Diagnosis, isolation and description of a novel amnoonvirus recovered from diseased fancy guppies, Poecilia reticulata.

The guppy, Poecilia reticulata, is one of the most common cultured ornamental fish species, and a popular pet fish highly desired by hobbyists worldwide due to its availability of many brilliantly coloured fish of many varieties. The susceptibility of guppies to diseases presents a remarkable concern for both breeders and hobbyists. In this study, we report the emergence of disease in fancy guppies caused by a previously uncharacterized virus in the USA. This virus was isolated from moribund guppies in two separate outbreaks in California and Alabama, from December 2021 to June 2023. The infected guppies presented with acute morbidity and mortality shortly after shipping, displaying nonspecific clinical signs and gross changes including lethargy, anorexia, swimming at the water surface, gill pallor, mild to moderate coelomic distension and occasional skin lesions including protruding scales, skin ulcers and hyperaemia. Histological changes in affected fish were mild and nonspecific; however, liver and testes from moribund fish were positive for Tilapia lake virus (TiLV), the single described member in the family Amnoonviridae, using immunohistochemistry and in situ hybridization, although the latter was weak. A virus was successfully recovered following tissue inoculation on epithelioma papulosum cyprini and snakehead fish cell lines. Whole genome sequencing and phylogenetic analyses revealed nucleotide and amino acid homologies from 78.3%-91.2%, and 78.2%-97.7%, respectively, when comparing the guppy virus genomes to TiLV isolates. Based on the criteria outlined herein, we propose the classification of this new virus, fancy tailed guppy virus (FTGV), as a member of the family Amnoonviridae, with the name Tilapinevirus poikilos (from the Greek 'poikilos', meaning of many colours; various sorts, akin to 'poecilia').

Confirmatory test of active IHHNV infection in shrimp by immunohistochemistry and IHHNV-LongAmp PCR.

The presence of endogenous viral elements (EVE) in the penaeid shrimp genome has been recently reported and suggested to be involved in the host recognition of viral invaders. Our previous report of a search for EVE of infectious hypodermal and haematopoietic necrosis virus (IHHNV-EVE) in the Thai Penaeus monodon whole genome sequence project (GenBank accession no. JABERT000000000) confirmed the presence of three clusters of EVE derived from IHHNV in the shrimp genome. This study aimed to compare an immunohistochemistry method (IHC) and a PCR method to detect infectious IHHNV infection in shrimp. First, specimens collected from farms were checked for IHHNV using three PCR methods; two methods were recommended by WOAH (309 and 389 methods), and a newly established long-range PCR for IHHNV (IHHNV-LA PCR) targeting almost the whole genome (>90%) of IHHNV. Among 29 specimens tested, 24 specimens were positive for WOAH methods (at least one method). Among 24 WOAH-positive specimens (WOAH+), there were 18 specimens with positive IHHNV-LA PCR method (WOAH+/LA+), six specimens with negative IHHNV-LA PCR method (WOAH+/LA-). Six specimens were negative for all methods (WOAH-/LA-). The positive signals detected by IHC method were found only in the specimens with WOAH+/LA+. The results suggest that the WOAH+/LA- specimens were not infected with IHHNV, and the positive WOAH method might result from the EVE-IHHNV. The study recommends combining the IHHNV-LA PCR method and IHC with positive PCR results from WOAH's recommended methods to confirm IHHNV infection.

Diagnosis of piscine francisellosis in Largemouth Bass from a public display exhibit in north-central Florida, USA.

OBJECTIVE: The Largemouth Bass Micropterus salmoides is an important freshwater fish that is native to the southeastern United States and is cultured for conservation, food, and for the sports fishing industry. Francisella orientalis is a globally distributed bacterial pathogen of warmwater fish species and is associated with granulomatous inflammation and high mortalities. Outbreaks of piscine francisellosis in the United States have been reported in only a few fish species. This study describes three case presentations of francisellosis in Largemouth Bass from a public display system in north-central Florida. Additionally, laboratory-controlled immersion challenges using an F. orientalis isolate from tilapia Oreochromis spp. evaluate susceptibility of Largemouth Bass fingerlings to F. orientalis infection and mortality through this exposure route. METHODS: Necropsy, histologic examination, immunohistochemistry, bacterial recovery and culture, and quantitative polymerase chain reaction were used as diagnostic tools to evaluate both the affected display fish and the immersion-challenged fingerlings. RESULT: Although the display fish and immersion-challenged fingerlings presented with nonspecific clinical signs, gross and histological changes were indicative of granulomatous disease. Immunohistochemical and molecular testing methods confirmed F. orientalis infection in affected fish. CONCLUSION: The three case presentations described here mark the first reporting of naturally occurring piscine francisellosis in Largemouth Bass that were held in a public display exhibit. Additionally, causality was proven in the Largemouth Bass fingerlings through the immersion challenges. These findings demonstrate susceptibility through immersion-based exposure and assert that francisellosis should be considered among the list of differential diagnoses for Largemouth Bass with granulomatous disease.

Early detection of Pseudocapillaria tomentosa by qPCR in four lines of zebrafish, Danio rerio (Hamilton 1882).

The intestinal nematode Pseudocapillaria tomentosa in zebrafish (Danio rerio) causes profound intestinal lesions, emaciation and death and is a promoter of a common intestinal cancer in zebrafish. This nematode has been detected in zebrafish from about 15% of the laboratories. Adult worms are readily detected about 3 weeks after exposure by either histology or wet mount preparations of the intestine, and larval worms are inconsistently observed in fish before this time. A quantitative PCR (qPCR) test was recently developed to detect the worm in fish and water, and here we determined that the test on zebrafish intestines was effective for earlier detection. Four lines of zebrafish (AB, TU, 5D and Casper) were experimentally infected and evaluated by wet mounts and qPCR at 8, 15-, 22-, 31- and 44-day post-exposure (dpe). At the first two time points, only 8% of the wet mounts from exposed fish were identified as infected, while the same intestines screened by qPCR showed 78% positivity, with low and consistent cycle threshold (Ct) values at these times. Wet mounts at later time points showed a high prevalence of infection, but this was still surpassed by qPCR.

Development of TaqMan quantitative PCR assay for detection of Nocardia seriolae in fish and the environment.

Nocardia seriolae is a gram-positive bacterium that causes nocardiosis, threatening fish farming. Advanced nocardiosis is challenging to control; thus, accurate detection methods of the causal agent in the early disease stage are required. In this study, we developed a TaqMan fluorescence quantitative PCR (qPCR) assay for quantitative detection of N. seriolae in fish tissues and water samples. A pair of highly specific primers and a TaqMan probe were designed based on the N. seriolae 16S23S rRNA internal transcribed spacer (ITS) region. A high correlation coefficient (R2 = 0.998) of the standard curve with a 99.5% efficiency was obtained. The qPCR detection limit of the method was as low as 19.8 copies/µL, 1000 times more sensitive than conventional PCR, and has a good performance in the detection of cultured bacteria (y = -3.750× + 48.075, R2 = 0.974). Even 1.42 CFU/mL N. seriolae collected from 500 mL of natural pond water can be detected. Furthermore, a linear model for the relationship between the log of bacteria load and Cq values in water was established (y = -3.239× + 40.978), and the R2 value was 0.979. This assay was used for accurate N. seriolae detection in fish tissues, water samples, feeds and soils. This study provides a valuable tool for the early detection and control of nocardiosis in aquaculture.

A Novel Specific Single-Chain Variable Fragment Diagnostic System for Viral Hemorrhagic Septicemia Virus.

Viral hemorrhagic septicemia virus (VHSV), one of the most important viral marine pathogens worldwide, has a broad range of hosts, such as members of the families Salmonidae and Paralichthyidae. In addition to being highly contagious, VHSV causes high lethality. The transmission of VHSV can be both vertical and horizontal. In fish, the resolution of VHSV infection is challenging. Thus, early diagnosis of VHSV infections is critical, especially in fish farms that have a high population of juvenile fish. Serological methods are commonly used to detect viral antigens. However, limited serological methods are available for marine viruses. In this study, a VHSV-specific single-chain variable fragment (scFv), E5, was selected using the yeast surface display and phage display systems. scFv, a type of recombinant antibody, comprises a variable heavy chain ([Formula: see text]) and a variable light chain ([Formula: see text]) connected by a polypeptide linker. An scFv clone was selected from the VHSV glycoprotein-expressing yeast cells using the bio-panning method. The scFv-encoding gene was subcloned and expressed in the Escherichia coli expression system. The binding affinity of the expressed and purified scFv protein was determined using an enzyme-linked immunosorbent assay and western blotting. Thus, this study reported a method to identify VHSV-specific scFv using bio-panning that can be utilized to develop a diagnostic system for other viruses.

Quick detection of Carassius auratus herpesvirus (CaHV) by recombinase-aid amplification lateral flow dipstick (RAA-LFD) method.

Crucian carp (Carassius auratus) is one of the major freshwater species and is also a common food fish in China. Recently, Carassius auratus herpesvirus (CaHV) could induce fatal viral disease with high mortality of crucian carp, which had caused huge economic losses. In this study, we described a rapid and simple recombinase-aid amplification (RAA) assay coupled with lateral flow dipstick (LFD), which could achieve sensitive diagnosis of tumor necrosis factor receptor (TNFR) of CaHV within 35 min at 40°C. Our RAA-LFD method had a satisfactory detection limit of 100 gene copies per reaction, which was 100-fold more sensitive than traditional PCR. In addition, no cross-reaction was observed with other viral pathogens, including koi herpesvirus (KHV), cyprinid herpesvirus 2 (CyHV-2), infectious hematopoietic necrosis virus (IHNV), spring viremia of carp virus (SVCV) and grass carp reovirus (GCRV). Furthermore, the overall cost of the method was cut in half compared to previous studies. In conclusion, RAA-LFD assay is therefore, a promising alternative for point-of-care testing (POCT) of CaHV, which is feasible and of certain value in application of aquatic disease control.

Anisakis spp, DNA detection in paraffin-embedded tissue biopsies recovered from patients with gastritis using real-time PCR in Bushehr, Persian Gulf, Iran.

Anisakiasis is a zoonotic fish-born parasitic disease caused by anisakid nematodes. Paraffin-embedded blocks containing biopsy samples taken from patients suffering gastritis with unknown causes were investigated by real-time PCR, in the Bushehr region, Iran; where human anisakiasis has not been reported, so far. A total of 50 paraffin-embedded blocks were randomly selected from 250 archived blocks of the patients with gastritis. A SYBER green-based real-time PCR targeting the ITS1 region was developed for the identification of Anisakis genus. An 86 bp partial fragment of the Anisakis spp. ITS1 gene was amplified successfully. A total of 3 out of 50 samples (6 %) had positive amplification in the samples and their pathology reports showed a significant finding of moderate chronic gastritis with or without ulcers. In conclusion, the developed qPCR could be used for detecting Anisakis spp. larval DNA in human biopsy blocks. This study showed the hidden human cases of anisakiasis in the Bushehr for the first time.

Concentration and quantification of Tilapia tilapinevirus from water using a simple iron flocculation coupled with probe-based RT-qPCR.

Background: Tilapia tilapinevirus, also known as tilapia lake virus (TiLV), is a significant virus that is responsible for the die-off of farmed tilapia across the globe. The detection and quantification of the virus using environmental RNA (eRNA) from pond water samples represents a potentially non-invasive and routine strategy for monitoring pathogens and early disease forecasting in aquaculture systems. Methods: Here, we report a simple iron flocculation method for concentrating viruses in water, together with a newly-developed hydrolysis probe quantitative RT-qPCR method for the detection and quantification of TiLV. Results: The RT-qPCR method designed to target a conserved region of the TiLV genome segment 9 has a detection limit of 10 viral copies per µL of template. The method had a 100% analytical specificity and sensitivity for TiLV. The optimized iron flocculation method was able to recover 16.11 ± 3.3% of the virus from water samples spiked with viral cultures. Tilapia and water samples were collected for use in the detection and quantification of TiLV disease during outbreaks in an open-caged river farming system and two earthen fish farms. TiLV was detected from both clinically sick and asymptomatic fish. Most importantly, the virus was successfully detected from water samples collected from different locations in the affected farms (i.e., river water samples from affected cages (8.50 × 103 to 2.79 × 105 copies/L) and fish-rearing water samples, sewage, and reservoir (4.29 × 103 to 3.53 × 104 copies/L)). By contrast, TiLV was not detected in fish or water samples collected from two farms that had previously experienced TiLV outbreaks and from one farm that had never experienced a TiLV outbreak. In summary, this study suggests that the eRNA detection system using iron flocculation, coupled with probe based-RT-qPCR, is feasible for use in the concentration and quantification of TiLV from water. This approach may be useful for the non-invasive monitoring of TiLV in tilapia aquaculture systems and may support evidence-based decisions on biosecurity interventions needed.

CRISPR-based platform for rapid, sensitive and field-deployable detection of scale drop disease virus in Asian sea bass (Lates calcarifer).

Scale drop disease virus (SDDV) is a major pathogen of Asian sea bass that has emerged in many countries across the Asia Pacific since 1992 and carries the potential to cause drastic economic losses to the aquaculture sector. The lack of an approved vaccine for SDDV necessitates timely prevention as the first line of defence against the disease, but current diagnostic platforms still face challenges that render them incompatible with field applications, particularly in resource-limited settings. Here, we developed a novel detection platform for SDDV based on a CRISPR-Cas12a-based nucleic acid detection technology combined with recombinase polymerase amplification (RPA-Cas12a). Using the viral adenosine triphosphatase (SDDV-ATPase) gene as a target, we achieved the detection limit of 40 copies per reaction and high specificity for SDDV. The coupling with fluorescence and lateral flow readouts enables naked-eye visualization and straightforward data interpretation requiring minimal scientific background. Compared with semi-nested PCR in field sample evaluation, our RPA-Cas12a assay is more sensitive and capable of detecting SDDV in asymptomatic fish. Importantly, the entire workflow can be carried out at a constant temperature of 37°C within an hour from start to finish, thus removing the need for an expensive thermal cycling apparatus and long turnaround times associated with PCR-based methods. Therefore, owing to its high accuracy, rapidity and user-friendliness, the developed RPA-Cas12a platform shows the potential for diagnosis of SDDV at point of need and could be a valuable tool to help protect fish farming communities from large-scale epidemics.

Aquaculture Reuse Water, Genetic Line, and Vaccination Affect Rainbow Trout (Oncorhynchus mykiss) Disease Susceptibility and Infection Dynamics.

Infectious hematopoietic necrosis virus (IHNV) and Flavobacterium psychrophilum are major pathogens of farmed rainbow trout. Improved control strategies are desired but the influence of on-farm environmental factors that lead to disease outbreaks remain poorly understood. Water reuse is an important environmental factor affecting disease. Prior studies have established a replicated outdoor-tank system capable of varying the exposure to reuse water by controlling water flow from commercial trout production raceways. The goal of this research was to evaluate the effect of constant or pulsed reuse water exposure on survival, pathogen prevalence, and pathogen load. Herein, we compared two commercial lines of rainbow trout, Clear Springs Food (CSF) and Troutex (Tx) that were either vaccinated against IHNV with a DNA vaccine or sham vaccinated. Over a 27-day experimental period in constant reuse water, all fish from both lines and treatments, died while mortality in control fish in spring water was <1%. Water reuse exposure, genetic line, vaccination, and the interaction between genetic line and water exposure affected survival (P<0.05). Compared to all other water sources, fish exposed to constant reuse water had 46- to 710-fold greater risk of death (P<0.0001). Tx fish had a 2.7-fold greater risk of death compared to CSF fish in constant reuse water (P ≤ 0.001), while risk of death did not differ in spring water (P=0.98). Sham-vaccinated fish had 2.1-fold greater risk of death compared to vaccinated fish (P=0.02). Both IHNV prevalence and load were lower in vaccinated fish compared to sham-vaccinated fish, and unexpectedly, F. psychrophilum load associated with fin/gill tissues from live-sampled fish was lower in vaccinated fish compared to sham-vaccinated fish. As a result, up to forty-five percent of unvaccinated fish were naturally co-infected with F. psychrophilum and IHNV and the coinfected fish exhibited the highest IHNV loads. Under laboratory challenge conditions, co-infection with F. psychrophilum and IHNV overwhelmed IHNV vaccine-induced protection. In summary, we demonstrate that exposure to reuse water or multi-pathogen challenge can initiate complex disease dynamics that can overwhelm both vaccination and host genetic resistance.

First evidence of fish nocardiosis in Mexico caused by Nocardia seriolae in farmed red drum (Sciaenops ocellatus, Linnaeus).

Between August and December 2013, the offshore cages of a commercial marine farm culturing red drum Sciaenops ocellatus in Campeche Bay Mexico were affected by an outbreak of an ulcerative granulomatous disease with up to 70% cumulative mortality. Thirty-one adults displaying open ulcers on the skin were submitted for diagnosis. At necropsy, multiple white-yellowish nodules (0.1-0.5 cm in diameter) were present in all internal organs, where the kidney and the spleen were the most severely affected. Histopathology evinced typical systemic granulomatous formations. Gram and Ziehl-Neelsen stains on tissue imprints, bacterial swabs and tissue sections revealed Gram-positive, acid-fast, branching beaded long rod filamentous bacteria. Tissue samples resulted positive for nocardiosis with a Nocardia genus-specific nested PCR. Definite identification at the species level and taxonomic positioning of the fastidious pathogen were achieved through a specific Nocardia seriolae PCR and by sequencing the gyrB gene of pure isolates. After administration of antibiotics during fry production, a posterior follow-up monitoring (from 2014 to 2017) detected mild but recurrent outbreaks of the bacteria with no seasonality pattern. To the extent of our knowledge, this is the first report of piscine nocardiosis in Mexico and the first time this disease is detected in red drum.

Systemic phaeohyphomycosis in splitnose rockfish (Sebastes diploproa) caused by Devriesia sp.

A novel pathogen was documented after two wild-caught, juvenile, splitnose rockfish presented with buphthalmia, grey corneal endothelial plaques and evidence of uveitis. Cytologic evaluation of ocular contents revealed fungal hyphae. Histologic evaluation identified multiple fungal granulomas and granulomatous inflammation in the globes, periocular tissue and heart. Fungi were slender, hyphenated and branched at angles, had parallel cell walls and had brown pigmentation in haematoxylin- and eosin-stained sections. Both fish were diagnosed with phaeohyphomycosis. Culture with nuclear ribosomal RNA internal transcribed spacer (ITS) segment identification further classified the fungus as Devriesia sp., which has not been previously documented as a cause of disease in animals.

Hook-shaped enterolith and secondary cachexia in a free-living grey nurse shark (Carcharias taurus, Rafinesque 1810).

The carcass of a critically endangered, juvenile female grey nurse shark (Carcharias taurus, Rafinesque 1810) was recovered from a south-eastern Australian beach and subjected to necropsy. The 1.98-m-long shark exhibited advanced cachexia with its total weight (19.0 kg) and liver weight (0.37 kg) reduced by 60% and 89%, respectively, compared with a healthy individual of the same length. Marked tissue decomposition was evident preventing histopathology and identification of a definitive cause of death. At necropsy, the abdominal organs were abnormally displaced and showed marked reductions in size compared with a healthy individual of the same size. Importantly, a hook-shaped enterolith (HSE), with a rough surface and cream in colour, was found within the spiral valve of the intestine and is to the authors' knowledge, the first description of such in any marine animal. X-ray diffractometry showed that the HSE comprised the minerals monohydrocalcite (Ca[CO3].H2O; ~70 wt%) and struvite (Mg [NH4 ] [PO4 ]. [H2 O]6 ; ~30 wt%). A CT scan showed concentric lamellate concretions around a 7/o offset J-hook that formed the nidus of the HSE. Nylon fishing line attached to the hook exited the HSE and was evident in the abdominal cavity through a perforation in the intestinal wall where the posterior intestinal artery merges. The most parsimonious reconstruction of events leading to enterolithiasis and secondary cachexia in this shark was the consumption of a hooked fish and subsequent hook migration causing perforations of the cardiac stomach wall followed by the thin, muscular wall of the apposed, sub-adjacent intestine.

Simultaneous detection of multiple bacterial and viral aquatic pathogens using a fluorogenic loop-mediated isothermal amplification-based dual-sample microfluidic chip.

Rapid and user-friendly diagnostic tests are necessary for early diagnosis and immediate detection of diseases, particularly for on-site screening of pathogenic microorganisms in aquaculture. In this study, we developed a dual-sample microfluidic chip integrated with a real-time fluorogenic loop-mediated isothermal amplification assay (dual-sample on-chip LAMP) to simultaneously detect 10 pathogenic microorganisms, that is Aeromonas hydrophila, Edwardsiella tarda, Vibrio harveyi, V. alginolyticus, V. anguillarum, V. parahaemolyticus, V. vulnificus, infectious hypodermal and haematopoietic necrosis virus, infectious spleen and kidney necrosis virus, and white spot syndrome virus. This on-chip LAMP provided a nearly automated protocol that can analyse two samples simultaneously, and the tests achieved limits of detection (LOD) ranging from 100 to 10-1  pg/µl for genomic DNA of tested bacteria and 10-4 to 10-5  pg/µl for recombinant plasmid DNA of tested viruses, with run times averaging less than 30 min. The coefficient of variation for the time-to-positive value was less than 10%, reflecting a robust reproducibility. The clinical sensitivity and specificity were 93.52% and 85.53%, respectively, compared to conventional microbiological or clinical methods. The on-chip LAMP assay provides an effective dual-sample and multiple pathogen analysis, and thus would be applicable to on-site detection and routine monitoring of multiple pathogens in aquaculture.

Epizootic neoplasia in a managed population of Atlantic bumper fish, Chloroscombrus chrysurus (Osteichthyes: Carangidae), at a public aquarium.

Advances in fish medicine and husbandry have increased the average lifespans of specimens in managed aquarium populations. As a result, an increased incidence and variety of neoplasia is expected. This work characterizes diverse neoplasms arising within a managed population of Atlantic bumper fish acquired via repeated collections from the Charleston Harbor region. A total of 76 neoplasms were evaluated histologically from 41 of 45 fish that died or were killed over a 46-month period, including cutaneous hemangiomas and hemangiosarcomas, lepidocytomas and lepidosarcomas, fibromas, vertebral body or cutaneous osteomas, disseminated lymphomas, testicular leiomyomas, cutaneous or branchial fibrosarcomas, myxomas, fibroblastic lepidosarcoma, teratoid medulloepithelioma, ganglioglioma, malignant nerve sheath tumour, cardiac rhabdomyoma, cutaneous rhabdomyosarcoma, soft tissue sarcoma and renal adenoma. Perioral and cutaneous lesions of vascular and scale origin were observed most frequently. Other, often malignant, neoplasms arose within these benign lesions, resulting in extensive local tissue invasion. However, excluding disseminated lymphomas, metastasis was only detected in one case of hemangiosarcoma. These findings suggest early surgical intervention may limit tissue destruction and loss of display quality. This report details a variety of common and rare neoplasms in fish, as well as the first characterizations of neoplasia in Atlantic bumper and ganglioglioma in fish.

Histological and immunohistochemical characterization of 17 gonadal tumours in koi carp (Cyprinus carpio koi).

Reports on abdominal tumours in koi carp are scarce and most are from the gonads. Their histological diagnosis is challenging due to the occurrence of mixed populations of neoplastic cells and the few availability of cross-reactive antibodies in fish tissues. The present study aims to provide a histopathological characterization of seventeen gonadal tumours, enriched by a wide antibody panel (vimentin, CD117, placental alkaline phosphatase-PLAP, AE1/AE3 cytokeratin, E-cadherin, proliferating cell nuclear antigen-PCNA, müllerian-inhibiting substance-MIS, GATA4 and Inhibin-α) applied on whole and tissue microarray (TMA) sections. Abdominal enlargement was associated with tumours filling 30%-80% of the abdominal cavity; frequently, the gonads had been completely replaced by neoplastic tissue. Twelve cases were characterized as sex cord-stromal tumours (SCSTs), three as germ cell tumours (GCTs), one as mixed germ cell sex cord-stromal tumour (MGCSCST) and one as carcinoma. By immunohistochemistry, PLAP enabled confirmation of GCTs, ovarian carcinoma and the objective identification of a further cell component in 8 out of the 12 SCSTs that were reclassified as mixed tumours. The use of an immunohistochemical panel can help in refining the histological diagnosis, but the morphological diagnosis still represents the main tool for the characterization of these tumours in koi carp.

Polyunsaturated Fatty Acids Influence LPS-Induced Inflammation of Fish Macrophages Through Differential Modulation of Pathogen Recognition and p38 MAPK/NF-κB Signaling.

Polyunsaturated fatty acids (PUFAs) not only serve as essential nutrients but also function as modulators of the immune response in marine fish. However, their immunomodulatory mechanism is poorly understood given that the underlying regulation of the innate immune response in fish has not been fully elucidated. Hence, study of the innate immunity of fish could help elucidate the mechanism by which PUFAs affect the fish immune response. Here, we used combined transcriptome analysis and in vitro experimentation to study the mechanism of LPS-induced inflammation. Transcriptome profiling indicated that LPS elicited strong pro-inflammatory responses featuring high expression levels of pathogen recognition receptors (PRRs) and cytokines along with the activation of NF-κB and MAPK signaling pathways. The transcription factor p65 alone could increase the transcription of IL1ß by binding to the promoter of IL1ß, and this promoting effect disappeared after mutation or deletion of its binding sites. We then examined the effects of PUFAs on the levels of gene expression and the abundance of proteins of critical kinases associated with LPS-induced inflammation. We found that LA exerts pro-inflammatory response while ALA, EPA, and DHA induced anti-inflammatory effects by modulating the expression of PRRs, phosphorylation of IKK and p38, and the nuclear translocation of p65. Overall, this study advances our understanding of the regulatory mechanisms by which PUFAs regulate LPS-induced inflammation in a non-model fish species.