using protein A-gold immunocytochemical electron microscopy. A homologous vitellogenin antisera was used along with 15-nm protein-A gold on thin sections of oocytes. Localization was significantly confined to organelles described as yolk spheres within the oocyte cytoplasm. This technique provides specific identification of yolk-containing organelles within the developing winter flounder oocyte.