Journal of fish diseases最新文献

Largemouth bass (Micropterus salmoides) is an economically important freshwater fish species in China that is frequently affected by disease outbreaks caused by Aeromonas hydrophila. This study employed an integrated transcriptomic and proteomic approach to investigate the response of the liver in largemouth bass at 72 h post-infection with A. hydrophila via intraperitoneal injection. We identified differentially expressed genes (DEGs; 2136 upregulated and 200 downregulated) and differentially expressed proteins (DEPs; 1683 upregulated and 1817 downregulated). Functional enrichment analysis indicated that the upregulated DEGs/DEPs were significantly involved in immune and inflammatory responses, encompassing pathways, such as TNF signalling, NF-kappa B signalling, complement and coagulation cascades and B cell receptor signalling, while the downregulated DEGs/DEPs were primarily associated with metabolic processes, especially lipid metabolism. Integrated transcriptomic and proteomic analysis identified 420 correlated DEG/DEP pairs, among which 154 were consistently upregulated and 19 consistently downregulated. The upregulation of eight key immune-related DEG/DEPs, including complement components (C1q, C7b, C1r-A), MYD88 and BCL10, was verified by RT-qPCR. Collectively, these findings provide valuable insights into the molecular basis of immune defence against A. hydrophila in largemouth bass and identify potential immune-related biomarkers that may be useful in the development of effective prevention and control strategies.

In vitro bacterial competition assays were performed to evaluate the ecological and antibacterial role of the PirAB^vp toxin complex produced by Vibrio parahaemolyticus Vp278. When co-cultured with V. cholerae, the PirAB^vp producing strains exhibited competitive dominance, eliminating V. cholerae after 19-24 h, whereas non-producing strains coexisted with V. cholerae throughout the incubation period. Quantification and immunodetection analyses confirmed a time-dependent increase in PirB^vp levels during competition, reaching up to 350 ng/mL, while non-producer strains showed no toxin production. Recombinant PirB^vp displayed direct antibacterial activity, causing up to 66.6% reduction in V. cholerae growth and a 5.2-log decrease in CFU at 20 μg/mL. Similarly, A. fischeri exhibited dose-dependent inhibition of bioluminescence and viability, with cell counts reduced from 3.0 × 10⁹ to 5.2 × 10⁵ CFU/mL at 20 μg/mL. The inhibitory effect was markedly reduced when PirB^vp was preincubated with IPTG and the glycoprotein Fetuin, suggesting that the jacalin-like lectin domain, which binds to amino sugars in the bacterial membrane, is essential for its antibacterial action. These findings demonstrate that PirB^vp contributes to interspecific competition and ecological dominance of V. parahaemolyticus through direct, lectin-mediated antibacterial activity.

Cell line misidentification or contamination undermines the reliability of research outcomes. In this study, we verified the species origin of commonly used fish cell lines, with a particular focus on the grass carp ovarian cell line (GCO). RNA-seq data from 146 samples of 11 widely used fish cell lines were collected from public databases (NCBI-SRA) and were mapped to the reference genomes of eight species. GCO RNA-seq samples from three Chinese laboratories showed a mapping rate of 21.64% ± 3.61% to the grass carp genome, but 89.81% ± 4.83% to the fathead minnow genome. Further transcriptomic similarity analysis revealed that these GCO RNA-seq samples clustered more closely with cell lines originating from fathead minnow. Targeted analysis of the COI gene amplification region confirmed that nearly all reads from GCO RNA-seq samples mapped exclusively to the fathead minnow COI amplicon. Collectively, these results indicate that GCO may have undergone cross-contamination or species misidentification. This study raises doubts about the true species origin of the GCO cell line and proposes that COI DNA barcoding can be used to discriminate the species origin of fish cell lines.

In this study, a multiplex RT-PCR assay was developed for the simultaneous detection of three major viral pathogens in salmonids: infectious haematopoietic necrosis virus (IHNV), infectious salmon anaemia virus (ISAV) and salmonid alphavirus (SAV). Primer pairs targeting highly conserved regions of each viral genome were designed based on specificity and broad genotype coverage and first validated through single RT-PCR assays. A β-actin gene was included as an internal control to monitor RNA extraction and cDNA synthesis efficiency. The limit of detection (LoD) in single RT-PCR assays was determined as 1.0 × 10⁰ copies/μL for IHNV, 1.0 × 10¹ copies/μL for ISAV and 1.0 × 10² copies/μL for SAV. The multiplex RT-PCR assay was optimised to detect all three viruses and β-actin simultaneously. Compared with WOAH singleplex PCRs, the assay achieved a LoD_95% of 5.9 × 10² copies/μL across targets. It also showed high specificity, with no cross-reactivity to salmonid bacteria and only the β-actin internal control amplified in fish tissues and cell lines. This assay exhibited high specificity in field samples, with IHNV detected in one of the farms tested. Furthermore, it successfully identified IHNV in infected specimens obtained from controlled infection experiments. Through robustness testing, the assay demonstrated consistent performance across all eight reagent kits, thereby confirming its reproducibility within the overall PCR reagent system. This assay offers a rapid, sensitive and specific diagnostic tool for detecting viral infections in salmonid aquaculture.

Populations of wild brown trout (Salmo trutta) have been in long-term decline across Central Europe, including the Czech Republic, with multiple factors, such as habitat alteration, climate change, predation and emerging diseases, implicated. Among the diseases, proliferative kidney disease (PKD), caused by the myxozoan Tetracapsuloides bryosalmonae, has gained increasing attention. Between 2020 and 2024, we investigated the occurrence of T. bryosalmonae and selected viral pathogens in wild brown trout populations from 34 streams (51 sites) across the three major Czech river basins (Elbe, Morava, Odra). In total, 501 fish were examined using pathology and molecular techniques. While T. bryosalmonae DNA was detected in 54.7% of fish and at 74.5% of localities, with highest prevalence in the Elbe basin (63.5%), gross kidney changes consistent with PKD were observed in just 7.4% of individuals. Significant associations were found between parasite occurrence and mean summer water temperature, with positive sites showing higher water temperatures. No mortalities were observed in the wild populations, though subclinical infections were common. Our findings demonstrate that not only is the PKD agent widespread in Czech trout populations but that temperature plays an important role in parasite dynamics, highlighting increasing risks posed by climate change. We suggest that careful fish stocking strategies will be essential in preventing further spread. The viral pathogens Salmonid novirhabdovirus, Piscine novirhabdovirus and Aquabirnavirus salmonidae were not detected, and piscine orthoreovirus genotype 3 was confirmed in two samples from the Odra basin, showing high sequence identity with previously reported Czech isolates.

Edwardsiella piscicida is a common marine pathogen infecting diverse fish species, causing hemorrhagic septicemia and substantial economic losses to the global aquaculture industry. The hemolysin proteins (EthA) and type III and VI secretion systems (T3SS/T6SS), controlled by the EsrA-EsrB two-component system (TCS), are essential for bacterial colonisation and host invasion. This study demonstrates that the UvrY-BarA TCS negatively regulates the expression of esrB, subsequently activates the expression of hemolysin proteins, and inhibits the secretion of T3SS/T6SS proteins. Importantly, when it detects environmental formate and acetate signals, the UvrY-BarA TCS controls virulence gene expression, affecting the colonisation ability of E. piscicida in turbot. In summary, our findings reveal a novel role for the UvrY-BarA TCS in mediating the regulatory effects of formate and acetate signals on bacterial virulence, providing deeper insights into the pathogenesis of E. piscicida.

Largemouth bass virus (LMBV), a member of the Ranavirus genus within the Iridoviridae family, is a significant pathogen causing high mortality and substantial economic losses in largemouth bass aquaculture. Therefore, developing effective antiviral agents against this viral infection is crucial for the largemouth bass aquaculture industry. Quercetin, a widely utilised flavonoid known for its therapeutic and prophylactic properties, was investigated for its antiviral efficacy against LMBV and underlying mechanisms in this study. The results demonstrated that quercetin significantly inhibited LMBV propagation in vitro and in vivo, and was effective in both prevention and treatment stages. Specifically, quercetin at 6.25 μg/mL suppressed LMBV (major capsid protein) MCP gene transcription by up to 92.09%, with the post-infection treatment achieving 99.33% inhibition in vitro. Mechanistic studies revealed that quercetin reduces LMBV infectivity by disrupting viral internalisation and replication, while also potentiating cellular immune responses mediated by the IFN system. In vivo experiments further confirmed that quercetin treatment reduced viral transcription in the liver, spleen and kidney by 97.49%, 47.45% and 98.42%, respectively, at 48 h post-infection and significantly enhanced the survival rate of infected juveniles. In conclusion, quercetin demonstrated potent antiviral activity against LMBV and might serve as a promising, eco-friendly antiviral agent.

To identify enriched pathogens and characterise the viral community associated with epidemic disease outbreaks in the freshwater mussel Hyriopsis cumingii, we performed metatranscriptomic sequencing combined with VirID-driven RNA-dependent RNA polymerase (RdRP) mining and phylogenetic analysis using hepatopancreas and intestinal samples from six severely infected individuals. Clinical observations were consistent with hallmark features of epidemic outbreaks. The sequencing yielded 86.2 Gb of raw data, of which 97.1% passed quality control, resulting in 77.7 Gb of high-quality clean data. Taxonomic annotation identified 182 viral species, predominantly unclassified viruses (45% Transcripts Per Million, TPM), followed by members of the phyla Lenarviricota (28%) and Uroviricota (17%). Phylogenetic analysis of RdRP sequences revealed 13 viral supergroups, with the Picorna-Calici supergroup showing the highest abundance (26.2% of annotated viruses) and reaching a prevalence of 39.3% in sample HcAV3. Notably, 89.6% of the identified viral RdRPs exhibited less than 70% amino acid identity to known viral sequences, highlighting the presence of extensive "viral dark matter" in this host species. This study establishes the first viral profile associated with epidemic disease in H. cumingii, providing a baseline for further etiological research on this high-mortality aquaculture disease.

Hepcidin is a cysteine-rich peptide that functions both as an antimicrobial peptide and a central regulator of iron homeostasis in vertebrates. Structurally, it consists of an N-terminal signal peptide, a propeptide region and a C-terminal bioactive mature peptide. The canonical mature hepcidin contains eight conserved cysteine residues that form four disulfide bonds, stabilising a characteristic β-sheet hairpin-like structure. In the Antarctic toothfish (Dissostichus mawsoni), which inhabits extremely cold marine environments, a unique hepcidin variant was identified possessing only four cysteine residues, designated dm_4cyshep. To evaluate the antimicrobial function of the dm_4cyshep peptide, the minimum inhibitory concentration (MIC) assays and flow cytometry analyses were performed. The peptide demonstrated binding affinity and bactericidal activity against a range of Gram-positive and Gram-negative bacteria, with distinct MIC values observed for different strains. Furthermore, overexpression of dm_4cyshep in a hepcidin-deficient zebrafish model markedly reduced systemic iron accumulation, indicating a conserved role in iron regulation. These results demonstrate that despite the reduced number of cysteine residues, dm_4cyshep retains both antimicrobial and iron-modulatory functions, underscoring its potential as an immune effector molecule. This study enhances the understanding of the structural and functional diversity of fish hepcidins and identifies a promising candidate for improving disease resistance in aquaculture.

The Chilean salmon farming industry faces major challenges from bacterial diseases, notably tenacibaculosis, which is primarily caused by Tenacibaculum maritimum and T. dicentrarchi. However, T. finnmarkense was recently identified as an emerging pathogen. This bacterium was previously described only in Norway, where two genomovars (finnmarkense and ulcerans) were reported. The present study focuses on the phenotypic and genetic characterisation of isolates recovered from salmonids in Chile to elucidate intraspecific diversity, identify the dominant genomovar, and evaluate pathogenic potential. Thirty-three isolates were analysed through phylogenetic analyses based on the 16S rRNA, fusA, and atpA genes, along with phenotypic characterisation, genotyping by RAPD and ERIC-PCR, and comparative genome analysis. All isolates were confirmed as T. finnmarkense and grouped within the genomovar ulcerans, which was further supported by whole-genome phylogenetic analysis of two representative isolates. Phenotypic characterisation was consistent with the general description of T. finnmarkense from previous studies, although some metabolic variations were observed compared to the homogeneity reported in Norwegian isolates. Genotypic analyses also revealed genetic variability among the presently studied isolates. Pathogenicity was assessed in Atlantic salmon (Salmo salar) through immersion challenge tests under controlled conditions, using isolates obtained from Coho salmon (Oncorhynchus kisutch; SC-T20) and Atlantic salmon (PB-L5). Koch's postulates were confirmed only with the SC-T20 isolate, which caused significant mortality (57.14%) in experimental infections. This study highlights the predominance of genomovar ulcerans in Chilean salmon farming and its varying pathogenic potential. The findings underscore the need to monitor the impacts of tenacibaculosis and to accurately identify the specific etiological agent responsible for skin lesions, particularly in cases of coinfection.