Review of cadmium toxicity effects on fish: Oxidative stress and immune responses.

Cadmium (Cd) in aquatic environments can cause environmental toxicity to fish and induce oxidative stress owing to an excessive production of reactive oxygen species in fish bodies. Fish have developed various antioxidant systems to protect themselves from reactive oxygen species; thus, a change in antioxidant responses in fish can be a criterion for evaluating oxidative stress resulting from Cd exposure. Because Cd exposure may be recognized as an exogenous substance by a fish body, it may lead to the stimulation or suppression of its immune system. Various immune responses can be assessed to evaluate Cd toxicity in fish. This review aimed to identify the impacts of Cd exposure on oxidative stress and immunotoxicity in fish as well as identify accurate indicators of Cd toxicity in aquatic ecosystems.

Toxic Effects on Oxidative Stress, Neurotoxicity, Stress, and Immune Responses in Juvenile Olive Flounder, Paralichthys olivaceus, Exposed to Waterborne Hexavalent Chromium.

Juvenile Paralichthys olivaceus were exposed to waterborne hexavalent chromium at various concentrations (0, 0.5, 1.0, and 2.0 mg/L) for 10 days. After chromium exposure, the activities of superoxide dismutase and glutathione S-transferase, which are oxidative stress indicators, were significantly increased; however, the glutathione level was significantly reduced. Acetylcholinesterase activity as a neurotoxicity marker was significantly inhibited upon chromium exposure. Other stress indicators, including plasma cortisol and heat shock protein 70, were significantly increased. The immune response markers (lysozyme and immunoglobulin M) were significantly decreased after chromium exposure. These results suggest that exposure to environmental toxicity in the form of waterborne chromium at concentrations higher than 1.0 mg/L causes significant alterations in antioxidant responses, neurotransmitters, stress, and immune responses in juvenile olive flounders. This study will provide a basis for an accurate assessment of the toxic effects of hexavalent chromium on aquatic organisms.

Toxic effects of waterborne cadmium exposure on hematological parameters, oxidative stress, neurotoxicity, and heat shock protein 70 in juvenile olive flounder, Paralichthysolivaceus.

Cadmium-induced toxicity can affect fish embryo development, ion homeostasis regulation, energy metabolism, maturation and growth, stress response, and immunity. However, studies on the toxic effects of cadmium exposure to aquatic animals, particularly olive flounder (Paralichthys olivaceus), are limited. In this study, juvenile P. olivaceus (mean length, 12.9 ± 1.3 cm; mean weight, 23.1 ± 3.2 g) was exposed to waterborne cadmium (0, 50, 100, 200, and 400 µg/L) for 10 d. Hematological parameters, including hematocrit value and hemoglobin level, in P. olivaceus were significantly decreased after waterborne cadmium exposure. Plasma components such as calcium, glucose, cholesterol, glutamic-oxaloacetic transaminase, and glutamic-pyruvic transaminase were significantly altered via cadmium exposure. The activities of antioxidant enzymes, such as superoxide dismutase, catalase, and glutathione S-transferase, were significantly altered in P. olivaceus after cadmium exposure. Acetylcholinesterase activity was significantly inhibited upon waterborne cadmium exposure. Hepatic heat shock protein 70 was significantly upregulated in P. olivaceus after waterborne cadmium exposure. Therefore, waterborne cadmium at concentrations of >100 or 200 µg/L can induce physiological toxicity in P. olivaceus via changes in hematological parameters, antioxidant enzymes, neurotransmitters, and stress indicators.

The correlations data between whether liposomes are coated or not and flatfish's (paralichthys olivaceus) Streptococcus parauberis antibody formation ability and survival rates.

Streptococcus parauberis is a known etiologic agent that causes damage leading to death in flatfish (paralichthys olivaceus). Liposomes were used to deliver streptococcal oral vaccines to the intestinal mucous membranes of paralichthys olivaceus. The liposomes were coated for stabilization, and stability was measured with high performance liquid chromatography (HPLC). The liposomes were stable until day nine and were orally administered to flatfish as a vaccine. The resultant antibody titers were analyzed. The titers resulting from the uncoated liposomes were highest two weeks after the oral administration, and those resulting from the coated liposomes were highest one week after boosting. In addition, the bacteria were subcutaneously injected to artificially infect flatfish and the survival rates and relative survival rates were analyzed. The coated liposomes were found to yield the highest survival rate.

Molecular characterization, expression and functional analysis of peptidoglycan recognition protein-SC2 from rock bream, Oplegnathus fasciatus.

Peptidoglycan recognition proteins are members of the family of pattern recognition receptors (PRRs), that play important roles in the recognition of peptidoglycan and various biological processes. In this study, we have characterized peptidoglycan recognition protein-SC2 (PGRP-SC2) in rock bream (Oplegnathus fasciatus) (RbPGRP-SC2) and analysed its expression in various tissues after pathogen challenge. A sequence alignment revealed that the residues essential to zinc binding of the deduced protein were highly conserved among all the organisms. Phylogenetic analysis revealed that RbPGRP-SC2 is most closely related to the large yellow croaker PGRP-SC2. RbPGRP-SC2 was ubiquitously expressed in all tissues analysed, predominantly distributed in muscle and skin. After challenge with microbial pathogens (Edwardsiella piscicida), Streptococcus iniae or red seabream iridovirus [RSIV]), RbPGRP-SC2 was up-regulated in all the tissues examined, especially in liver. We produced recombinant RbPGRP-SC2 (rRbPGRP-SC2) using an Escherichia coli expression system. The rRbPGRP-SC2 had agglutination activity towards both Gram-negative (E. piscicida) and Gram-positive bacteria (S. iniae). In addition, rRbPGRP-SC2 induced leukocyte apoptosis and promoted leukocyte phagocytosis. These results suggest that the RbPGRP-SC2 plays an important role in the immune system and in maintaining cellular homeostasis of rock bream.

Stability of virus-like particles of red-spotted grouper nervous necrosis virus in the aqueous state, and the vaccine potential of lyophilized particles.

Virus-like particles (VLPs) are multi protein complexes mimicking the structural properties of the native virus. The development of freeze-dried formulations of such complex protein structures remains a challenge. Red-spotted grouper nervous necrosis virus (RGNNV) causes mass mortality in fish culture, and RGNNV VLPs have been suggested to be promising vaccine candidates. In the present study, the stability of RGNNV VLPs in the liquid state was investigated over a 4-week period, along with the influence of freeze-drying on VLP stability. RGNNV VLPs were completely degraded after one week at 37 °C followed by 3 weeks at ambient temperature, and they were partially degraded after 4 weeks at 4 °C. Therefore, the inherent stability of RGNNV VLP in an aqueous milieu is insufficient for long-term storage. When RGNNV VLPs were freeze-dried in the presence or absence of sugar stabilizers, sorbitol was found to improve VLP stability whereas mannitol reduced it. VLP preparations freeze-dried with sorbitol or without stabilizer were as immunogenic as control (non-freeze dried) VLPs, whereas VLPs freeze-dried in mannitol were less immunogenic. These results indicate that freeze-dried RGNNV VLPs have potential as vaccines.

Oral vaccination through voluntary consumption of the convict grouper Epinephelus septemfasciatus with yeast producing the capsid protein of red-spotted grouper nervous necrosis virus.

Nervous necrosis viruses (NNV) cause serious economic losses in marine fish cultivation. The red-spotted grouper NNV (RGNNV) is the most common species of NNV worldwide. There have been many efforts to develop prophylactic NNV vaccines, and various types of vaccine candidate have been suggested. However, most were designed as injectable vaccines, which are not suitable for large-scale vaccination and cause too much stress to the fish. Oral vaccination through voluntary feeding is an ideal way to provide protective immunity to fish. In the present study, recombinant Saccharomyces cerevisiae producing RGNNV capsid protein was used as oral vaccine. The recombinant yeast was prepared in freeze-dried form after disruption. Convict groupers were divided into three groups, control, and oral and parenteral vaccination groups, each consisting of 700 fishes. The control group received no treatment, the parenteral group received one intraperitoneal injection of RGNNV virus-like particles, and the oral vaccination group consumed feed containing the lysed recombinant yeast; voluntary intake was allowed four times at one-week intervals. Both vaccination groups produced serum RGNNV neutralizing antibody titers of >103 (log 2, 9.96), sustained for at least 95days post-immunization. In addition, in response to challenge with RGNNV both groups suffered significantly reduced mortality and had reduced brain RGNNV titers. These results indicate that recombinant yeast-based oral fish vaccines have great potential for large-scale vaccination.

The first report of CD2 associated protein gene, in a teleost (Rock bream, Oplegnathus fasciatus): An investigation of the immune response upon infection with several pathogens.

CD2 is expressed on the surfaces of virtually all T cells and natural killer (NK) cells. In mammals, the CD2 molecule is 50 kDa. The cytoplasmic tail of CD2 interacts with CD2-associated protein (CD2AP), which plays an important role in mediating the trigger signal in outer magnetic pole cells. In this study, we identified CD2AP from rock bream and investigated its gene expression. The ORF of CD2AP (1950 bp) encodes 650 amino acids (aa). CD2AP has a Src homology 3 (SH3) domain. Quantitative real-time PCR analysis revealed that CD2AP shows higher expression in the gills and skin. Under experimental challenge, CD2AP gene expression was increased as relative to the control after 7 days. This result will improve our understanding of blood vessels in teleost fish, and will provide a basis for the study of CD2-related genes.

Molecular cloning and expression of cDNAs for two distinct granulocyte colony stimulating factor genes from black rockfish Sebastes schlegelii.

Granulocyte-colony stimulating factor (G-CSF) is a cytokine that stimulates the proliferation and differentiation of hematopoietic progenitor cells committed to the neutrophil/granulocyte lineage. Here, we report the two distinct granulocyte colony stimulating factor homologues from black rockfish Sebastes schlegelii. The G-CSF homologue cDNAs were isolated from the black rockfish LPS or Con A/PMA stimulated leukocyte cDNA libraries. The cDNA for the Black rockfish G-CSF-1 homologue predicts a peptide of 202 amino acids that is the closest to the Bastard halibut (Paralichthys olivaceus) G-CSF, whereas the cDNA of the Black rockfish G-CSF-2 homologue predicts a peptide of 212 amino acids that is the closest to the Fugu (Takifugu rubripes). In a healthy fish, the mRNAs of both G-CSF homologues were predominantly expressed in leukocytes, spleen, and gill. Expression of the black rockfish G-CSF-1 homologue was induced in peripheral blood leukocytes (PBLs) after stimulation with LPS, Con A/PMA, or Poly I:C, and G-CSF-2 homologue was strongly induced in PBLs after stimulation with Con A/PMA for 24 h only.