Tissue factor pathway inhibitors disrupt structures of rhabdovirus/ranairidovirus and inhibit viral infection in Chinese perch, Siniperca chuatsi.

Viral diseases have caused great economic losses to the aquaculture industry. However, there are currently no specific drugs to treat these diseases. Herein, we utilized Siniperca chuatsi as an experimental model, and successfully extracted two tissue factor pathway inhibitors (TFPIs) that were highly distributed in different tissues. We then designed four novel peptides based on the TFPIs, named TS20, TS25, TS16, and TS30. Among them, TS25 and TS30 showed good biosafety and high antiviral activity. Further studies showed that TS25 and TS30 exerted their antiviral functions by preventing viruses from invading Chinese perch brain (CPB) cells and disrupting Siniperca chuatsi rhabdovirus (SCRV)/Siniperca chuatsi ranairidovirus (SCRIV) viral structures. Additionally, compared with the control group, TS25 and TS30 could significantly reduce the mortality of Siniperca chuatsi, the relative protection rates of TS25 against SCRV and SCRIV were 71.25 % and 53.85 % respectively, and the relative protection rate of TS30 against SCRIV was 69.23 %, indicating that they also had significant antiviral activity in vivo. This study provided an approach for designing peptides with biosafety and antiviral activity based on host proteins, which had potential applications in the prevention and treatment of viral diseases.

The Stability and Efficency of CPB Cells Were Acclimated for Virus Proliferation.

BACKGROUND: Vaccinations are still the most effective means of preventing and controlling fish viral diseases, and cells are an important substrate for the production of a viral vaccine. Therefore, the rapid-stable growth and virus sensitivity of cells are urgently needed. METHODS: Chinese perch brain 100th passage (CPB p100) were acclimated in a low serum with 5% FBS L-15 for 50 passages, then transferred to 8% FBS L-15 for 150 passages. Additionally, the morphology and cell type of CPB 300th passage (CPB p300) cells were identified. We analyzed the transfection efficiency and virus sensitivity of CPB p300 cells, and then optimized the conditions of ISKNV, SCRV, and LMBV multiplication in CPB cells. RESULTS: CPB p300 cells were more homogeneous, and the spread diameter (20-30) µm in CPB p300 cells became the dominant population. The doubling time of CPB p300 was 1.5 times shorter than that of CPB p100.However, multiplication rate of CPB p300 was 1.37 times higher than CPB p100. CPB p300 cells were susceptible to ISKNV, SCRV, and LMBV, and the optimal conditions of ISKNV, SCRV, and LMBV multiplication were simultaneous incubation, 0.6 × 105 cells/cm2 and MOI = 0.1; infection at 48 h, 0.8 × 105 cells/cm2 and MOI = 0.01; simultaneous incubation, 0.7 × 105 cells/cm2 and MOI = 0.05, respectively. The time and economic costs of ISKNV, SCRV, and LMBV multiplication in CPB p300 cells were significantly reduced. CONCLUSIONS: The acquisition of CPB p300 cells laid a good material foundation for the production of ISKNV, SCRV, and LMBV vaccines.

A Novel and Effective Therapeutic Method for Treating Aeromonas schubertii Infection in Channa maculata.

Aeromonas schubertii is a pathogen that severely affects aquatic animals, including the snakehead, Channa maculata. Lytic bacteriophages have been recognized as effective alternatives to antibiotics for controlling bacterial infections. However, there have been no reports of A. schubertii phages as far as we know. In this study, a lytic bacteriophage SD04, which could effectively infect A. schubertii, was isolated from pond water cultured with diseased snakehead. The SD04 phage formed small, round plaques on Petri dishes. Electron microscopy revealed a hexagonal head and a contractile tail. Based on its morphology, it may belong to the Myoviridae family. Two major protein bands with molecular weights of 50 and 38 kilodaltons were observed after the phage was subjected to SDS-PAGE. The phage showed a large average burst size, high specificity, and a broad host range. When stored at 4 °C, phage SD04 had high stability over 12 months and showed almost no variation within the first six months. All fish were healthy after both intraperitoneal injection and immersion administration of SD04, indicating the safety of the phage. After treatment with SD04, Channa maculata in both phage therapy groups and prevention groups showed high survival rates (i.e., 83.3 ± 3.3% and 100 ± 1.3%, respectively). Phage therapy inhibits bacterial growth in the liver, the target organ of the infected Channa maculat. The experimental results indicate the potential use of phage SD04 for preventing A. schubertii infection in Channa maculata.

An Avirulent Largemouth Bass Birnavirus Vaccine Candidate Protects Largemouth Bass against Birnavirus Infection.

BACKGROUND: Largemouth bass birnavirus (LBBV) disease outbreaks in largemouth bass fingerlings lead to high mortality in China. Therefore, the development of immersion immunization strategies is paramount. METHODS: An avirulent LBBV strain was screened using a fish challenge assay. The proliferation dynamics of the avirulent strain were determined in vitro and in vivo. The efficacy of the avirulent vaccine was evaluated using immune gene expression, viral load, and a virus challenge, and the safety was also assessed using a reversion to virulence test. RESULTS: An avirulent virus strain, designated as largemouth bass birnavirus Guangdong Sanshui (LBBV-GDSS-20180701), was selected from five fish birnavirus isolates. The proliferation peak titer was 109.01 TCID50/mL at 24 hpi in CPB cells and the peak viral load was 2.5 × 104 copies/mg at 4 dpi in the head kidneys and spleens of largemouth bass. The largemouth bass that were immersed within an avirulent vaccine or injected with an inactivated vaccine were protected from the virulent LBBV challenge with a relative percent survival (RPS) of 75% or 42.9%, respectively. The expression levels of IL-12, MHCI, MHCII, CD8, CD4, and IgM in the avirulent group were significantly upregulated at a partial time point compared to the inactivated vaccine group. Moreover, the viral load in the avirulent vaccine group was significantly lower than those in the inactivated vaccine group and control group using real-time PCR. CONCLUSIONS: LBBV-GDSS-20180701 is a potential live vaccine candidate against LBBV disease.