Evidence of Natural Nervous Necrosis Virus Infection in Juvenile Greenfin Horse-Faced Filefish (Thamnaconus septentrionalis) During an Aquaculture Mortality Event

Viral nervous necrosis (VNN), caused by nervous necrosis virus (NNV), is a devastating viral disease affecting marine fish aquaculture worldwide, particularly during larval and juvenile stages. The virus primarily targets the central nervous system and retina, often resulting in severe neurological symptoms and high mortality. To clarify the relationship between greenfin horse-faced filefish (Thamnaconus septentrionalis) and NNV, this study investigated an outbreak of abnormal mortality in juvenile fish from a marine aquaculture farm in Rizhao, Shandong Province, China. A comprehensive etiological diagnosis was conducted using bacterial isolation and identification, real-time quantitative RT-PCR (RT-qPCR), histopathological analysis, transmission electron microscopy (TEM), and phylogenetic analysis. Diseased fish exhibited typical neurological signs, including spiral swimming, anorexia, and body darkening. Several opportunistic bacteria were isolated from affected fish; however, they were not considered the primary cause of mortality. RT-qPCR results demonstrated that all diseased samples were positive for NNV nucleic acid, with high viral loads detected in neural tissues. Histopathological examination revealed pronounced vacuolation and neuronal nuclear pyknosis in the brain and retinal tissues, while TEM identified abundant non-enveloped, icosahedral viral particles approximately 25–26 nm in diameter, consistent with the ultrastructural characteristics of NNV. Phylogenetic analysis based on the RNA2 capsid protein gene revealed that the NNV isolate clustered within the red-spotted grouper nervous necrosis virus (RGNNV) genotype and showed a close evolutionary relationship with RGNNV strains previously reported from groupers and barramundi. Based on clinical manifestations, molecular detection, histopathological lesions, ultrastructural evidence, and phylogenetic characterization, and in accordance with the criteria of the World Organisation for Animal Health (WOAH) for defining susceptible hosts, this study confirms that T. septentrionalis can be naturally infected by NNV and represents a newly confirmed susceptible host. These findings expand the known host range of NNV and provide essential baseline information for disease surveillance, molecular epidemiology, and biosecurity management in filefish aquaculture. Moreover, the addition of T. septentrionalis to the susceptible species allows further inference of the susceptibility of 109 species in family Monacanthidae to infection with NNV, according to the WOAH standard.