Aeromonas salmonicida is a predominant pathogen that infects fish. The pathogen A. salmonicida subsp. masoucida (ASM) was isolated for the first time from diseased starry flounders (Platichthys stellatus). Our study aimed to isolate, characterize, and investigate the pathogenicity of ASM. Bacterial species were identified using 16s rRNA, gyrB, dnaJ, and vapA analyses. Phylogenetic tree analysis revealed that the ASM strains were clustered with the ASM ATCC strain and other strains isolated from black rockfish. In the antimicrobial susceptibility test, the three ASM strains were considered non-wild types for enrofloxacin, florfenicol, flumequine, oxolinic acid, and oxytetracycline susceptibility. Histopathological analysis revealed bacterial colonies in the secondary lamella and heart, indicating that ASM strains are highly virulent in fish. Comparative analysis and annotation via genome sequencing revealed that, among the 1156 factors, adherence factors were the most prevalent putative virulence determinants, followed by the effector delivery system and adherence. ASM was found to possess 43 type III secretion systems, 22 type VI secretion systems, 11 antimicrobial resistance genes, 3 stress genes, and prophage regions. These findings provide new insights into the virulence profile of ASM and highlight the risk posed by emerging pathogenic strains to starry flounders.

Japanese eel endothelial cells-infecting virus (JEECV) and Anguillid herpesvirus 1 (AnHV) are major pathogens in farmed eels. JEECV causes eel viral endothelial cell necrosis (VECNE), while AnHV leads to symptoms such as head erythema and gill necrosis. Both viruses cause severe mortality alone or in combination, necessitating rapid and early detection of their presences. In this study, we developed a highly efficient duplex quantitative PCR method (r² = 0.999) using hydrolysis probes for the rapid and simultaneous detections of AnHV and JEECV. This new diagnostic method demonstrated a 1.7-fold higher detection rate for AnHV and a 2.5-fold higher detection rate for JEECV than conventional PCR. Quantitative analysis of water and eel tissue samples from aquaculture facilities revealed that the two viruses could be detected in water 1–3 months prior to mortality, enabling their early identification of infections through water testing alone. Notably, the method reliably detected low viral loads (< 1 copy) in both water and tissue samples, facilitating preclinical detection and proactive disease management. This approach reduces the risk of mass mortality and economic losses in eel farming. This study underscores the critical role of advanced molecular diagnostic technologies in enhancing health management in aquaculture.

The Japanese eel endothelial cell-infecting virus (JEECV) and Anguilla herpesvirus (AnHV) are major pathogens in farmed eels. JEECV causes eel viral endothelial cell necrosis (VECNE), while AnHV leads to symptoms such as head erythema and gill necrosis, respectively. Both viruses can result in severe mortality, either independently or in combination, necessitating rapid and early detection of their presence. In this study, we developed a highly efficient duplex quantitative PCR method (R² = 0.999) using TaqMan probes for the rapid and simultaneous detection of AnHV and JEECV. This new diagnostic method demonstrated a 1.7-fold higher detection rate for AnHV and a 2.5-fold higher detection rate for JEECV than those of conventional PCR. Quantitative analysis of water and eel tissue samples from aquaculture facilities revealed that viruses could be detected in water 1–3 months prior to mortality, enabling early identification of infections through water testing alone. Moreover, the method reliably detected low viral loads (<1 copy) in both water and tissue samples, facilitating preclinical detection and proactive disease management. This approach reduces the risk of mass mortality and economic losses in eel farming. These findings underscore the critical role of advanced molecular diagnostic technologies in enhancing health management in aquaculture.

Coastal sea-cage aquaculture environments support dynamic microbial communities, yet their viral components remain poorly characterized. This study profiled DNA viral-like reads recovered from total metagenomes of iron-flocculated seawater concentrates (operationally termed DNA virome) at a rock bream (Oplegnathus fasciatus) sea-cage farm off Geoje Island, Korea, at five time points between July 2024 and May 2025 using long-read Nanopore metagenomics. Multivariate analyses of DNA virome composition indicated temporal patterning in virome composition; samples collected within the cages tended to group together. The DNA virome was dominated overall by bacteriophages of abundant marine bacteria and by viruses infecting picoeukaryotic phytoplankton, and it showed marked temporal turnover during the investigated year: Micromonas-associated viruses became prominent in December, whereas Ostreococcus- and Bathycoccus-associated viruses increased during seasonal rewarming in May, suggesting that environmentally driven changes in microbial host communities may contribute to temporal virome turnover. Pelagibacter phage abundance was positively correlated with that of its potential host Pelagibacter ubique, and both Synechococcus T7-like virus and Synechococcus virus were positively correlated with Parasynechococcus marenigrum, associations that are consistent with, but do not demonstrate, taxon-specific virus–host relationships. Nanopore reads assigned to infectious spleen and kidney necrosis virus, now classified within the species Megalocytivirus pagrus 1, were recovered from samples collected in September 2024 (12,960 bp) and May 2025 (340 bp). Average nucleotide identity analysis indicated that the seawater-derived sequence was most closely related to red seabream iridovirus subtype II, the subtype predominant in Korean cultured marine fish. The seawater virome results and the bioinformatic pipelines developed in this study together may provide a non-invasive approach for tracking temporal virome dynamics and for environmental screening of viral pathogen DNA in sea-cage aquaculture.

Edwardsiella piscicida causes severe edwardsiellosis in farmed finfish, particularly Paralichthys olivaceus. Lacking clinical breakpoints, epidemiological cut-off values (ECVs) are crucial for distinguishing wild-type (WT) populations from non-wild-type (NWT) strains. We evaluated 68 E. piscicida isolates from South Korea (2022–2025) across six aquaculture antimicrobials. Tentative ECVs derived via Normalized Resistance Interpretation were established at 8/4 µg/mL for amoxicillin/clavulanic acid (64.7% WT), 2 µg/mL for florfenicol (92.6% WT), 0.5 µg/mL for oxytetracycline (38.2% WT), and 0.25/4.75 µg/mL for trimethoprim/sulfadiazine (51.5% WT). Due to pronounced bimodal distributions and mathematical fitting artifacts, objective ECVs could not be reliably determined for enrofloxacin and flumequine via unimodal modeling. ARG screening revealed high prevalence of bla_TEM (54.4%) and qnrS (52.9%). Tetracycline resistance genes were harbored by 60.3% of isolates (tetD, 45.6%; tetA, 7.4%; tetB, 5.9%; tetB&tetD, 1.5%; tetE, 0%), showing strong statistical concordance with the oxytetracycline NWT phenotype. Furthermore, floR (7.4%) and sul2 (14.7%) strongly correlated with florfenicol and potentiated sulfonamide NWT phenotypes, respectively. Presumptive multidrug resistance (pMDR) remained low at 4.4% (n = 3). These findings establish empirical baseline ECVs for East Asian aquaculture, supporting responsible antimicrobial stewardship and diagnostic harmonization.

Emaciation disease, primarily caused by the myxosporean parasite Enteromyxum leei, poses a significant economic threat to the olive flounder (Paralichthys olivaceus) aquaculture industry. Research on E. leei has been severely restricted by its status as an unculturable obligate parasite and its extreme sensitivity to cryopreservation. In this study, we developed a standardized in vitro culture system that achieved stable maintenance of E. leei for over 12 months for the first time. To overcome the challenges of primary isolation, a physical aggregation technique was implemented, which effectively concentrated the parasites while reducing bacterial contamination by 99.1%, as validated by qPCR. The established culture, maintained in minimum essential medium supplemented with 30% FBS at 25 °C, exhibited a consistent 20-day asexual proliferative cycle consisting of six distinct developmental stages. Morphological stability and genetic identity were rigorously monitored, showing no phenotypic drift or genetic divergence throughout the 12-month period. Our results demonstrate that the entire proliferative phase of E. leei can be recapitulated in vitro without host intervention. Given that cryopreservation remains non-viable, this continuous culture system provides a critical, unique platform for high-throughput drug screening and myxozoan developmental biology, offering a robust tool to mitigate emaciation syndrome in marine fish farms.