Identification and pathological study of Cyprinid herpesvirus II in Sichuan

In October 2018, crucian carp (Carassius auratus) in a farm located in Zigong, Sichuan Province, developed a communicable disease with a mortality rate of 80%. In order to explore the cause of this disease, sick fish were subjected to necropsy, pathological observation, molecular biological examination and artificial infection test. The clinical manifestations of diseased fish included leaving the shoal of fish to swim alone; staying on the water surface; bodies blackened and body surface had haemorrhages. The necropsy showed haemorrhage, swelling and necrosis of organs such as gills, liver and kidney; swimming bladder had a lot of blutene chloaide. Histopathological changes showed that the respiratory epithelial cells of gill were swollen, necrosis and sloughting. The kidneys showed focal necrosis. The spleen tissue was extensively degenerated and necrotic, and the hematopoietic system collapsed. Vacuolar degeneration and necrosis occurred in the liver. Transmission electron microscopy showed that there were four different sizes of virus particles in the spleen and kidney, which were virus DNA core, empty viral capsid, solid capsid and enveloped mature virions. It can be seen that immature virions are present in the nucleus, and mature virions are present in the cytoplasm. Cyprinid herpesvirus Ⅱ (CyHV-2) completed nucleic acid replication and nucleocapsid assembly in the nucleus, and the envelope protein was obtained after transmembrane membrane. After filtering the diseased fish tissue homogenate, it was inoculated into EPC and FHM cell lines, and there was no cytopathic effect after 3 generations of blind transmission. The helicase gene fragment of CyHV-2 was amplified by PCR, and a positive band appeared. The artificial infection test was carried out by intraperitoneal injection of tissue homogenate. The test group showed the same clinical symptoms as the natural infection fish and the control group was normal. The cumulative mortality rate of the test group was 80%. Phylogenetic analysis of helicase gene by the neighbor-joining method revealed that the homology with CyHV-2 (YZ-01) was 99%. The results demonstrated that CyHV-2 contributed to this disease outbreak.