Distribution patterns of settlement-stage juveniles of <Emphasis Type="Italic">Girella punctata</Emphasis> and <Emphasis Type="Italic">Girella leonina</Emphasis> on the rocky coast of the Kanto–Izu region,Japan

The early life history of girellid fishes in Japanese waters is unclear, and little is known about their species-specific reproductive strategies. We examined seasonal changes of distribution patterns for settlement-stage juveniles of Girella punctata and Girella leonina on the rocky shore in the regions of Kanto and Izu, Japan, to infer the influence of the Kuroshio Current on their reproduction. We collected 813 settlement-stage juveniles mainly in Sagami Bay and genetically identified the species. The juveniles of G. punctata were collected on the rocky shore in Sagami Bay during April to August, with the abundant catch in May and June. Thus, we infer that juvenile G. punctata ubiquitously inhabit the rocky shore in the area in spring and summer. By contrast, juveniles of G. leonina were rarely collected in Sagami Bay, with a total catch of only 66. Notably, no juveniles were collected during the wintertime in Sagami Bay, although an abundant catch of G. leonina had been previously reported for Sagami Nada off Sagami Bay during January to March. This clear-cut difference between the areas likely reflects the difference in proximity to the path of the Kuroshio Current. We expect that the Kuroshio Current strongly influences the reproductive success of G. leonina.     

Effects of dietary riboflavin levels on antioxidant defense of the juvenile grouper <Emphasis Type="Italic">Epinephelus coioides</Emphasis>

This study was conducted to evaluate the effects of dietary riboflavin on antioxidant defense in the juvenile grouper Epinephelus coioides. Graded levels of riboflavin (0.9, 1.6, 4.4, 6.7, 12.9 and 19.4 mg kg⁻¹ dry diet) were fed to grouper juveniles (mean weight: 14.90 ± 0.46 g) for 12 weeks. Higher levels of liver thiobarbituric acid reactive substances (TBARS) content were observed in grouper fed low doses (0.9 and 1.6 mg kg⁻¹ diet) of riboflavin. Both liver glutathione reductase (GR) activity and its activation coefficient (GR-AC) poorly responded to riboflavin deficiency. In addition, other indices of the glutathione-dependent defense system, including the activities of glutathione peroxidase (GSH-PX) and glutathione-S-transferase (GST), and the content of glutathione (GSH), were also non-significantly affected by dietary riboflavin levels. However, the activities of liver superoxide dismutase (SOD) and catalase (CAT) were significantly lower in fish fed 0.9 mg kg⁻¹ diet, with a positive correlation between the different groups. In conclusion, the present study indicated that the juvenile grouper fed the riboflavin-unsupplemented diet was susceptible to lipid peroxidation (LPO), with lower SOD and CAT activities in the liver. However, the glutathione-dependent defense system of grouper was not affected by dietary riboflavin levels.     

The effects of hairtail protein hydrolysate–Fe<Superscript>2+</Superscript> complexes on growth and non-specific immune response of red swamp crayfish (<Emphasis Type="Italic">Procambarus clarkii</Emphasis>)

The experiment was conducted to investigate the effects of prepared hairtail protein hydrolysate–Fe²⁺ (PH–Fe²⁺) complexes on growth and non-specific immune responses of crayfish (Procambarus clarkii). The diets with five different levels of PH–Fe²⁺ (200, 400, 600, 800, and 1000 mg/kg) were fed to crayfish for 60 days. The results indicated that the survival rate, weight gain percentage, specific growth rate, and muscle index of crayfish with 400–1000 mg/kg of PH–Fe²⁺ feeding were significantly higher, 9.94–10.10, 8.55–8.72, 0.24–0.29 %/day, and 2.39–3.05 %, respectively, than the control values after 60 days of feeding. Additionally, after 30 days of feeding, 200–400 mg/kg of PH–Fe²⁺ showed no significant (P > 0.05) improvement effect on activities of superoxide dismutase (SOD), phenoloxidase (PO), lysozyme (LSZ), and acid phosphatase (ACP) in serum or hepatopancreas of crayfish. However, significant improvement effects were observed in 800–1000 mg/kg groups. After 60 days of feeding, 400–600 mg/kg of PH–Fe²⁺ significantly (P < 0.05) improved the activity of SOD and LSZ, but did not affect the activity of PO and ACP significantly (P > 0.05). Moreover, the activities of SOD, PO, LSZ, and ACP in serum and hepatopancreas were all significantly enhanced upon treatment with 800–1000 mg/kg of PH–Fe²⁺. For these reasons, PH–Fe²⁺ can be recommended as a supplement in crayfish feed to increase the growth and immunity.     

Ontogenetic development of digestive enzymes and effect of starvation in miiuy croaker <Emphasis Type="Italic">Miichthys miiuy</Emphasis> larvae

The ontogenetic development of the digestive enzymes amylase, lipase, trypsin, and alkaline phosphatase and the effect of starvation in miiuy croaker Miichthys miiuy larvae were studied. The activities of these enzymes were detected prior to exogenous feeding, but their developmental patterns differed remarkably. Trypsin activity continuously increased from 2 days after hatching (dah), peaked on 20 dah, and decreased to 25 dah at weaning. Alkaline phosphatase activity oscillated at low levels within a small range after the first feeding on 3 dah. In contrast, amylase and lipase activities followed the general developmental pattern that has been characterized in fish larvae, with a succession of increases or decreases. Amylase, lipase, and trypsin activities generally started to increase or decrease at transitions from endogenous to exogenous feeding or diet changes, suggesting that these enzymatic activities can be modulated by feeding modes. The activities of all the enzymes remained stable from 25 dah onwards, coinciding with the formation of gastric glands and pyloric caecum. These results imply that specific activities of these enzymes underwent changes due to morphological and physiological modifications or diet shift during larval development but that they became stable after the development of the digestive organs and associated glands was fully completed and the organs/glands functioned. Trypsin and alkaline phosphatase were more sensitive to starvation than amylase and lipase because delayed feeding up to 2 days after mouth opening was able to adversely affect their activities. Enzyme activities did not significantly differ among feeding groups during endogenous feeding; however, all activities were remarkably reduced when delayed feeding was within 3 days after mouth opening. Initiation of larvae feeding should occur within 2 days after mouth opening so that good growth and survival can be obtained in the culture.     

Effect of nitrite exposure on oxygen-carrying capacity and gene expression of <Emphasis Type="Italic">NF-κB</Emphasis>/<Emphasis Type="Italic">HIF-1α</Emphasis> pathway in gill of bighead carp (<Emphasis Type="Italic">Aristichthys nobilis</Emphasis>)

Nitrite (NO₂^?) contamination of water can severely impact the health of aquatic life and is a major concern for commercial aquaculture. In order to study the effect of nitrite on Aristichthys nobilis, we investigated the oxygen-carrying capacity, NF-κB/HIF-1α pathway, and the gill tissue structure under nitrite stress. In the current study, bighead carp (initial weight 180.05?±?0.092 g) were exposed to nitrite (48.634 mg/L) for 96 h and then for 96 h recovery test. After nitrite exposure for 6 h, hypoxia-inducible factor-1α (HIF-1α) and nuclear factor-κB (NF-κB) mRNA expression increased significantly in the gill of bighead carp (P?<?0.05). After nitrite exposure for 12 h, hemoglobin (Hb) and methemoglobin reductase (MHBR) content in blood decreased significantly (P?<?0.05); TLR4 mRNA expression increased significantly (P?<?0.05). After nitrite exposure for 24 h, methemoglobin (MetHb) content increased significantly (P?<?0.05). After recovery test, all the indicators except TLR4 mRNA expression level recovered to initial level. In conclusion, nitrite exposure can affect hemoglobin dynamics, as oxidization of nitrite by hemoglobin results in the reduction of Hb to MetHb leading to hypoxia and nitrite exposure can also result into gill tissue damage. In the face of nitrite exposure, NF-κB and HIF-1α mRNA expression level increased immediately to protect the body from oxidative damage and eased hypoxic condition caused by nitrite. It was also observed that nitrite damage is recoverable in Aristichthys nobilis, but it may be need more than 96 h.     

Effects of <Emphasis Type="Italic">Euphorbia hirta</Emphasis> plant leaf extract on growth performance,hematological and organosomatic indices of hybrid catfish, <Emphasis Type="Italic">Clarias macrocephalus </Emphasis>× <Emphasis Type="Italic">C. gariepinus</Emphasis>

This study was conducted to evaluate the effects of Euphorbia hirta leaf extract on the growth performance, hematological and organosomatic indices of hybrid catfish, Clarias macrocephalus?×?C. gariepinus. The fish were treated with 0 (control), 300, 500 and 800 mg/kg (ppm) for 90 days. The weight gain, average daily growth rate, and specific growth rate were at significantly higher levels in fish from all the treatment groups on days 75 and 90, while the feed efficiency and protein efficiency ratio were consistently higher in fish from all the treatment groups from day 60 up until day 90. The feed conversion rate significantly decreased from day 60 until day 90 in all treatment groups when compared with the control group, and the survival rate was significantly different from day 30 until day 90; a consistently higher rate was observed in fish fed 800 mg/kg. The highest viscerosomatic index and intraperitoneal fat were observed in the group fed 500 mg/kg (p?<?0.05). The hepatosomatic index was significantly increased alongside increasing levels of E. hirta extract. The total white blood cell count in the control group was significantly higher on day 30, but on day 90 all the treatment groups showed higher levels. Hematocrit percentage was significantly different on days 30 and 90. Lymphocyte, eosinophil and thrombocyte levels were shown to be significantly different (p?<?0.05) when different groups of fish were compared. In conclusion, 300 mg/kg of E. hirta leaf extract could improve growth performance, hematological and some organosomatic indices in hybrid catfish.     

尼罗罗非鱼无乳链球菌基因缺失株I>cpsE和I>neuA的构建及其生物学特性

为探究尼罗罗非鱼无乳链球菌(GBS)荚膜多糖合成基因cpsE和neuA对菌株生物学特性的影响,本研究利用同源重组的方法,构建了GBS的cpsE与neuA的单基因缺失突变株.具体方法为:用Infusion-PCR的方法分别构建带有氯霉素抗性基因的cpsE与neuA基因敲除重组质粒pSET4s-cpsE和pSET4s-neuA.将构建好的质粒电转化入GBS感受态细胞中,通过改变培养温度实现双交换和质粒丢失,最后经氯霉素抗性筛选获得疑似敲除株.通过菌落PCR、RT-PCR及DNA测序等方法对疑似敲除株进行验证.结果显示GBS的两个突变株△cpsE和△neuA被成功构建.在此基础上,通过生物学功能分析比较基因缺失突变株△cpsE、△neuA与野生株在菌株生长速率、荚膜多糖厚度、唾液酸含量和毒力方面的差异.结果发现缺失突变株△cpsE和△neuA的生长速度与野生株无显著差异,但荚膜多糖厚度、唾液酸含量和菌株毒力均显著低于野生株.进一步研究显示,cpsE是鱼源GBS荚膜多糖合成的关键基因,neuA基因则是荚膜多糖唾液酸化的关键基因,它们的缺失导致了GBS荚膜唾液酸含量的降低,且显著降低了菌株的毒力.     

A comparison of growth and biochemical composition of <Emphasis Type="Italic">Mytilus galloprovincialis</Emphasis> (Lmk.) and <Emphasis Type="Italic">Mytilus edulis</Emphasis> (L.) on the West coast of Cotentin,Normandy, France 1999–2000

The blue mussel, Mytilus edulis has been reared along the Normandy coast line since the 1960s. The gonadal cycle of this mussel species shows a sharp decrease in meat quality during the winter period after spawning. This decline in meat quality is so severe that sales have to be suspended from December to July. Another species of mussel, Mytilus galloprovincialis, which is reared in the lagoons along the French shore of the Mediterranean Sea has a different spawning cycle. An experiment was undertaken to study the meat quality of M. Galloprovincialis throughout the year after the spat were transferred from the Mediterranean Sea to the Normandy coast. This species showed an immediate adaptation to the gonadal cycle of M. edulis. Despite suggestions from researchers, no interest was expressed to consider such transfers of M. galloprovincialis in the future.     

Development of digestive enzyme activity in larvae of spotted sand bass <Emphasis Type="Italic">Paralabrax maculatofasciatus</Emphasis> II: Electrophoretic analysis

The activities of several digestive enzymes during larval development of the spotted sand bass (Paralabrax maculatofasciatus) were evaluated using electrophoretic techniques. The results show the presence of three isoforms of alkaline protease from day 2 after hatching (ah) and the early appearance of one pepsin-like band from day 12 ah onwards. In addition, two lipase bands first appeared on day 2 ah, and there was a change in the molecular weight of one band from day 15 ah onwards. Several α-amylase isoforms were observed from hatching up to day 5 ah. These results indicate that the important digestive enzymes develop rapidly in these larvae, supporting the possibility of early weaning at day 12 ah using artificial diets.     

Evaluation of interferon gamma (IFN-γ) of <Emphasis Type="Italic">Labeo rohita</Emphasis> as an immunomodulator: in vitro expression model

Interferon gamma (IFN-γ) or type II interferon is a cytokine that is critical for innate and adaptive immunity against viral and some bacterial and protozoal infections. The importance of IFN-γ in the immune system lies in its ability to inhibit viral replication directly and most importantly from its immunomodulatory effects. Previously, we successfully co-administered IFN-γ along with GAPDH gene of Edwardsiella tarda as bicistronic DNA vaccine in Labeo rohita. In order to ascertain the individual role of IFN-γ, the present study involves cloning and expression of 552-bp IFN-γ open-reading frame (ORF) of L. rohita in striped snakehead (SSN-1) cell line using eukaryotic expression vector system (pQE-TriSystem) followed by transfection in peripheral blood lymphocytes (PBMCs) to evaluate its immunomodulatory ability in comparison to polyinosinic-polycytidylic acid (Poly I:C)-treated PBMCs. The 18.7-kDa protein, expressed in the pQE-IFNγ-transfected SSN-1 cells, reacted with anti-His antibody in Western blot confirming it to be recombinant IFN-γ, whereas the relative expression of IFN-γ, iNOS, Mx, and IL-1β genes in PBMCs was quantified at 24 h and 48 h post treatment by qPCR. The comparative kinetics of all four genes showed significantly (p?<?0.05) high upregulation pattern in both pQE-IFNγ-transfected cell group and Poly I:C-treated cell group demonstrating recombinant IFN-γ as an equally efficient inducer like Poly I:C. Thus, our in vitro experiment results highlight the immunomodulatory potential of recombinant IFN-γ as an analogue to synthetic Poly I:C which warranted future studies to further explore the potential of recombinant IFN-γ as an effective vaccine adjuvant against different microbial invasion.     

Influence of cell volume changes on protein synthesis in isolated hepatocytes of air-breathing walking catfish (<Emphasis Type="Italic">Clarias batrachus</Emphasis>)

The present study aimed at determining the effect of cell volume changes on protein synthesis, measured as the incorporation of [³H]leucine into acid-precipitable protein, in isolated hepatocytes of air-breathing walking catfish (Clarias batrachus). The rate of protein synthesis, which was recorded to be 10.02 ± 0.10 (n = 25) nmoles mg⁻¹ cell protein h⁻¹ in isotonic incubation conditions, increased/decreased significantly by 18 and 48%, respectively, following hypo- (−80 mOsmol l⁻¹)/hypertonic (+80 mOsmol l⁻¹) incubation conditions (adjusted with NaCl), with an accompanying increase/decrease of hepatic cell volume by 12 and 20%, respectively. Similar cell volume-sensitive changes of protein synthesis were also observed when the anisotonicity of incubation medium was adjusted with mannitol. Increase of hepatic cell volume by 9%, due to addition of glutamine plus glycine (5 mM each) to the isotonic control incubation medium, led to a significant increase of protein synthesis by 14%. Decrease of hepatic cell volume by 15 and 18%, due to addition of dibutyl-cAMP and adenosine in isotonic control incubation medium, led to a significant decrease of protein synthesis by 30 and 34%, respectively. Thus, it appears that the increase/decrease of hepatic cell volume, caused either by changing the extracellular osmolarity or by the presence of amino acids or certain other metabolites, leads to increase/decrease of protein synthesis, respectively, and shows a direct correction (r = 0.99) between the hepatic cell volume and protein synthesis in walking catfish. These cell volume-sensitive changes of protein synthesis probably help this walking catfish in fine tuning the different metabolic pathways for better adaptation during cell volume changes and also to avoid the adverse affects of osmotic stress. This is the first report of cell volume-sensitive changes of protein synthesis in hepatic cells of any teleosts.     

Biochemical properties of the sensitivity to GABA<Subscript>A</Subscript>ergic ligands,Cl<Superscript>−</Superscript>/HCO<Subscript>3</Subscript><Superscript>−</Superscript>-ATPase isolated from fish (<Emphasis Type="Italic">Cyprinus carpio</Emphasis>) olfactory mucosa and brain

This paper presents a comparative study of the roles of Cl^? and HCO₃ ^? in the functioning of the GABA_AR-associated Cl^?/HCO₃ ^?-ATPase of the plasma membranes of the olfactory sensory neurons (OSNs) and mature brain neurons (MBNs) of fish. The ATPase activity of OSNs and its dephosphorylation were increased twofold by Cl^?(15–30 mmol l^?1), whereas the enzyme from MBNs was not significantly affected by Cl^?. By contrast, HCO₃ ^?(15–30 mmol l^?1) significantly activated the MBN enzyme and its dephosphorylation, but had no effect on the OSN ATPase. The maximum ATPase activity and protein dephosphorylation was observed in the presence of both Cl^?(15 mmol l^?1)/HCO₃ ^?(27 mmol l^?1) and these activities were inhibited in the presence of picrotoxin (100 μmol l^?1), bumetanide (150 μmol l^?1), and DIDS (1000 μmol l^?1). SDS-PAGE revealed that ATPases purified from the neuronal membrane have a subunit with molecular mass of ~?56 kDa that binds [³H]muscimol and [³H]flunitrazepam. Direct phosphorylation of the enzymes in the presence of ATP-γ-³²P and Mg²⁺, as well as Cl^?/HCO₃ ^? sensitive dephosphorylation, is also associated with this 56 kDa peptide. Both preparations also showed one subunit with molecular mass 56 kDa that was immunoreactive with GABA_AR β3 subunit. The use of a fluorescent dye for Cl^? demonstrated that HCO₃ ^?(27 mmol l^?1) causes a twofold increase in Cl^? influx into proteoliposomes containing reconstituted ATPases from MBNs, but HCO₃ ^? had no effect on the reconstituted enzyme from OSNs. These data are the first to demonstrate a differential effect of Cl^? and HCO₃ ^? in the regulation of the Cl^?/HCO₃ ^?-ATPases functioning in neurons with different specializations.     

Are hatchery‐reared abalone naïve of predators? Comparing the behaviours of wild and hatchery‐reared northern abalone,Haliotis kamtschatkana (Jonas, 1845)

Abalone populations have declined worldwide, generating interest in enhancement using hatchery‐reared individuals. In many cases, such restoration efforts have met with limited success due to high predator‐induced mortality rates. Furthermore, the mortality rates of outplanted hatchery abalone are often considerably higher than for wild individuals. This study uses northern abalone (Haliotis kamtschatkana) as a case study to determine whether hatchery‐reared abalone behave differently than their wild counterparts. In the field, outplanted hatchery‐reared abalone were significantly less responsive than wild abalone, in terms of number of abalone responding and intensity of response, to nearby movement and to physical contact with an inert probe. Also, when encountering a cue to which all abalone responded (a seastar predator), hatchery‐reared individuals remained subdued. Anti‐predator behavioural deficits in hatchery‐reared abalone were more pronounced in 4‐year‐old individuals than in 1‐year‐old individuals, suggesting an influence of either age or amount of time spent in the hatchery environment. These behavioural differences are expected to increase the vulnerability of hatchery‐reared abalone to predators, and are likely a major cause of their elevated predator‐induced mortality when outplanted.     

PGF<Subscript>2α</Subscript> and gonadal steroid plasma levels of successful and unsuccessful spawning <Emphasis Type="Italic">Piaractus mesopotamicus</Emphasis> (Teleostei,Characiformes) females

Gonadal steroid and prostaglandin F₂α (PGF_2α) plasma levels were evaluated in successfully (SP) and unsuccessfully ovulated (UN) female Piaractus mesopotamicus. Forty-one females were injected with crude carp pituitary extract (0.6 and 5.4 mg kg^?1 with a 24-h interval between the doses) and sampled to determine the plasma concentration of 17β-estradiol (E₂), 17α-hydroxyprogesterone (17α-OHP), 17α,20β-dihydroxy-4-pregnen-3-one (DHP), PGF_2α, and testosterone (T) after each injection (first—A1 and second—A2), and at the time of ovulation for SP and UN. Two clusters were obtained using multivariate analysis: 1—composed of all A1, all A2, and some UN; and 2—composed of all SP and some UN. Median values of E₂ plasma levels were similar between clusters; however, plasma levels of T, 17α-OHP, DHP, and PGF_2α of cluster 2 (predominantly formed by SP) were higher than those of cluster 1. Since cluster 2 contained all SP and females of this cluster presented higher levels of PGF_2α, T, 17α-OHP, and DHP, here we evidently shown in an unprecedented manner that concomitant increased levels of these substances were associated with successful ovulation in this species, but such an increase was not determinant for successful ovulation due to the presence of some UN females in the same cluster 2. These findings highlight the unexplored potential of PGF_2α to be used as an accessory tool for inducing successful ovulation for fish farming purposes.     

Protective effect of squilla chitosan–silver nanoparticles for <Emphasis Type="Italic">Dicentrarchus labrax</Emphasis> larvae infected with <Emphasis Type="Italic">Vibrio anguillarum</Emphasis>

Antimicrobial nanoparticle therapy was proposed as an alternative strategy to reduce the use of antibiotics in larval-rearing systems. Antibacterial potential of the prepared squilla chitosan–silver nanoparticles and its protective effect on Dicentrarchus labrax (sea bass) larvae in the early stages were studied against Vibrio angularium. Different concentrations of squilla chitosan (Csq) and squilla chitosan–silver nanoparticles (Csq–AgNps) (1, 2, 5, 10, and 20 %) were, in vitro, tested against V.anguillarum and expressed as a role of Log₁₀ mean. Sea bass larvae were treated using: 10 % Csq and 5 % Csq–AgNps as effective inhibitory concentrations against the pathogen either encapsulated during the feeding regime or added directly to the model system via the water from the onset of 4 weeks. The long-term administration of Csq–AgNps through enriched food for both non-infected and infected systems had survival % of 74.5 ± 1.5 and 72.5 ± 2.5, respectively. Larval clinical observations using Csq–AgNps were studied compared with the two controls. The current study found that 5 % encapsulated Csq–AgNps was enough to suppress infection and considered as an alternative to antibiotics in controlling virulent fish pathogens.     

Species differences and effects of soft coral extracts from <Emphasis Type="Italic">Sinnularia maximus</Emphasis> on the expression of cytochrome P4501A and 2N in butterflyfishes (<Emphasis Type="Italic">Chaetodon</Emphasis> spp.)

Cytochrome P450 (CYP) has been shown to confer resistance in numerous terrestrial insects that consume potentially toxic secondary metabolites in plants, but fewer studies have examined the role of critical biotransformation enzymes in allowing marine organisms to consume chemically defended foods. This study examined the expression of CYP1A and CYP2N mRNAs in several butterflyfish species, which can feed on numerous chemically defended soft and hard corals. In addition, the effect of an extract from a soft coral (Sinnularia maxima) on expression of hepatic CYP1A and CYP2 mRNAs was also examined. Fish were fed extracts on days 1, 3 and 5, and expression was examined on day 5. Phylogenetic analyses of the CYP1A cDNA from 12 species of butterflyfish (DNA, amino acid) indicate well-separated groupings according to their feeding strategies. The non-coralline feeding Chaetodon xanthurus exhibited a 7-fold higher basal expression of CYP2N8 relative to the other species studied. Although induction of CYP2N7 expression was observed in C. punctatofasciatus, CYP1A and CYP2N was largely unaffected or diminished by extract treatment in the other species of butterflyfish. These results indicated groupings of feeding strategy with CYP1A phylogeny in Chaetodon, but generally unaltered expression of CYP1A and CYP2N following dietary treatment with an extract from a chemically defended soft coral suggesting an inconclusive role of these isoforms in the detoxification of chemicals in these extracts.     

Nucleotide sequence and mRNA expression analysis of <Emphasis Type="Italic">β</Emphasis>-actin gene in the orange-spotted grouper <Emphasis Type="Italic">Epinephelus coioides</Emphasis>

The full-length cDNA, encoding the orange-spotted grouper β-actin and spanning 1920 bp including a poly (A) tail, was cloned from its brain cDNA library. The open reading frame encodes a protein of 375 amino acids. Sequence analysis indicated that it contained the typical structural features of cytoplasmic actins, and showed higher homology with other vertebrate β-actin than any other members of the actin family. The partial genomic sequence indicated that the organization of the β-actin gene in the orange-spotted grouper might also be conserved. Northern blot analysis indicated that it was expressed at high levels in the brain, spleen, adipose tissue, ovary, and liver, but at low levels in the gill filament and heart, and at a very low level in the kidney. The expression of β-actin gene in the skeletal muscle was barely detectable. These results indicated that the expression of the orange-spotted grouper β-actin gene showed significant variation in different tissues. Therefore, caution should be taken when using β-actin gene as an internal control in the normalization of gene expression among tissues. Whereas, semi-quantitative RT-PCR analysis indicated that treatment with 17α–methyltestosterone (MT) had little effect on the mRNA expression of β-actin gene in the in vitro incubated hypothalamus, pituitary, and ovary fragments of the orange-spotted grouper, suggesting β-actin can be used as an internal control for RT-PCR analysis of MT effects on gene expression in these tissues.