Sustained exercise improves vertebral histomorphometry and modulates hormonal levels in rainbow trout

Abnormal compressions and fusions of vertebral bodies are frequently observed in reared rainbow trout and could result from chronic and unbearable muscle pressures acting on the axial skeleton during intensive growth. Sustained swimming at moderate speeds was shown to induce many positive effects on growth and swimming performances in salmonids, but yet little is known about its effects on vertebral remodeling processes and related hormonal regulation. Rainbow trout were subjected to three different swimming speeds (0, 1.0 and 1.5 body length (BL) s^− 1), starting one month after they were first fed (65 mm) and ending when they reached 260 mm in size (market-size of 275 g). At the end of the experiment, 20 trout were sampled in each lot (N = 60) and blood samples were taken. Vertebrae abnormalities were assessed by radiological examinations. Vertebrae from the middle axial region (V32–38) were selected to evaluate bone mineralization (BM) and total bone area (Tt-B.Ar.) on radiographed transverse sections (125 ± 10 μm). Assays were performed to evaluate mineral homeostasis (calcemia and phosphatemia), bone cell activities (alkaline phosphatase, ALP, and tartrate-resistant acid phosphatase, TRAP) and bone regulating hormones (calcitonin, CT and thyroid hormones, THs). Sustained exercise reduced the appearance of fused vertebrae, enhanced vertebral BM and decreased vertebral Tt-B.Ar., while it increased circulating CT and TH levels. No variations were observed on mineral homeostasis and bone cell activities. Increasing the swimming speed up to 1.5 BL s^− 1 had positive effects on the vertebral skeleton, and therefore, seems to be a suitable approach to prevent aggravation of vertebral abnormalities in juvenile trout. The changes observed in vertebral features are interpreted as a compromise between the necessity to mobilize vertebral mineral ions in response to various physiological demands and to maintain vertebral strength against mechanical constraints.     

Assessment of “discreet” vertebral abnormalities, bone mineralization and bone compactness in farmed rainbow trout

The present study aimed to evaluate the importance of “discreet” vertebral abnormalities in normally-shaped rainbow trout in relation to vertebral bone condition in French fish farms. A total of 373 trout (262 ± 2 mm in total length) from 20 fish farms sampled were studied. The fish were radiographed and the axial skeleton examined for vertebral abnormalities. Vertebrae from the middle axial region (V32-38) were selected to evaluate vertebral bone condition. Bone mineralization (BM, %) was estimated by the ratio of ash and dry weight. Bone compactness (BC,%) was measured using Bone Profiler 3.23 images software on digitized radiographs of transverse sections (125 ± 10 μm). Statistical analyses were performed to test the relationships between the occurrence of vertebral abnormalities, and BM and BC. The occurrence of affected trout ranged from 0 to 55% depending on the farm. Trout displayed vertebrae with low BM (< 54.6%) and low BC (< 28.1%) in 40% and 55% of the farm, respectively. No relationships were observed between bone condition parameters (BM, BC) and the occurrence of vertebral abnormalities. These results could be explained by a wide and variable plastic response of bone characters (i.e., vertebral abnormalities, BM and BC) to the various rearing conditions in the fish farms sampled.     

The mineral content affects vertebral morphology in underyearling smolt of Atlantic salmon (Salmo salar L.)

This study investigated the individual variation in vertebral morphology and mineral content (as % of bone dry weight) in rapid growing underyearling smolt (n = 33) twelve weeks after transfer to seawater. The smolt, which were reared in triplicate tanks, had a large individual variation in vertebral bone mineral content (Tank A: 36.2% min, 45.2% max, 39.6% mean. Tank B: 32.6% min, 46.6% max, 40.0% mean. Tank C: 38.5% min, 47.9% max, 42.4% mean), and were subdivided into two groups based on mean within each tank: high mineral content (HMC, above tank mean, n = 15) and low mineral content (LMC, below tank mean, n = 18). Vertebrae of smolt with a low mineral content were more compressed in the anterior–posterior direction (HMC vertebral length/dorso-ventral diameter ratio 0.83, LMC 0.80). This was related to a change in the growth direction of the vertebral cone. This change occurred around transfer to seawater and was characterised by an increased angle between the wall of the cone of the vertebrae and the cranial–caudal axis, and gave the vertebrae a compressed appearance. Smolt with a high mineral content were significantly longer (HMC 28.1 cm, LMC 26.5 cm) and heavier (HMC 270 g, LMC 210 g) twelve weeks after transfer to seawater.     

美洲鲥仔稚鱼脊柱及附肢骨骼系统的早期发育

使用软骨-硬骨双染色技术,描述了美洲鲥(Alosa sapidissima)仔稚鱼(1~51日龄)脊柱、胸鳍、尾鳍、背鳍等附肢骨骼的形态发育特征。结果显示,脊柱的发育开始于10日龄仔鱼尾部的髓弓、脉弓和尾下骨的出现,16日龄髓弓和脉弓延伸形成髓棘和脉棘,19日龄脊柱出现分节的硬骨环,23日龄所有椎体形成。各附鳍支鳍骨发育顺序先后依次为胸鳍、尾鳍、背鳍、臀鳍和腹鳍。胸鳍在2日龄时出现乌喙骨,13日龄形成软骨质的胸鳍支鳍骨,19日龄仔鱼肩带和上匙骨开始骨化;尾鳍的尾下骨最早出现在5日龄,12日龄尾鳍形成2枚尾上骨、1枚尾杆骨和6枚尾下骨,19日龄仔鱼尾椎和尾鳍率先开始骨化,直至23日龄尾鳍骨骼系统钙化完全。最终背鳍和臀鳍分别形成18和22根鳍条。美洲鲥骨骼发育研究对其早期发育功能趋向、环境优化及分类鉴定有重要作用。     

Increased dietary phosphorous prevents vertebral deformities in triploid Atlantic salmon (Salmo salar L.)

Triplicate groups of triploid and diploid Atlantic salmon were fed diets with a low (LP, total P: 7.1 g kg^?1), medium (MP, total P: 9.4 g kg^?1) or high (HP, total P: 16.3 g kg^?1) phosphorous (P) level from first feeding (0.18 g) to transfer to sea water (~50 g, duration: 203 days) and subsequently fed a commercial diet in sea water for 426 days (~3 kg). This study examined the short‐ and long‐term effects of dietary P on freshwater performance (mortality, growth), vertebral deformities (radiology), bone cell activity (ALP and TRACP enzyme activity in vertebrae and scales, and fgf23, bgp and igf‐I relative gene expression in vertebrae), bone mineralization (ash content) and some parameters related to fish condition (heart and liver size). Irrespective of ploidy, at seawater transfer, fish fed the MP diet had significantly highest length and weight and those fed the LP diet significantly lowest length and weight, while those fed the HP diet had intermediate lengths and weights. Increased dietary phosphorus reduced deformities in both ploidies at seawater transfer; however, triploids fed the LP and MP diets had more deformities than diploids fed the respective diets, while there was no ploidy effect observed for fish fed the HP diet. The vertebral bone ash content at seawater transfer was significantly higher in diploids than in triploids when fed the MP diet only. Alkaline phosphatase (ALP) and tartrate‐resistant acid phosphatase (TRACP) enzyme activities and relative gene expression of bone hormones involved in metabolism of plasma phosphate (fgf23) and bone growth (bgp) were not affected by ploidy at seawater transfer, but by dietary P level; LP increased ALP activity and reduced TRACP activity and fgf23 and bgp expression levels in vertebral bone. In scales, LP increased both ALP and TRACP activity. At the termination of the seawater period, the group‐wise pattern in occurrence of vertebral deformities was the same as at seawater transfer. The present results on mortality, growth, bone mineralization and development of skeletal deformities all demonstrate that triploids have a higher P requirement than diploids in fresh water. This study shows that an optimalization of P nutrition for triploid Atlantic salmon can improve health and welfare and reduce down‐grading of triploid salmon.     

The prevalence of vertebral deformities is increased with higher egg incubation temperatures and triploidy in Atlantic salmon Salmo salar L.

Suboptimal egg incubation temperature is a risk factor for the development of skeletal deformities in teleosts. Triplicate diploid and triploid Atlantic salmon, Salmo salar L., egg batches were incubated at 6, 8 and 10 °C up until first feeding, whereupon fish were reared on a natural temperature before examination for externally visible skeletal deformities (jaw and spine) and radiographed for vertebral deformities and morphology at the parr stage. Increasing incubation temperatures and triploidy increased the number of fish showing one or more deformed vertebrae. Triploids had significantly higher mean vertebrae cranio‐caudal length (L) and dorsal‐ventral height (H) ratio at 6 and 10 °C than diploids, but triploidy had no effect on mean vertebrae centra area. Triploids demonstrated an increase in lower jaw deformities with increased incubation temperature, whereas jaw deformities were rare in diploids. Fish incubated at 10 °C had a significantly lower mean vertebral number than fish incubated at 6 °C, and triploids had lower mean vertebral numbers than diploids. Diploid fish with 58 vertebrae had a significantly higher mean vertebral centra area than fish with 59 vertebrae, but vertebral number did not affect the mean vertebral L/H ratio. The results are discussed with respect to the welfare and production of farmed salmonids.     

Estimation of heritability and genetic correlation of number of abdominal and caudal vertebrae in masu salmon

ABSTRACT:   Ten sibship masu salmon Oncorhynchus masou masou families were produced to estimate the heritability of the number of abdominal vertebrae (AV) and caudal vertebrae (CV), and the genetic correlation between the two traits were calculated. The AV and CV of both the parents and their offspring were counted separately, and the heritability was estimated by performing parent–offspring regression analysis. The mean vertebral number of the offspring differed among families although they were reared under identical water temperature conditions and had the same parental vertebral number type in crossbreeding. The heritabilities were estimated as 0.65 for AV and 0.84 for CV. The genetic correlation between the number of AV and CV was calculated as −0.92. These results suggest that AV and CV are governed by genetic components even when the fish are reared under identical environmental conditions, and the total vertebral number is expected to converge within a certain range. It is necessary to analyze separately the variations in the number of AV and CV when investigating the vertebral number in masu salmon.     

Characterization of differentially expressed genes in liver in response to the rearing temperature of rainbow trout Oncorhynchus mykiss and their heritable differences

To characterize thermal-responsive genes in fish, firstly, juvenile rainbow trout were reared in four different temperature conditions (average temperatures were 10, 14, 18, and 22 °C, respectively) and differentially expressed genes were identified. Gene expression in the liver was analyzed by the differential display method, followed by validation using real-time PCR. Subsequently, to examine whether the identified genes show heritable differences, the gene expression levels were compared among juveniles of three genetically distinct lines of rainbow trout (a strain and two closed colonies) by rearing at two different temperature conditions (average 14 and 22 °C). By rearing at 22 °C, growth retardation was observed compared with fish reared at 14 and 18 °C, and six genes were identified as differentially expressed genes in response to the rearing temperature in the gene expression analyses. With the increase in rearing temperature, gene expressions of a complement C1q and two ribosomal proteins were significantly up-regulated. On the other hand, three metabolic genes (betaine homocysteine methyltransferase, triosephosphate isomerase, and glucose-6-phosphatase) were down-regulated, indicating a metabolic depression due to high temperature. In the subsequent analyses, in response to the rearing temperature (14 and 22 °C), there was a trend that the complement C1q and glucose-6-phosphatase genes showed different expression patterns among the three rainbow trout lines, suggesting heritable differences in these genes. Our study provides information on thermal-responsive genes in fish, and we anticipate it will facilitate further investigation in the thermal biology of fish.     

Effects of Temperature on Production and Specificity of Antibodies in Rainbow Trout (Oncorhynchus mykiss)

Abstract.— The effect of temperature on production and affinity of antibodies against antigens from the parasitic ciliate Ichthyophthirius multifiliis were studied in rainbow trout ( Oncorhynchus mykiss ). Fish were immunized with I. multifiliis antigens and reared at three different temperatures, 5, 12, and 20 C for 56 d. The production of specific antibodies was examined by enzyme-linked immunosorbent assay (ELISA). The results showed that temperature has a pronounced effect on the assay result of the ELISA technique. Plasma samples analyzed at 5 C showed that the specific antibody response of fish reared at 5 C was similar to fish reared at 12 and 20 C. However, when samples were assayed at 12 and 20 C, the measured antibody response tended to be higher for the samples from trout reared at 12 and 20 C. Additionally, it was found that rainbow trout reared at 5 C showed a delayed but not hampered antibody response compared to fish reared at 12 and 20 C, indicating a correlation between low temperature and delayed antibody production.     

Impact of smolt production strategy on vertebral growth and mineralisation during smoltification and the early seawater phase in Atlantic salmon (Salmo salar, L.)

This study investigates the effect of different smolt production strategies on vertebral morphology (radiology), composition (mineral content) and mechanical strength (load-deformation testing) in Atlantic salmon (Salmo salar). Rapid-growing underyearling (0+) smolt were compared with slower-growing yearling (1+) smolt and a reference group of wild smolt (w). The underyearling and yearling smolt were transferred to seawater in October 2002 and May 2003, respectively. The underyearling smolt were reared under continuous light and the yearling smolt under natural light during the first twelve weeks in seawater, at ambient temperatures. Thus, the underyearling smolt hit seawater at 13 °C and were reared at 10-13 °C during the early seawater phase, whereas the yearling smolt hit seawater at 7 °C and were reared at 7-10 °C during the early seawater phase. All groups displayed increased longitudinal growth (up to 9% increase in relative length) of the caudal vertebrae during parr-smolt transformation. However, at transfer to seawater, the underyearling smolt had significantly lower vertebral mineral content (0+ 44%, 1+ 47%, w 50%) and higher incidence of deformed vertebrae (0+ 1.5%, 1+ 0%, w 0%), and at twelve weeks after transfer to seawater significantly lower vertebral mineral content (0+ 36%, 1+ 41%, w 43%), yield-load (0+ 6492 g, 1+ 8797 g, w 9150 g) and stiffness (0+ 7578 g/mm, 1+ 15,161 g/mm, w 20,523 g/mm), and significantly higher incidence of deformed vertebrae (0+ 2.5%, 1+ 0.3%, w 0%). There was a significant correlation between the mineral content and mechanical properties of the vertebrae. The underyearling smolt had significantly elevated plasma concentrations of total Ca, and P and Ca²⁺ during the parr-smolt transformation and in the early seawater phase.The results show that underyearling smolt may have an increased risk of developing vertebral deformities. It is possible that this risk can be reduced by postponing the start of the short-day treatment. This will reduce the temperature during smoltification, the temperature and daylength during the early seawater phase, and increase the age at smoltification.     

Qualitative and quantitative changes of F<Subscript>o</Subscript>F<Subscript>1</Subscript>-ATPase in Japanese flounder and red sea bream associated with rearing temperatures

ABSTRACT:   Fast skeletal muscles of Japanese flounder Paralichthys olivaceus and red sea bream Pagrus major were examined for quantitative and qualitative changes of mitochondrial ATP synthase (F_oF₁-ATPase) in association with rearing temperatures. The specific activities of F_oF₁-ATPase from Japanese flounder reared at 10°C, 15°C and 25°C for 4 weeks were determined to be 81 ± 11, 74 ± 13 and 83 ± 11 nmol/min·mg mitochondrial protein, respectively. The corresponding activity from red sea bream reared at 8°C for 5 weeks was determined to be 65 ± 9 nmol/min·mg mitochondrial protein, which was higher than 33 ± 9 nmol/min·mg mitochondrial protein in fish reared at 23°C. The contents of α- and β-F₁-ATPase in total mitochondrial proteins were not significantly different between fish reared at different temperatures for the two fish species. However, the contents of β-F₁-ATPase in the total fast skeletal muscle extracts, prepared from Japanese flounder reared at 10°C, were 2.1- and 2.9-fold higher than those for fish reared at 15°C and 25°C, respectively. The corresponding content from red seabream reared at 8°C was 2.2-fold higher than that for fish reared at 23°C. Therefore, the changes in F_oF₁-ATPase depending on rearing temperatures were species-specific.     

Effects of rearing temperature and dietary short‐chain fructooligosaccharides supplementation on allochthonous gut microbiota,digestive enzymes activities and intestine health of turbot (Scophthalmus maximus L.) juveniles

The gut microbiomes of fish play important roles in host development, digestion and health. Evidence suggests that abiotic factors, such as diet and rearing temperature, could affect fish gut microbiota. In this study, the effect of dietary short‐chain fructooligosaccharides (scFOS) on turbot intestinal health, microbiota and digestive enzymes was investigated at two rearing temperatures: 15 and 20 °C. Four practical diets were supplemented with scFOS at 0, 5, 10 and 20 g kg^?1. scFOS did not affect fish performance. PCR‐DGGE did not show differences in bacterial profiles between dietary treatments; however, the number of operational taxonomic units, richness and diversity were higher at 20 °C. Enzyme activities in the foregut were not affected by rearing temperature, while in the hindgut, enzyme activities were higher at 15 °C. Total alkaline protease, α‐amylase and lipase activities in the foregut were higher in fish fed 20 g kg^?1 scFOS. Prebiotic supplementation had no effect on hindgut α‐amylase activity, while lipase activity of fish reared at 20 °C was higher in fish fed diet with 5 g kg^?1 scFOS. No differences were observed in intestinal morphology. This was the first study to simultaneously evaluate the effect of dietary prebiotic level and rearing temperature on fish intestinal microbiota and health.     

Effects of light spectrum on growth and physiological status of gilthead seabream Sparus aurata and rainbow trout Oncorhynchus mykiss reared under recirculating system conditions

Previously reported data clearly indicate that depending on species, ambient light spectrum can affect fish growth, physiology, behaviour, reproduction, etc. Since light spectrum can be easily controlled in intensive indoor fish farming facilities, the present study aimed to investigate the effects of coloured light on growth performance (body weight, growth rate, food utilization, carcass composition, etc.) and physiological status (blood and plasma parameters, brain neurotransmitters, tissue fatty acid composition, etc.) of two widely reared fish species, gilthead seabream Sparus aurata and rainbow trout Oncorhynchus mykiss. For this purpose, 240 specimens of S. aurata (29.8 ± 0.13 g) and 60 specimens of O. mykiss (71.4 ± 0.30 g) were exposed to white (full-spectrum, fluorescent lamps), red (605 nm) and blue (480 nm) light (lamps covered with appropriate filters) for 11 weeks under recirculating water systems. Blue light had a significantly negative impact on O. mykiss growth performance accompanied with reduced liver total lipids and plasma glucose and increased brain serotonergic and dopaminergic activity. In the case of S. aurata, red light significantly increased brain dopaminergic activity, while a tendency towards reduced growth was also observed. Since these results indicated the establishment of stressful conditions, O. mykiss and S. aurata should not be reared under blue and red light, respectively. For each species, the effect of the remaining light colours tested, was not clearly differentiated so that an appropriate light spectrum for the most efficient farming of O. mykiss and S. aurata could not be suggested, at least for the time period examined. Nevertheless, present results suggest that light colour should be regarded as a rearing factor worth to be further investigated, especially when recirculating systems are concerned.     

Rainbow trout (Oncorhynchus mykiss,Walbaum ) develop a more robust body shape under organic rearing

Prolonged exposure to captive conditions has led to the development of a rainbow trout ‘farmed’ phenotype, which is different from that of wild trout. Selection for desirable productive traits in hatcheries has resulted in the development of some morphological traits that are maladaptive in nature. The recent development of organic aquaculture, guided by the well‐being of the fish, could potentially produce a new farmed phenotype that would be more adaptive in nature. In this study, rainbow trout reared in intensive and organic farms were compared by means of shape analysis, to detect patterns of shape variation associated with rearing environment. The results of this study highlight a significant effect of the rearing method on rainbow trout shape: organically reared trout showed a higher body profile, in particular in the head and trunk regions, shorter median fins and a deeper caudal peduncle. A combined effect of density and habitat complexity could have contributed to the observed shape differences: in organic rearing systems, lower densities and steady water could increase territoriality and aggressive interactions, promoting body designs more functional in rapid attacks and escapes.     

Seawater tolerance and morphological assessment of yearling sea trout (Salmo trutta L.)

To study seawater tolerance and make a morphological assessment of yearling sea trout, their maturation and smoltification signs were estimated in two different fish rearing systems in Latvia,—in recirculating and flow-through system. For yearling sea trout (Salmo trutta L.), fish hypo-osmoregulatory ability was evaluated using seawater tolerance test, also fish morphological parameters were analyzed from January to May. April and May are months when smoltification occurs for wild sea trout in nature. Sea trout from recirculating system initially showed better growth, higher survival rate, and hypo-osmoregulatory ability, due to the elevated rearing water temperature. However, the situation completely changed in May when natural smoltification peaked and wild sea trout migration to the sea occurred—survival rate for yearling sea trout reared in recirculating system dropped to zero. Nevertheless, survival of fish from flow-through system increased, reaching 33%. Furthermore, in flow-through systems, sea trout that survived had significantly lower condition factor, also silvering level was higher compared those who did not survive. Weight was not a useful factor for determination of sea trout smoltification. The most part of yearling sea trout did not smoltify at the age of 1 year and should be reared for one more year before release in natural watercourses foreseen for migration to the sea.

     

Effects of rearing temperature on growth,metabolism and thermal tolerance of juvenile sea cucumber,Apostichopus japonicus Selenka: critical thermal maximum (CTmax) and hsps gene expression

Effects of different rearing temperatures (16, 21 and 26°C) on growth, metabolic performance and thermal tolerance of juvenile sea cucumber Apostichopus japonicus (initial body weight 7.72 ± 0.96 g, mean ±SD) were investigated in this study. During the 40‐day experiment, growth, metabolic performance, food intake and energy budget at different reared temperatures were determined. Sea cucumbers rearing at 16°C obtained better growth (final body weight 11.96 ± 0.35 g) than those reared at 21 (10.33 ± 0.41 g) and 26°C (8.31 ± 0.19 g) (P < 0.05), and more energy was allocated for growth at 16°C (162.73 ±11.85 J g^?1 d^?1) than those at 21(79.61 ± 6.76 J g^?1 d^?1) and 26°C (27.07 ± 4.30 J g^?1 d^?1) (P < 0.05). Critical thermal maxima (CTmax) values of juvenile sea cucumbers reared at 16, 21 and 26°C were 33.1, 34.1 and 36.6°C, respectively, and the upregulation of hsps in sea cucumbers reared at 26°C was higher than those acclimated at lower temperatures (16 and 21°C), indicating that temperature acclimation could change the thermal tolerance of the sea cucumber, and CTmax and hsps were sensitive indicators of the sea cucumber's thermal tolerance.     

Effects of two culturing techniques on the growth, survival and larval quality of Dentex dentex Linnaeus, 1758

Common dentex larvae were reared using two culturing techniques, mesocosms and intensive rearing, to determine the principal culture parameters involved in the differences observed in growth, skeletal deformations and survival between the two rearing techniques. In growth, only dry weight of larvae of 40 days post‐hatching (dph) from mesocosms was significantly higher than larvae from intensive rearing. Significant differences were observed in survival at 40 dph (6.58% in mesocosms and 1.58% in intensive rearing) and in the incidence of skeletal deformations, both for percentage of deformed larvae and for some deformation types such as those related to vertebral column and to the caudal complex. Initial larval density and initial prey density and quality are the factors suspected to affect growth and survival performance, while skeletal deformities might also be affected by tank hydrodynamics.     

Ontogeny of thyroid tissue and tissue thyroid hormone clearance in rainbow trout embryos reared at two temperatures

The development of the thyroid tissue and clearance of thyroid hormones was studied in rainbow trout embryos reared at 8.5 and 5.5 °C, in order to determine if these events are affected by temperature in the same manner as growth rate. Although the change in growth rate was found to be predictable using the degree-day model, the pattern of whole body thyroid hormone clearance was not similarly related to rearing temperature. Further, although the thyroid follicles appeared at different times in embryos reared at the two temperatures, the differences could not be predicted using the degree-day model. These results suggest that the development of the thyroid tissue and the clearance of yolk hormones are not directly related to rearing temperature. Immunohistochemical staining of the thyroid tissue indicated two phases of thyroid activity, one associated with hormonogenesis, and the other a later phase of hormone release. However, there was no increase in 'whole body' thyroid hormone level associated with the time of appearance of colloid in the follicles. Further, evidence was presented to suggest that the extraction method used did not remove thyroid hormone associated with the colloid in the follicles. There was also evidence of sporadic increases in 'whole body' thyroid hormone content, after the development of phase 2 thyroid follicles which might indicate pulsatile release of T4 and T3.     

The effect of dietary vitamin K (phylloquinone and menadione) levels on the vertebral formation in mummichog Fundulus heteroclitus

SUMMARY: The effect of different dosages of vitamin K, either as phylloquinone or menadione sodium bisulfite (MSB), on vertebrae formation during growth of mummichog Fundulus heteroclitus was investigated. Diets without vitamin K caused a significantly higher incidence of bone deformity in fish than diets supplemented with MSB or phylloquinone ( P < 0.01). These results indicate that vitamin K is essential for mummichog. A massive dose of MSB caused a high incidence of bone deformity compared with a lower dose, while an increased dose of phylloquinone brought about no significant difference in deformity incidence. The vertebral deformity was significantly higher ( P < 0.01) in fish fed a vitamin K-free diet (Control Group) and MSB-supplemented diet (Group MSB25) compared to fish fed a phylloquinone-supplemented diet (Group PK1). These results indicate that phylloquinone is more suitable than MSB as a vitamin K source in fish feed.