Perkinsus olseni is a major pathogenic parasite that threatens shellfish aquaculture and can trigger mass mortality and considerable economic losses. Conventional culture-based assays and PCR-based detection protocols for P. olseni recommended by the World Organization for Animal Health (OIE) involve long assay durations and heavy dependence on sophisticated instruments, making them unsuitable for on-site rapid screening. To establish a simple, rapid and highly sensitive detection method for early warning of P. olseni at aquaculture sites, this study integrated isothermal recombinase polymerase amplification (RPA) with the specific-recognition capacity of PfAgo nuclease to construct an RPA-PfAgo detection assay. The assay achieved favourable specificity with a limit of detection as low as 30 copies. Its practical applicability was validated using 55 real-world shellfish samples. Performing at constant temperatures, the RPA-PfAgo detection assay features a simple procedure and short turnaround time, effectively addressing the drawbacks of conventional detection approaches. This assay is suitable for shellfish seed quarantine and routine pathogen surveillance in shellfish farms, and provides solid support for the early prevention and control of perkinsosis caused by P. olseni.
Decapod iridescent Virus 1 (DIV1) poses a major threat to crustacean aquaculture, yet effective antiviral agents remain limited. In this study, a biochemical screening assay was established using recombinant DIV1 dUTPase, a predicted viral enzyme selected as a biochemical screening target. Screening of 108 laboratory-preserved medicinal-plant crude extracts and subsequent evaluation of monomeric compounds identified chrysin as a candidate inhibitor. In the recombinant-enzyme assay, chrysin inhibited DIV1 dUTPase activity. In DIV1-infected Litopenaeus vannamei, chrysin treatment reduced viral loads and improved survival after challenge. Relative to the DIV1-infected control, chrysin treatment was also associated with higher SOD and CAT activities, GSH content and T-AOC. These findings support further evaluation of chrysin as an anti-DIV1 lead compound.
Recently, a disease outbreak was observed in farmed yellow catfish (Tachysurus fulvidraco) in Tianjin, China. Naturally infected fish exhibited surface haemorrhages, abdominal distension, anal redness, and hemorrhagic ascites. Histopathological examination revealed systemic lesions in the stomach, intestine, liver, and spleen. Two Gram-negative bacterial isolates, designated PFL and PFS, were isolated from the liver and spleen of diseased fish, respectively. The isolates were identified as Aeromonas veronii and Citrobacter braakii by morphological and biochemical characterisation combined with 16S rRNA and gyrB gene sequence analyses. Experimental infection with single or combined isolates demonstrated that both A. veronii and C. braakii were pathogenic to yellow catfish, with A. veronii showing higher virulence. Fish infected with A. veronii or co-infected with the two isolates displayed similar external clinical signs with the naturally diseased fish. Hemolytic assays revealed beta-hemolysis in A. veronii and gamma-hemolysis in C. braakii. These findings indicate that both A. veronii and C. braakii contribute to disease outbreaks in yellow catfish.
Francisella halioticida has emerged as a significant pathogen of cultured molluscs, causing substantial economic losses due to mass mortalities. The Francisella Pathogenicity Island encodes a Type VI secretion system essential for virulence and intracellular persistence. However, the F. halioticida pathogenicity island shows marked sequence divergence from well-characterized Francisella strains, limiting the use of conventional sequence-based annotation. This study performed a comprehensive structure-based annotation of the F. halioticida pathogenicity island using AlphaFold3 predictions combined with systematic structural similarity searches. This approach identified five previously uncharacterized FPI components and their respective T6SS homologues. Notably, F. halioticida lacks the IglF protein and the N-terminal alpha-helix extension of IglG but retains a nearly complete and functionally coherent Type VI secretion system architecture.
Northern snakehead (Channa argus) is a bimodal air-breathing fish in which erythrocyte-associated parameters remain relatively stable during physiological challenges such as aerial exposure. However, whether this characteristic is maintained during systemic infection remains unclear. In this study, temporal host responses were characterized following experimental Aeromonas salmonicida subsp. salmonicida infection by integrating haematological, leukocyte morphological, biochemical, and multi-tissue transcriptional analyses. Most erythrocyte-associated indices, including RBC, HGB, HCT, MCV, MCH, and RDW, showed no significant temporal changes, whereas MON, GRA, and PLT exhibited significant responses. Morphologically abnormal leukocytes increased from approximately 7% in healthy fish to 56% at 6 days post-infection and remained above 50% through 8 days, indicating persistent leukocyte-associated cellular alterations during infection. Biochemical responses were also selective, with changes primarily in protein- and metabolism-associated parameters. Transcriptional responses differed among peripheral blood, head kidney, spleen, and liver, including sustained casp7 suppression in the head kidney and distinct inflammatory and apoptosis-associated responses across tissues. Two-way ANOVA further identified significant tissue & times; time interactions for casp8, tnf alpha, and casp3. Collectively, A. salmonicida infection in northern snakehead was characterized by comparatively limited temporal variation in most erythrocyte-associated indices, alongside selective leukocyte, biochemical, and tissue-dependent transcriptional responses. These findings highlight a contrasting blood response characterized by persistent leukocyte alterations despite comparatively limited temporal variation in most erythrocyte-associated indices during bacterial infection in northern snakehead.
