In vitro characteristics of cyprinid herpesvirus 2: effect of kidney extract supplementation on growth

Diseases of Aquatic Organisms

Teruyuki Nakanishi¹  Azusa Nanjo¹  Takafumi Ito¹  Takashi Sakamoto¹  Tomoya Shibata¹  Masato Saito¹  Motohiko Sano¹  Keisuke Yoshii¹ 

关键词: Goldfish hematopoietic necrosis virus;    GFHNV;    Cyprinid herpesvirus 2;    Culture;    Kidney extract;    Growth temperature;    Cell susceptibility;    Carassius;   

DOI  :  10.3354/dao02885

学科分类：生物科学（综合）

来源: Inter-Research

【 摘 要 】

ABSTRACT: Herpesviral hematopoietic necrosis caused by goldfish hematopoietic necrosis virus (now identified as cyprinid herpesvirus 2, CyHV-2) has contributed to economic losses in goldfish Carassius auratus culture and is becoming a major obstacle in Prussian carp C. gibelio aquaculture in China. Several reports have described difficulties in culturing the virus, with the total loss of infectivity within several passages in cell culture. We succeeded in propagating CyHV-2 with a high infectious titer in a RyuF-2 cell line newly derived from the fin of the Ryukin goldfish variety using culture medium supplemented with 0.2% healthy goldfish kidney extract. The addition of kidney extract to the medium enabled rapid virus growth, resulting in the completion of cytopathic effect (CPE) within 4 to 6 d at 25°C. The extract also enabled reproducible virus culture with a titer of 10^5-6 TCID₅₀ ml^-1. The virus cultured using this protocol showed pathogenicity in goldfish after intraperitoneal injection. The virus grew in RyuF-2 cells at 15, 20, 25, 30, and 32°C but not at 34°C or higher. Higher incubation temperatures allowed earlier development of CPE, but culture at 30 and 32°C yielded a lower virus titer than that obtained at other temperatures because of heat inactivation of the propagated virus during cultivation. Cell lines derived from goldfish and ginbuna C. langsdorfii showed high susceptibility to the virus; cell lines from carp were susceptible to the virus using a medium containing goldfish kidney extract, but EPC, FHM, and BF-2 cell lines did not produce any CPE, even in the presence of the extract.

【 授权许可】

Unknown