Tilapia tilapinevirus or tilapia lake virus (TiLV) is an emerging virus that inflicts significant mortality on farmed tilapia globally. Previous studies reported detection of the virus in multiple organs of the infected fish; however, little is known about the in‐depth localization of the virus in the central nervous system. Herein, we determined the distribution of TiLV in the entire brain of experimentally infected Nile tilapia. In situ hybridization (ISH) using TiLV‐specific probes revealed that the virus was broadly distributed throughout the brain. The strongest positive signals were dominantly detected in the forebrain (responsible for learning, appetitive behaviour and attention) and the hindbrain (involved in controlling locomotion and basal physiology). The permissive cell zones for viral infection were observed mostly to be along the blood vessels and the ventricles. This indicates that the virus may productively enter into the brain through the circulatory system and widen broad regions, possibly through the cerebrospinal fluid along the ventricles, and subsequently induce the brain dysfunction. Understanding the pattern of viral localization in the brain may help elucidate the neurological disorders of the diseased fish. This study revealed the distribution of TiLV in the whole infected brain, providing new insights into fish–virus interactions and neuropathogenesis.

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A rapid increase in multi‐drug‐resistant (MDR) bacteria in aquaculture highlights the risk of production losses due to diseases and potential public health concerns. Previously, we reported that ozone nanobubbles (NB‐O3) were effective at reducing concentrations of pathogenic bacteria in water and modulating fish immunity against pathogens; however, multiple treatments with direct NB‐O3 exposures caused alterations to the gills of exposed fish. Here, we set up a modified recirculation system (MRS) assembled with an NB‐O3 device (MRS‐NB‐O3) to investigate whether MRS‐NB‐O3 (a) were safe for tilapia (Oreochromis niloticus), (b) were effective at reducing bacterial load in rearing water and (c) improved survivability of Nile tilapia following an immersion challenge with a lethal dose of MDR Aeromonas hydrophila. The results showed no behavioural abnormalities or mortality of Nile tilapia during the 14‐day study using the MRS‐NB‐O3 system. In the immersion challenge, although high bacterial concentration (~2 × 107 CFU/ml) was used, multiple NB‐O3 treatments in the first two days reduced the bacteria between 15.9% and 35.6% of bacterial load in water, while bacterial concentration increased from 13.1% to 27.9% in the untreated control. There was slight up‐regulation of non‐specific immune‐related genes in the gills of the fish receiving NB‐O3 treatments. Most importantly, this treatment significantly improved survivability of Nile tilapia with relative percentage survival (RPS) of 64.7% – 66.7% in treated fish and surviving fish developed specific antibody against MDR A. hydrophila. In summary, the result suggests that NB‐O3 is a promising non‐antibiotic approach to control bacterial diseases, including MDR bacteria, and has high potential for application in recirculation aquaculture system (RAS).

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