Gudgeon Novirhabdovirus Diagnostic Protocol

Siniperca chuatsi rhabdovirus (SCRV), also known as Gudgeon rhabdovirus, is a virus that mainly affects the snakehead fish (Siniperca chuatsi) and belongs to the Rhabdoviridae family. SCRV S3 exhibits strong pathogenicity to zebrafish (Danio rerio), with mortality rates of 53% for blue zebrafish and 73% for red zebrafish at 28°C. Zebrafish can serve as an infection model for SCRV. The snakehead fish is a valuable freshwater fish endemic to China, with high annual production and value. SCRV, as a significant disease, has caused substantial economic losses to the snakehead fish industry.

Purpose

This protocol aims to establish an effective detection and diagnostic process for Siniperca chuatsi Rhabdovirus, in order to promptly identify virus infections, prevent the spread of epidemics, and safeguard the healthy development of the aquaculture industry.

Pathological Characteristics

Siniperca chuatsi Rhabdovirus is a single-stranded negative-sense RNA virus belonging to the Novirhabdovirus family. This virus mainly spreads through water and can cause hemorrhagic septicemia in gudgeon, leading to significant mortality in severe cases.

Sample Collection

1. Sampling Targets: Gudgeon showing suspected symptoms such as surface hemorrhage, exophthalmos, pale gills, etc.
2. Sampling Sites: Blood, internal organs (especially liver and spleen), and gill tissue.
3. Sampling Tools: Sterile syringe, scissors, forceps, etc., must be disinfected beforehand.
4. Sample Volume: At least 1-2 grams of tissue or a suitable amount of blood per sample.

Laboratory Testing

Routine Pathological Examination

1. Prepare tissue sections and observe cellular pathological features.
2. Staining Methods: HE staining or other suitable staining methods.

Virus Isolation Cultivation

1. Select sensitive cell lines for virus isolation.
2. Observe cytopathic effects (CPE).

Molecular Biology Detection

1. PCR: Design specific primers to amplify viral gene segments.
2. Real-time quantitative PCR (qPCR): Quantitative analysis of viral load.

Immunological Detection

1. ELISA: Use specific antibodies to detect viral antigens.
2. Immunofluorescence (IFA): Directly detect viral antigens in tissues.

Result Interpretation

1. If pathological examination reveals typical Novirhabdovirus cellular pathology and molecular biology or immunological testing is positive, a diagnosis of Gudgeon Novirhabdovirus infection can be made.
2. If only pathological examination is abnormal and other tests are negative, clinical symptoms and epidemiological data should be considered together for comprehensive judgment.

Reporting and Management

1. Issue detailed test reports, including sample information, testing methods, results, and recommendations.
2. For confirmed cases, implement isolation, and disinfection measures promptly to prevent further virus spread.
3. Conduct a comprehensive assessment of the farming environment and develop corresponding prevention and control strategies.

Quality Control

1. Regularly calibrate equipment.
2. Validate methods using standard materials.
3. Participate in external quality assessment programs.

Precautions

1. Personnel must wear protective gear to avoid cross-contamination.
2. Dispose of all waste according to biosafety regulations.
3. Adhere to local laws, regulations, and industry standards.