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The mechanism for the elimination of xenobiotics undergoes three different phases of reactions in organisms. Among these, glutathione S-transferases (GSTs) are classified as phase II detoxification enzymes, catalyzing the conjugation of electrophilic substrates to glutathione or reduced hydroperoxides. This study aimed to investigate the molecular characteristics, detoxification functions, and immune responses of GST omega (LhGSTO1) and kappa (LhGSTK1) in redlip mullet. The open reading frames of LhGSTO1 (720 bp) and LhGSTK1 (687 bp) encoded proteins of 239 and 228 amino acids, respectively. Sequence analysis revealed that LhGSTO1 and LhGSTK1 possessed GSH-binding sites in their N-terminal domains. Substrate-binding sites in the C-terminal domain were exclusively identified in LhGSTO1. In the tissue-specific transcription profile analysis, both LhGSTO1 and LhGSTK1 were ubiquitously expressed in all tissues of healthy mullets. Temporal expression analysis of LhGSTO1 and LhGSTK1 in the blood showed that their expression was significantly modulated by polyinosinic:polycytidylic (poly I:C), lipopolysaccharide (LPS), and Lactococcus garvieae. Different chemical and cellular assays were performed to assess the detoxification and cellular protective abilities of the two proteins. A substrate specificity test using the recombinant proteins revealed that both proteins possessed specific activity toward 1-chloro-2,4-dinitrobenzene (CDNB). In the disk diffusion assay, the smallest clearance zones were observed for LhGSTO1 and LGSTK1 against CdCl2. In the cell protection assay, both LhGSTO1 and LhGSTK1 showed significant Cd detoxification ability compared to the control. Collectively, these results demonstrate that GST omega and kappa are involved in host defense against immune stimulants and xenobiotics in redlip mullet.

Scuticociliatosis, caused by the parasitic pathogen Miamiensis avidus, poses a significant threat to olive flounder farms in South Korea. Infected fish suffer from severe systemic infections affecting various organs, with potential secondary bacterial diseases. This study investigated the emergence of different M. avidus serotypes in 20 olive flounder farms on Jeju island, South Korea, from 2015 to 2020. Additionally, we identified Tenacibaculum maritimum as a co-infecting bacteria. Based on serotyping and monitoring data, we developed a trivalent vaccine targeting two serotypes of M. avidus and one strain of T. maritimum. The efficacy of the vaccine was evaluated under laboratory conditions and demonstrated a relative percentage of survival (RPS) of 75%, 80%, and 93% for M. avidus serotype I, M. avidus serotype II and T. maritimum, respectively. Furthermore, successful field trials conducted on four different olive flounder farms resulted in significantly higher survival rates (52%–76% RPS) and weight gains in vaccinated fish. Overall, this study presents an effective vaccine against M. avidus and T. maritimum infections in farmed olive flounder, making a valuable contribution to sustainable aquaculture in South Korea.

Lysozymes are well-known antibacterial enzymes that mainly target the peptidoglycan layer of the bacterial cell wall. Animal lysozymes are mainly categorized as g-type, c-type, and i-type based on protein sequence and structural differences. In this study, c-type (AcLysC) and g-like-type (AcLysG-like) lysozymes from Amphiprion clarkii were characterized in silico via expressional and functional approaches. According to in silico analysis, open reading frames of AcLysC and AcLysG-like were 429 bp and 570 bp, respectively, encoding the corresponding polypeptide chains with 142 and 189 amino acids. Elevated expression levels of AcLysC and AcLysG-like were observed in the liver and the heart tissues, respectively, as evidenced by quantitative real-time polymerase chain reaction assays. AcLysC and AcLysG-like transcript levels were upregulated in gills, head kidney, and blood cells following experimental immune stimulation. Recombinant AcLysC exhibited potent lytic activity against Vibrio anguillarum, whereas recombinant AcLysG-like showed remarkable antibacterial activity against Vibrio harveyi and Streptococcus parauberis, which was further evidenced by scanning electron microscopic imaging of destructed bacterial cell walls. The findings of this study collectively suggest the potential roles of AcLysC and AcLysG-like in host immune defense.

Enteromyxum leei has been identified as the causative agent of emaciation disease in a wide range of marine fish hosts. In this study, we aimed to determine the effect of the parasitic infection of Enteromyxum species on starry flounder that were cultured in aquaculture farms of Jeju island in Korea. As the mortality of cultured olive flounder Paralichthys olivaceus because of E. leei infection increased, some fish farms on Jeju island attempted to culture the starry flounder Platichthys stellatus, as an alternative. Myxosporeans with a developmental stage similar to E. leei were found in the intestines of cultured starry flounders. The partial 18S rDNA of myxosporeans showed 100% similarity with E. leei. To reveal the effect of E. leei infection on starry flounder, the intensity of E. leei infection measured using quantitative polymerase chain reaction, and the condition factor (CF) of fish were measured and analyzed statistically. The results showed that high-intensity E. leei infection significantly decreased the CF of the starry flounder. However, the pathogenicity of E. leei to starry flounder is low, considering its mortality and clinical signs.

Galectins, a family of β-galactoside-binding lectins, have emerged as soluble mediators in infected cells and pattern recognition receptors (PRRs) responsible for evoking and regulating innate immunity. The present study aimed to evaluate the role of galectin-1 in the host immune response of redlip mullet (Liza haematocheilia). We established a cDNA database for redlip mullet, and the cDNA sequence of galectin-1 (LhGal-1) was characterized. In silico analysis was performed, and the spatial and temporal expression patterns in gills and blood in response to lipopolysaccharide polyinosinic:polycytidylic acid, and Lactococcus garvieae were estimated via quantitative real-time PCR. Functional assays were conducted using recombinant protein to investigate carbohydrate binding, bacterial binding, and bacterial agglutination activity. LhGal-1 was composed of 135 amino acids. Conserved motifs (H-NPR, -N- and -W-E-R) within the carbohydrate recognition domain were found in LhGal-1. The tissue distribution revealed that the healthy stomach expressed high levels of LhGal-1. The temporal monitoring of LhGal-1 mRNA expression in the gill and blood showed its significant upregulation in response to immune challenges with different stimulants. rLhGal-1 exhibited binding activity in response to carbohydrates and bacteria. Moreover, the agglutination of rLhGal-1 against Escherichia coli was observed. Collectively, our findings suggest that LhGal-1 may function as a PRR in redlip mullet. Furthermore, LhGal-1 can be considered a significant gene to play a protective role in redlip mullet immune system.

Abstract Olive flounder (Paralichthys olivaceus) is the major species developed for aquaculture in South Korea. Over the long history of olive flounder aquaculture, complex and diverse diseases have been a major problem, negatively impacting industrial production. Vibriosis is a prolific disease which continuously damages olive flounder aquaculture. A bacterial disease survey was performed from January to June 2017 on 20 olive flounder farms on Jeju Island. A total of 1710 fish were sampled, and bacteria from the external and internal organs of 560 fish were collected. Bacterial strains were identified using 16 s rRNA sequencing. Twenty-seven species and 184 strains of Vibrio were isolated during this survey, and phylogenetic analysis was performed. Bacterial isolates were investigated for the distribution of pathogenic and non-pathogenic species, as well as bacterial presence in tested organs was characterized. V. gigantis and V. scophthalmi were the dominant non-pathogenic and pathogenic strains isolated during this survey, respectively. This study provides data on specific Vibrio spp. isolated from cultured olive flounder in an effort to provide direction for future research and inform aquaculture management practices.

Abstract Stimulator of interferon gene (STING) is induced by various inflammatory agents, such as lipopolysaccharide and microbial pathogens, including virus and bacteria. In this study, we obtained a full-length cDNA of a STING homolog from olive flounder using rapid amplification of cDNA ends PCR technique. The full-length cDNA of Paralichthys olivaceus STING (PoSTING) was 1442 bp in length and contained a 1209-bp open reading frame that translated into 402 amino acids. The theoretical molecular mass of the predicted protein sequence was 45.09 kDa. In the PoSTING protein, three transmembrane domains and the STING superfamily domain were identified as characteristic features. Quantitative real-time PCR revealed that PoSTING expressed in all the tissues analyzed, but showed the highest level in the spleen. Temporal expression analysis examined the significantly upregulated expression of PoSTING mRNA after viral hemorrhagic septicemia virus (VHSV) stimulation. In contrast, no significant changes in the PoSTING expression were detected in Edwardsiella tarda-challenged group compared to the un-injected control. The expression of P. olivaceus type I interferon (PoIFN-I) was also highly upregulated upon VHSV challenge. These results suggest that STING might be involved in the essential immune defense against viral infection together with the activation of IFN-I in olive flounder.