Journal of fish diseases最新文献

Megalocytivirus pagrus 1 is a major viral pathogen that causes high mortality and economic losses in aquaculture worldwide. Although PCR-based diagnostics are highly sensitive and specific, their equipment dependence and long assay times limit field applicability. In this study, a digital multienzyme isothermal rapid amplification with exonuclease probe (digital MIRA-EXO) assay was developed and validated for rapid and specific detection of red sea bream iridovirus (RSIV), infectious spleen and kidney necrosis virus (ISKNV), and turbot reddish body iridovirus (TRBIV). The digital MIRA-EXO assay specifically identified Megalocytivirus pagrus 1 without cross-reactivity with non-target samples, including four fish cell lines and 13 different fish pathogens. Analytical sensitivity, expressed as 95% limit of detection, ranged from 146.87 to 201.6 copies/μL across the three viruses. Diagnostic performance evaluation of 180 fish samples showed high sensitivity (92.22%), specificity (100%), and overall accuracy (96.11%), with an area under the receiver operating characteristic (ROC) curve of 0.971. Substantial agreement with reference assays was observed for experimentally infected (κ = 0.786) and field samples (κ = 0.792), and between column-based and rapid DNA extraction methods (κ = 0.783). Overall, the digital MIRA-EXO assay provides a rapid, accurate, and field-deployable diagnostic tool for Megalocytivirus pagrus 1 detection.

Piscine lactococcosis, a major bacterial disease in aquaculture causing haemorrhagic septicaemia and economic losses, was traditionally attributed to Lactococcus garvieae but now includes emerging pathogens like Lactococcus petauri. Amid rising antibiotic resistance, bacteriophage therapy offers a sustainable alternative. This study aimed to isolate, characterise, and evaluate the therapeutic potential of a lytic phage targeting L. petauri. Phage Lp ACM616_1 was isolated from aquaculture effluents in Türkiye using L. petauri as the host. Whole-genome analysis and transmission electron microscopy were used to identify it as a member of the Caudoviricetes class. It has an icosahedral head (~31 nm) and a long tail (~107 nm). Host range testing showed specificity to 10 of 11 L. petauri strains, with no activity against other pathogens. Lp ACM616_1 exhibited robust stability (70%-90% viability at -20°C to 50°C; > 75% at pH 4-10), rapid adsorption (> 85% in 30 min), a 10-20 min latent period, and high burst size. In vitro, it suppressed bacterial growth at low MOIs and showed no cytotoxicity in EPC cells. Genomic characterisation revealed a 23,157 bp double-stranded DNA genome with 36.3% GC content and 40 predicted coding sequences, supporting its classification as a novel species within the genus Chertseyvirus lacking virulence and antimicrobial resistance genes. In rainbow trout challenge trials, intraperitoneal phage injection of the phage post-infection significantly improved survival to 77% compared to 40% in the untreated control group. In contrast, oral delivery via feed was found to be ineffective, yielding a survival rate of 42%. This first L. petauri-specific phage demonstrates promising biocontrol potential, advocating for cocktails and encapsulation to enhance aquaculture applications.

Nervous necrosis virus (NNV) is the causative agent of viral nervous necrosis, otherwise known as viral encephalopathy and retinopathy in larval and juvenile marine fish worldwide. The pandemic outbreaks have caused nearly 100% mortality in hatcheries, leading to significant economic losses in the aquaculture industry. Since NNV attacks the insufficient immune competence fish at early developmental stages, there is an urgent need for effective antiviral drugs. Several nucleoside analogues, including Molnupiravir, have been developed to target viral RNA-dependent RNA polymerase (RdRp). This study examines the antiviral activity of Molnupiravir against NNV isolated from giant grouper (GG), Epinephelus lanceolatus (Bloch), using grouper brain (GB) cells as an infection model. The inhibition concentration for 50% of maximal effect (EC₅₀) of Molnupiravir on GGNNV propagation in GB cells was determined as 1.87 μM. Surprisingly, the titre of GGNNV reduced 50,000 at 100 μM of Molnupiravir treatment at 4-day post-infection, whereas treatment with 200 μM of Molnupiravir resulted in a 10⁵ reduction in viral titre. The inhibition of viral replication and translation was further examined using RNA-fluorescence in situ hybridization (FISH) and immunocytochemistry detection, respectively. Moreover, administration of 100 μM Molnupiravir at early time 0 hpi resulted in a dramatic decrease in viral load, reducing cell-associated virus by approximately 4000-fold and released virus by 6000-fold. Finally, the molecular docking simulation model supports the interaction between NHC-TP and GGNNV RdRp for the successful therapeutics. These results demonstrated that Molnupiravir is a very effective therapeutic drug for controlling viral nervous necrosis caused by NNV infection.

Kuruma shrimp (Penaeus japonicus) is an economically important shrimp perennially affected by diseases. In 2022, White Muscle Disease (WMD) was first characterised in this Penaeid species, caused by Photobacterium damselae subsp. damselae (Pdd). In this study, muscular and gut microbiome dynamics and their function in the disease progression are investigated by 16S rRNA metagenome sequencing using Illumina sequencing technologies. Alpha diversity indices showed that Pdd infection in the muscle, stomach, and intestine did not significantly change bacterial diversity between control and infected groups at all time points observed (Days 0, 1, 3, 5, 7 and 10). In the infected samples, the Shannon and Simpson indices increased starting Day 5 (D5), in congruence with the first observation of muscle whitening. Bacterial composition for the infected group at the genus level revealed that Photobacterium and Vibrio have increased their relative abundance in the muscle at Day 5 (D5) until Day 7 (D7), but declined at Day 10 (D10). As for stomach samples, Photobacterium declined in abundance and later increased significantly at Day 7 (D7). Photobacterium in the intestinal samples from the infected group increased at Day 5 (D5) but later decreased at Day 7 (D7). Meanwhile, linear discriminant analysis Effect Size (LEfSe) identified that most taxa belong to phylum Pseudomonadota, which can be potential markers for WMD. Moreover, the temporal dynamics of the amplicon sequencing variant ASV2, confirmed to be 100% homologous to the WMD-P3 strain used in this study, were characterised. For all tissues, the logarithmic relative abundance is considered high and very apparent in infected samples collected at Day 7 (D7). Overall, our study provides an understanding of the muscle and gut microbial community, specifically at the genus level, distinguished between WMD-infected and healthy Kuruma shrimps.

Perkinsus spp. are pathogenic protistan parasites that pose a significant threat to the aquaculture of the Hong Kong oyster (Crassostrea hongkongensis). This study investigated the transcriptional response of the Hong Kong oyster digestive gland to natural Perkinsus spp. infection. The parasite species was identified via PCR, while infection prevalence, tissue-specific abundance, and infection grade were assessed using Ray's Fluid Thioglycollate Medium (RFTM) culture. Transcriptomic analysis was performed to compare gene expression profiles among uninfected (Neg), lightly infected (L), and light-to-moderately infected (LM) oyster digestive glands. Results confirmed the presence of P. beihaiensis, with an overall infection prevalence of 85.5%. The digestive gland exhibited the highest infection prevalence (80.1%) and abundance (1880.0 ± 144.2 cells/g) compared to other tissues (gills, mantle, and siphon). Transcriptomic analysis of 12 samples (4 biological replicates per Neg, L, and LM group) using an Illumina Novaseq 6000 platform generated 102,827 unigenes. A total of 531 differentially expressed genes (DEGs) were common to both infected groups relative to the Neg group. GO and KEGG enrichment analyses highlighted key biological processes and pathways: up-regulated DEGs were associated with the cell cycle, antigen processing and presentation, and signal transduction, while down-regulated DEGs were linked to cholesterol metabolism, phagosome function, and fat digestion and absorption. qRT-PCR validation of 8 DEGs confirmed consistent expression patterns with RNA-seq data. These findings elucidate the tissue-specific infection patterns of P. beihaiensis in Hong Kong oysters in the digestive gland and provide critical insights into host-parasite interactions.

The study consisted of two experiments. First, a PK experiment was set up with a single oral dose at 20 mg/kg body weight, and second, a withdrawal time determination study, where the fish were fed once a day for 5 consecutive days at the same dosage. The DOX concentration in medicated feed, fish muscle, plasma, liver, and kidney was analysed by LC-MS/MS. Phoenix 8.1 and WT 1.4 software were used in the calculation for PK parameters and withdrawal time, respectively. DOX was absorbed quickly into plasma after oral medication with an absorption half-life T_1/2abs of 0.12 h, reaching the C_max (116 μg/L) at 1 h (T_max). DOX distribution was highest in the kidney, followed by the liver and plasma. The high DOX concentration in the kidney and liver supports the possibility of using DOX to treat relevant bacterial infections in snakehead. In conclusion, DOX is rapidly absorbed in snakehead after oral medication, has a significant tissue distribution, followed by slow elimination. Finally, after feeding medicated feed for 5 consecutive days, the withdrawal times of DOX in composite muscle and skin samples of snakehead were 4 days and 6 days (at 28.5°C) according to the maximum residue limits set by the European Commission (100 μg/kg for bovine, porcine and poultry) and Japanese authority (50 μg/kg), respectively.

A captive-reared cohort of Laurel Dace, Chrosomus saylori (Skelton 2001) (Cypriniformes: Leuciscidae), a federally-endangered species endemic to several streams in eastern Tennessee's Cumberland Plateau, exhibited signs of morbidity and trickling mortalities in a recirculating aquaculture system. Ziehl-Neelsen (ZN) staining revealed acid-fast rod-shaped bacteria within necrotic and intracellular lesions affecting the eye, swim bladder, spleen, and ovary. Histopathology showed granulomatous inflammation characterised by macrophages, multinucleated giant cells, and eosinophilic exudates. The eye, swim bladder, and ovary were particularly impacted: the choroid plexus of the eye displayed haemorrhages and dense cellular infiltrates; the swim bladder exhibited granulomatous inflammation with extensive fibrosis; the ovary contained caseous granulomas; and the choroid plexus of the eye displayed haemorrhages and dense cellular infiltrates. PCR amplification of the hsp65 gene (316 bp) from formalin-fixed and paraffin-embedded lesioned tissues was identical (100% similar; 424 bp fragment) to that of the type strain of Mycobacterium gordonae Bojalil et al. 1962 (AF547840). This is the first report of an infection by M. gordonae in Laurel Dace; the first record of this bacterium causing a lesion in the swim bladder, eye, or ovary; and the first record of M. gordonae from a freshwater fish in North America.

Vibrio species are the main causative agents of vibriosis, the most prevalent bacterial disease affecting the aquaculture industry. Since 2009, Vibrio parahaemolyticus has been identified as the key pathogen responsible for Acute Hepatopancreatic Necrosis Disease (AHPND), leading to widespread outbreaks across several Southeast Asian countries. Although V. parahaemolyticus is the primary causative agent of AHPND, recent studies have shown that other Vibrio species, such as V. campbellii and V. harveyi, can also cause AHPND when they acquire the PirAB toxin-encoding plasmid. This study aimed to develop DNA aptamers that specifically target the LuxP receptor protein, a key component of the quorum sensing system. QS regulates the expression of multiple virulence factors, including novel toxins involved in Vibrio infection. Using Ni-NTA Systematic Evolution of Ligand by Exponential Enrichment, the DNA aptamer, VPQ1, was developed, which binds to the LuxP receptor. From the SELEX cycles, a total of 1114 aptamer candidates were isolated and sequenced. Thermodynamic analyses demonstrated that the interactions between recombinant LuxP and the selected aptamers VPQ1 and VPQ2 were exothermic, with a dissociation constant of 23.3 nanomolar and 24.2 nanomolar, respectively. Functional assays showed that the treatment with VPQ1 led to downregulation of genes associated with bacterial metabolism, quorum sensing, flagellar assembly and biofilm formation. These findings suggest that aptamer VPQ1 may serve as a promising anti-virulence agent to disrupt QS and mitigate Vibrio infection in aquaculture systems.

The composition and structure of the helminth fauna in the intestine of the anostomid fish Rhytiodus microlepis was studied during two distinct periods in a floodplain lake in Central Amazonia. Additionally, the tissue damage in the infected intestine caused by the endoparasites was observed. Of the 38 fish analysed, a high prevalence of infection by Digenea (metacercariae) and Acanthocephala (> 90%) was observed; the metacercariae preferred the initial portions of the intestine, with a high frequency of occurrence in the lamina propria of the mucosa (75.9%), while the acanthocephalans occurred in the intermediate portions of intestine, often free in the lumen (99.5%). The histological alterations caused by the presence of the acanthocephalans were slight compression of the villi, desquamation/abrasion of the mucosal epithelium and hemorrhagic foci. Villi hypertrophy, intense leukocyte infiltration, formation of fibrous capsules and necrosis were the main responses associated with metacercariae. The composition and structure of infracommunities of intestinal helminths in R. microlepis showed seasonal variations, indicating that the hosts are susceptible to infections in the low and in the high-water seasons, with high values of intensity, abundance and pathogenicity (IAH value: 358.8 ± 99.22) of metacercariae during the low waters, suggesting that fish are more vulnerable to infections by these parasites in this period.