This study provides a detailed pathological characterization of skeletal neoplasms in the Indian oil sardine (Sardinella longiceps) from the southeast coast of India. A total of 237,370 landed fish examined at Parangipettai landing center between 2022 and 2024 formed the surveillance population, among which 721 tumour cases were documented. The overall apparent prevalence of grossly detected tumour cases among landed fish examined was 0.30%, with seasonal values ranging from 0.02% during monsoon to 0.72% during summer. Among the tumour-bearing fish, 136 lesions showed skeletal involvement and constituted the focus of the present study. Grossly, these lesions appeared as firm, irregular bony swellings involving predominantly cranial and fin-associated skeletal regions, with lesion diameters ranging from 1.5 to 14.5 mm. Radiographic examination of skeletal cases demonstrated well-defined radiopaque lesions consistent with mineralized skeletal proliferation. Of the 136 skeletal lesions, 112 were suitable for histopathological examination. Histologically, 97 lesions (86.61%) were classified as compact (ivory) osteoma, 12 (10.71%) as cancellous osteoma, and 3 (2.68%) as chondrosarcoma. Osteomas were characterized by well-differentiated mature lamellar bone without cytological atypia or malignant features, whereas chondrosarcomas exhibited increased cellularity, nuclear atypia, a lobulated chondroid matrix, and focal infiltrative growth within the available sections. Because complete tumour margins and surrounding tissues were not available in all chondrosarcoma cases, the extent of invasion could not be comprehensively assessed. These findings provide baseline pathological evidence of skeletal neoplasia in S. longiceps and support continued pathological surveillance and further investigation into the biological and environmental factors potentially associated with tumour development in wild marine fish.
Spring viremia of carp virus (SVCV), that can infect cyprinoid and koi causing an acute hemorrhagic symptom, with a feature of widespread epidemic, which poses a severe threat to the global aquaculture industry. Given the absence of specific effective medicine or vaccines, the early, accurate diagnosis is critical in the disease prevention. Many SVCV detection methods have been established, but most require specialized equipment and skilled operators, which limits their field application. In this study, we established a recombinase-mediated isothermal amplification (RAA) in conjunction with the CRISPR/Cas12a system targeting the screened conserved region of the G gene in the genome to identify SVCV. The optimal combination of the primer pairs and crRNA was screened via the fluorescence intensity. In addition, the test exhibited high sensitivity and specificity, which could detect at the limit of 1.243 × 101 copies/μL and show no cross-reactivity with other common freshwater fish pathogens. Furthermore, the result could be visible by excitation of fluorophores by blue light at 39°C within 30 min. The integrated method combines the advantages of RAA and CRISPR-based signal transduction, offering a diagnosis project in resource-limited aquaculture environments and a field-applicable early warning of SVCV.
Tenacibaculosis, caused by Tenacibaculum dicentrarchi, is an emerging bacterial disease affecting salmonid aquaculture worldwide; however, host-pathogen interactions and the immune response of Atlantic salmon (Salmo salar) remain poorly understood. This study evaluated the expression of 20 immune-related genes in Atlantic salmon experimentally infected by immersion with T. dicentrarchi and in uninfected controls under identical conditions. The 10-day challenge trial included sampling every two days (skin/muscle, gills, spleen and head kidney) for gene expression analyses. Koch's postulates were fulfilled in the challenge with the T. dicentrarchi TdCh06, which induced 40% cumulative mortality; deaths began at 2 days post-infection and continued until Day 7. Of the 20 genes analysed by RT-qPCR, seven (MHC-I, MHC-II, Arg-1, IL-1β, IL-2, IL-10 and IFN-α1) were consistently detected across tissues and used for comparative analysis. The results show that T. dicentrarchi induces a rapid but heterogeneous immune response in Atlantic salmon: the spleen exhibits an early, transient systemic response; the head kidney shows delayed but strong transcriptional activation; skin/muscle displays early local activation with sustained IL-1β and IFN-α1 expression; and gills show early mucosal activation with persistent IL-1β induction. Overall, infection triggers a tissue-specific response involving inflammatory, antigen-presentation, interferon-associated and macrophage-related pathways, suggesting sustained immune stimulation rather than effective bacterial clearance. These findings shed light on the host-pathogen interaction during tenacibaculosis and identify key immunological pathways involved in disease progression. Notably, the genes IL-1β, MHC-I, MHC-II, IFN-α1, IL-2, IL-10 and Arg-1 are proposed as potential transcriptional markers of infection, offering valuable tools to characterise host responses. Importantly, these markers may also be useful for evaluating vaccine efficacy against T. dicentrarchi in future studies.
In teleosts, mesenchymal tumours like fibromas are frequently observed, whereas their malignant counterparts, fibrosarcomas, occur only sporadically. In our study, an adult female greater amberjack (Seriola dumerili) reared at the Aquarium of Genoa developed a slow-growing mass protruding from the ventral right side of the head. The tissue was firm and white greyish with localized cranial haemorrhaging on the external surface. Cut-section examination showed a homogeneous, dry, whitish and richly vascularized tissue layout. Tissue samples from the mass were stained with Masson's Trichrome, Alcian Blue-PAS (pH 2.5), and Toluidine blue for differential diagnosis. Microscopically, it was characterized by alternating collagenous spindle-cell septa and hypocellular metachromatic myxoid areas positive for acidic mucopolysaccharides and glycosaminoglycans. The absence of cellular atypia, mitotic activity and necrosis confirms the diagnosis of fibromyxoma. Documenting rare tumours in aquarium fish expands comparative pathology literature and highlights public aquaria as valuable research platforms for long-term health monitoring under controlled husbandry conditions.
Poikilothermic organisms like fish are key economic drivers in aquaculture and the ornamental fish trade. Climate change impacts the health of fish because poikilothermic hosts experience the same temperature as their environment, making temperature a critical determinant of parasite infection dynamics. Pseudocapillaria tomentosa, a capillarid nematode infecting approximately 20% of zebrafish research facilities, provides an excellent model for investigating temperature effects on nematode development because zebrafish can tolerate a wide temperature range. We examined how water temperature influences egg development and worm maturation across the zebrafish's tolerable thermal range (13°C-35°C). Eggs held at 4°C or 13°C failed to larvate but resumed development when returned to 28°C. Larvation increased at 35°C, though infectivity was reduced in larval zebrafish. In adult zebrafish, cold water (13°C) slowed worm development for up to 3 months; only immature larvae were observed even at 102 days post-exposure. Returning these fish to 28°C triggered rapid maturation, with gravid females appearing within 1 month. Conversely, fish held at 35°C had reduced worm burdens and arrested maturation compared to 28°C controls, with development resuming after temperature reduction. These findings align with field observations of seasonal P. tomentosa dynamics in temperate climates.
This research was designed to explore the immunogenicity of the yersiniosis vaccine and the effect of Ulvan as a natural adjuvant on immune, antioxidant and survival parameters in Oncorhynchus mykiss. A total of 450 O. mykiss (mean body weight 52.90 ± 2.02 g) were allocated to three experimental treatments, each with three replicate tanks. Treatments were: (1) injectable inactivated Yersinia ruckeri vaccine (Vac), (2) the same vaccine formulated with Ulvan as an adjuvant (Vac + U) and (3) an unvaccinated control group (Ctrl) receiving phosphate-buffered saline (PBS). White blood cells were significantly elevated in all vaccine-treated groups compared with the Ctrl group on days 30 and 60 (p < 0.0001). In contrast, no significant treatment effects were observed for red blood cells (p = 0.1078), haemoglobin (p = 0.2646), or haematocrit (p = 0.2352) at either sampling time. There were no significant treatment impacts on total protein (p = 0.3330) or serum globulin (p = 0.6521) at the 30- or 60-day samplings. Conversely, serum albumin was notably elevated in the Vac + U group relative to the Ctrl at both 30 and 60 days (p = 0.0244). Vaccination induced pronounced increases in both serum immune (lysozyme, total immunoglobulin, NBT) and antioxidant (catalase, superoxide dismutase, glutathione peroxidase) parameters at 30 and 60 days post-immunization compared with Ctrls (p < 0.05). Antibody titers against yersiniosis were significantly higher in all vaccine-treated groups than in the Ctrl group at 30 and 60 days post-immunization (p < 0.0001). Survival rate after a two-week Y. ruckeri challenge was 73.33% (Vac), 86.66% (Vac + U) and 16.66% (Ctrl); survival in the vaccine-treated groups was notably higher than in the Ctrl group (p < 0.0001). The results indicated that both the vaccine and the Vac + U formulation enhanced biochemical markers, immune-antioxidant responses, serum antibody titers and survival following challenge with Y. ruckeri infection. Among the tested groups, the Vac + U demonstrated the greatest effectiveness, likely due to the presence of Ulvan as an adjuvant.
Non-medicinal approaches for controlling sea lice (Lepeophtheirus salmonis) infestations, such as freshwater (FW) treatments, have been increasingly implemented to support more sustainable parasite control in aquaculture. This study evaluated the efficacy of FW treatment against sea lice under controlled experimental conditions and assessed the potential development of resistance across multiple generations. Sea lice populations were maintained over five consecutive generations, and FW efficacy was evaluated by quantifying lice abundance on Atlantic salmon before and after treatment. Resistance assessment involved comparative bioassays using copepodid and adult life stages from two distinct populations, one exposed to FW and the other a saltwater (SW) control group. Our results demonstrated that a 4 h FW treatment significantly reduced total lice abundance, while an extended 18-day exposure resulted in complete detachment of all lice from salmon. Importantly, bioassay comparisons revealed no evidence of resistance development, as both FW-exposed and SW-control populations maintained similar susceptibility across generations. The absence of measurable changes in salinity tolerance over multiple generations suggests that FW protocols may remain a viable and sustainable management strategy. However, the potential for genetic or epigenetic variation in FW tolerance among lice populations warrants further investigation to better understand long-term implications for treatment efficacy.
Infectious diseases among fish farms present significant challenges to fish health and production. Horizontal transmission serves as one of the main mechanisms driving outbreaks in these settings. The dynamics of horizontal transmission are influenced by pathogen properties, host characteristics and environmental factors. Additionally, the implementation of epidemiological interventions can further shape the patterns and consequences of infectious disease spread. However, there is a paucity of studies assessing how specific epidemiological measures, such as timely disease surveillance and strategic depopulation, influence transmission and outbreak outcomes. This study utilized a spatiotemporal hybrid model to assess pathogen dissemination following an infectious disease event in Tongyeong, a prominent area for floating net cage aquaculture in Korea. We investigated how variations in sampling intensity affected the timing of disease detection and analysed the impact of two control strategies (immediate culling upon detection versus no culling) on the progression of disease and fish mortality. Findings revealed significant differences in disease detection timing and probability based on the surveillance sampling size. Early detection of disease was achieved when 100 fish were sampled, as opposed to a delayed response when only 5 fish were tested. Furthermore, the conditions under which surveillance occurred had a notable impact on both the spatial spread and the success of depopulation measures. Generally, prompt detection followed by immediate depopulation led to a decrease in disease-related mortality over a wider area. However, this method also necessitated culling a substantially greater number of fish. Although the impact of these interventions may be influenced by geographical and environmental variables, our results suggest that the utility of depopulation strategies is closely linked to the efficacy of surveillance efforts and the epidemiological profile of the infection, such as its associated mortality rate and transmissibility.
In 2023, all but one Swedish restocking farm rearing Baltic salmon and sea trout fry experienced massive mortality caused by lipoid liver disease (LLD). The struck farms used the same feed, and this was suspected to be the cause of the outbreak. We performed two parallel feed trials in three salmon populations as well as fatty acid and antioxidant analyses of feed and fish to investigate the causal relationship between the feed and the LLD mortality. Significant mortality was induced in one population and LLD histopathology was induced in all three populations in groups fed the suspected feed (test feed) but not in the control groups, linking the test feed to the outbreak in 2023. Fatty acid analysis revealed a difference in bound fatty acid composition between the suspected feed and the control feeds, including higher amounts of erucic acid. Erucic acid has negative health effects for fish by altering lipid metabolism. It is present in for example, capelin, a main ingredient in this feed. The test feed also contained lower levels of several antioxidants, especially vitamin A. Antioxidative status of the fish could not be fully analysed but results indicated a difference in antioxidative status between the populations.
Singapore grouper iridovirus (SGIV) is a major viral pathogen threatening grouper aquaculture. The lack of specific antiviral drugs underscores the urgent need to develop safe and effective control strategies. This study demonstrates that tannic acid (TA) exhibits significant anti-SGIV activity in vitro. In grouper spleen (GS) cells, TA inhibited SGIV replication in a concentration-dependent manner. Time-of-addition assays confirmed its antiviral efficacy when applied either before or after SGIV infection. Mechanistically, TA directly interfered with SGIV particle infectivity, viral adsorption, entry and intracellular replication. Network pharmacology analysis suggested TA's targets are associated with host cellular antioxidant pathways, among which Peroxiredoxin 6 (PRDX6) was identified as a potential target of TA. Further experiments confirmed that TA upregulated the Nrf2 antioxidant pathway and suppressed the expression of pro-inflammatory cytokines (IL-1β, IL-8). Notably, TA enhanced the expression of PRDX6, and overexpression of PRDX6 significantly inhibited SGIV replication, indicating that PRDX6 contributes to the antiviral effect of TA. In vivo, TA treatment not only restored PRDX6 expression in infected groupers but also significantly reduced viral load in liver and spleen following SGIV infection. These findings demonstrate that TA inhibits SGIV through direct viral interference and PRDX6-mediated antioxidant modulation, highlighting its potential as a natural therapeutic agent against SGIV.
Hyperostosis is a non-pathological condition widely reported in marine teleosts, yet its developmental dynamics and underlying drivers remain poorly understood. This study investigates the ontogenetic progression and anatomical distribution of hyperostosis in Caranx hippos based on a growth series of 39 specimens collected along the Brazilian coast. Individuals were categorised into two size classes (class 1 < 400 mm and class 2 ≥ 400 mm), and skeletal alterations were assessed through radiographic imaging, morphometric analyses and histology. Hyperostosis was detected in 87.2% of specimens and affected nine skeletal regions, including dorsal and anal pterygiophores, cleithrum, ribs, pelvic bones, neural and hemal spines and for the first dorsal fin pterygiophore, preopercle and opercle. The dorsal pterygiophores were the earliest and most consistently affected elements, with detectable changes in individuals as small as 184 mm TL, suggesting an early onset of the condition. Other regions exhibited a size-dependent pattern, with hyperostosis appearing sequentially and increasing in frequency and extent with growth. Morphological variation among skeletal elements indicates region-specific expression, with pronounced differences in thickness, width and mass between hyperostotic and non-hyperostotic bones. Comparisons with previous studies confirm a species-specific pattern of hyperostotic distribution in C. hippos, with minor variation in the onset and extent of affected regions. The observed ontogenetic progression supports the hypothesis that hyperostosis develops sequentially across skeletal elements rather than simultaneously. These findings, combined with ecological and biomechanical considerations, suggest that hyperostosis results from multiple interacting factors, including growth-related mechanical loading and environmental variability. Overall, this study provides a comprehensive characterisation of hyperostotic development in C. hippos, highlighting the importance of ontogenetic approaches for understanding the biological and functional significance of this condition in marine fishes.
Aeromonas hydrophila is a major zoonotic bacterial pathogen responsible for haemorrhagic septicemia, substantial economic losses in aquaculture and increasing antimicrobial resistance among field isolates. It is also one of the main bacterial species associated with mortality outbreaks in pirapitinga, Piaractus brachypomus, culture in the Amazon region of Latin America. Therefore, this study evaluated the immunogenicity and protective efficacy of the first autogenous vaccine against A. hydrophila developed for P. brachypomus, formulated with or without a Montanide adjuvant. A total of 150 fish were vaccinated by intraperitoneal injection with a formalin-inactivated homologous bacterin and, at 35 days post-vaccination, challenged with a homologous strain of A. hydrophila at the median lethal dose (LD50). Following bacterial challenge, biological samples were collected at 24 h post-challenge, and survival was monitored for 15 days (50-day trial period). The vaccinated groups showed enhanced immune responses, characterized by increased ROS levels, lysozyme activity, agglutinating antibody titers and IgM levels, particularly in the fish vaccinated with the adjuvant. Haematological evaluation revealed lymphocytosis and, morphologically, lower lesion severity and preservation of renal architecture in vaccinated groups. In contrast, infected non-vaccinated fish exhibited leukopenia, tubular disorganization, intense mononuclear inflammatory infiltrates, glomerular alterations and lesions suggestive of tissue necrosis. These findings were reflected in enhanced protection following the bacterial challenge, with relative percent survival (RPS) values of 76.19% for fish vaccinated without the adjuvant and 85.71% for those vaccinated with the adjuvant. Overall, the autogenous vaccine was safe, immunogenic and provided effective protection against Aeromonas hydrophila infection in Piaractus brachypomus.
Decapod iridovirus 1 (DIV1) is a highly lethal pathogen that infects decapod crustaceans including Litopenaeus vannamei, causing mass mortality in cultured shrimp and severe economic losses worldwide. The ATPase gene is a highly conserved region within the DIV1 genome, plays a critical role in viral replication and represents an ideal target for molecular diagnostic development. In this study, we established a rapid, sensitive and field-adaptable detection platform for DIV1 by integrating recombinase polymerase amplification (RPA) with the CRISPR/Cas12a system. RPA enables efficient isothermal amplification of target nucleic acids, achieving exponential enrichment of the target nucleic acids and exerting the function of signal amplification. While the CRISPR/Cas12a system upon crRNA-guided specific recognition of the amplicon, triggers robust trans-cleavage activity against reporter probes for signal generation and readout. After systematic optimization, the RPA reaction was performed at 38°C for 10 min and the CRISPR-Cas12a reaction was conducted at 37°C for 20 min. The integrated two-step workflow completed detection within 40 min, with a limit of detection of 2.3 × 101 copies/μL. Specificity evaluation confirmed that the RPA-CRISPR/Cas12a assay exclusively recognised DIV1 without cross-reaction with other major shrimp pathogens. Further validation using clinical shrimp samples demonstrated stable and reliable performance, supporting its practical utility in aquaculture settings. In conclusion, the established CRISPR/Cas12a-based detection platform provides a robust technical tool for early warning and on-site rapid screening of DIV1, facilitating timely disease control and risk management in shrimp farming.
Examination of round sardinella Sardinella aurita Valenciennes, 1847 (Clupeidae) from the Gulf of Tunis (Tunisia) revealed the first local and Mediterranean record of Anthobothrium sp. 1 sensu Jensen & Bullard, 2010 (Tetraphyllidea incertae sedis, Anthobothrium). Nodular lesions containing multiple larvae were detected at the proximal end of the pyloric caeca, and free plerocercoids were observed in the intestinal lumen and stomach. The newly generated LSU rDNA sequences shared 100% nucleotide identity with adult Anthobothrium sp. 1B from the blacktip shark Carcharhinus limbatus Valenciennes, 1839 and larvae from the largehead hairtail Trichiurus lepturus L., both collected from the Gulf of Mexico (Florida and Mississippi, USA). Furthermore, they exhibited a minor 2.1% sequence divergence from related lineages within the same complex found in the Atlantic sharpnose shark Rhizoprionodon terranovae Richardson, 1836 and the hardhead sea catfish Ariopsis felis (Linnaeus, 1766) from the same Gulf of Mexico geolocations. COI sequences showed low variability, with p-distances ranging from 0.18% to 0.92% (corresponding to 1-5 nucleotide differences across 544 base pairs), consistent with the presence of a single species. Scanning electron microscopy of the cestode larva revealed aristate gladiate spinitriches and capilliform filitriches within the acetabula, and capilliform filitriches on the scolex peduncle. Histopathological analysis of nodular lesions showed clusters of multiple larvae, predominantly located within the caecal lumen. Host tissues exhibited localised mucosal alterations, characterised by elements of mucosal atrophy, villus compression and epithelial degeneration. These changes were primarily the result of mechanical pressure exerted by the larvae within the caecal lumen. A focal accumulation of mucous cells (presumptive goblet cells) was observed in affected areas. While primary structural alterations were confined to the mucosa, inflammatory infiltration within the submucosa appeared minimal. Despite this typical localised tissue response, S. aurita appears to be a regular second intermediate host, whereas other clupeids may also act as second intermediate hosts and clupeid-feeding teleosts as paratenic hosts. This finding extends the distribution of Anthobothrium sp. 1 from the western Atlantic to the western Mediterranean Sea, where the definitive host remains unknown.
Viral hemorrhagic septicemia virus (VHSV) is a highly pathogenic rhabdovirus that primarily spreads through horizontal transmission. Characterizing viral shedding dynamics is therefore critical for understanding its epidemiology and improving disease control strategies in aquaculture. In this study, the median lethal dose (LD₅₀) of VHSV in olive flounder (Paralichthys olivaceus) was determined to be 103.7TCID50/100 μL. We subsequently investigated viral shedding patterns at both individual and group levels under different initial infection ratios. Two distinct peaks in viral shedding were consistently observed at 6 to 8 and 10 to 12 days post-challenge (dpc). In group tanks, the first peak was more pronounced, whereas in individual tanks, the second peak predominated, suggesting that the timing of mortality strongly influences shedding dynamics. Notably, moribund or deceased fish released substantially higher viral loads, with some individuals reaching 106 copies/400 mL on day 10, whereas surviving fish maintained relatively stable and much lower levels (< 104 copies/400 mL) throughout the study. Among different infection ratios, cumulative mortality was highest in the 50% infection group (approximately 60% ~ 70% by 20 dpc). However, the 30% infection group exhibited the greatest viral shedding, indicating that viral release does not necessarily scale with infection prevalence. Instead, transmission dynamics appear to be disproportionately driven by a limited number of high-shedding individuals capable of sustaining infection at the population level. Collectively, these findings demonstrate that viral shedding is concentrated in severely affected individuals and that individual shedding capacity, rather than infection prevalence alone, plays a dominant role in VHSV transmission. Early intervention targeting high-shedding individuals may therefore be critical for effective disease control in aquaculture systems.
Aeromonas hydrophila is a major pathogen of Motile Aeromonad Septicemia (MAS) in Nile tilapia, with pathogenesis and immunity poorly understood. In early 2026, a > 70% mortality outbreak occurred at a Nanning tilapia farm. The dominant strain NN0116 was identified as A. hydrophila by 16S rRNA and whole-genome ANI analysis. It exhibited high virulence (LD50 = 5.2 × 103 CFU/fish). Its 5.37 Mb genome encodes 5065 proteins and contains T2SS, T3SS, T6SS, and pore-forming toxins. Virulence and antimicrobial resistance genes co-localized on genomic islands GI2 and GI7. The strain was resistant to 12 antibiotic classes but susceptible to third-generation cephalosporins and fluoroquinolones. Head kidney transcriptomics at 24 h identified 1066 differentially expressed genes, including 61 immunoglobulin (Ig) genes. KEGG enrichment revealed 43 significant pathways; the top ten were driven by 26 Ig genes. Network analysis of the top 20 pathways identified B cell receptor signalling (ko04662) and COVID-19 (ko05171) as central nodes, alongside 10 secondary core pathways, all Ig-driven. Integration of bacterial genomic and host transcriptomic data characterizes hypervirulent A. hydrophila NN0116 and demonstrates that Ig-mediated B cell responses are central to host defence, offering targets for vaccines and therapeutics.
This study reports the optimization of a cost-effective one-step PCR assay for the detection of White Spot Syndrome Virus (WSSV) in wild shrimp populations, providing an accessible alternative to expensive commercial nested PCR kits in Mozambique. Primer sets targeting VP19, VP24, VP26 and VP28 were evaluated, with the VP28_2 primer set demonstrating the highest sensitivity and no cross-reactivity with Infectious Hypodermal and Haematopoietic Necrosis Virus (IHHNV) or Decapod Iridescent Virus 1 (DIV1). Cost analysis showed that the optimized assay is approximately three times less expensive than commercial nested PCR (USD 5.57 vs. USD 15.75 per reaction). The assay was applied to 300 wild shrimp specimens (Penaeus indicus and Penaeus monodon) collected during a cross-sectional survey conducted in the rainy season (October-November 2022) at 15 landing sites across six provinces. WSSV was detected in 60.7% of samples, with significant regional variation: 78.0% in the South, 58.0% in the Centre and 48.1% in the North. Although the findings indicate widespread WSSV occurrence in wild shrimp populations, the cross-sectional design, single-season sampling and sample size limit inference on temporal trends. Nevertheless, the optimized assay provides a practical and affordable tool for routine WSSV surveillance under existing resource constraints.
Monitoring antimicrobial resistance (AMR) in fish pathogens is essential for supporting sustainable aquaculture. In this study, susceptibility testing and genome sequencing were used to characterize the AMR profile of seven Flavobacterium psychrophilum originating from Danish rainbow trout farms (2019-2022). To establish epidemiological context and baseline virulence, isolates were also subjected to genome-based Multi-Locus Sequence Typing (MLST), serotyping and, for a subset, in vivo LD50 testing. The analysis revealed changes in the susceptibility to antimicrobials used in Danish aquaculture. Most importantly, reduced susceptibility to florfenicol was observed in two isolates originating from a farm experiencing ineffective treatment (disc diffusion diameter of 7-8 mm, MIC > 16 μg mL-1). These isolates (serotype Th) belonged to a clonal complex previously unreported in the Nordic countries (CC-ST90), and the genome analysis revealed a floR gene (413 aa) lying in a large genomic island, putatively conferring this property. The high diversity of the floR GC content (57.4%) from the floR-harbouring genomes (32.7%-32.8%) suggested its recent acquisition. Since florfenicol is the drug of choice for this infection in Denmark, these observations are highly relevant. However, in vitro lab and in vivo animal experiments are needed to confirm the activity of floR and support the clinical relevance of these findings. Also, a larger bacterial panel is needed to evaluate potential trends across farms/regions.
ABSTRACT Tenacibaculosis is an important disease of marine fishes caused by bacteria of the genus Tenacibaculum . Pathogenic Tenacibaculum species or serotypes, including Tenacibaculum maritimum , T. dicentrarchi , T. finnmarkense and T. soleae , have been isolated from farmed Chinook salmon ( Oncorhynchus tshawytscha ) during disease outbreaks in Aotearoa‐New Zealand. We investigated the cytotoxic effects of extracellular products (ECP) produced by these bacteria on the Chinook salmon gill epithelial cell line, OTgill1PFR. ECP from T. maritimum had the greatest cytotoxic effect on confluency and metabolism of OTgill1PFR following 24‐h exposure. ECP of T. dicentrarchi also caused significant but more moderate effects, while T. finnmarkense and T. soleae had minimal effects on the cell confluency or metabolism. These in vitro results were consistent with whole‐animal challenge findings, where T. maritimum caused the most severe clinical disease and mortality. A finding of note was that OTgill1PFR exposure to ECP from all Tenacibaculum species resulted in minimal detectable membrane disruption as assessed by end‐point assay and continuous live‐cell imaging, suggesting the observed cytotoxic effects on confluency and metabolic activity may occur via mechanisms not primarily associated with acute membrane lysis. The OTgill1PFR cell model was sensitive enough to detect both interspecific and intraspecific differences in Tenacibaculum ECP activity, proving its utility as an in vitro platform for investigating host‐pathogen interactions and virulence mechanisms in salmonid tenacibaculosis.
