Nocardia seriolae infection is one of the most serious bacterial diseases in Seriola species in Japanese aquaculture. In this study, the absorption-enhancing effect of citric acid on oral oxytetracycline (OTC) treatment against nocardiosis was evaluated with experimentally infected juvenile yellowtail, Seriola quinqueradiata, followed by serum, kidney and spleen OTC level analysis. When 50 mg/kg body weight (BW) of OTC was administered orally in combination with 1250 mg/kg BW of citric acid, the serum OTC level increased 3-fold at 6 h post single-dose administration, while no effect was observed when the dose of citric acid was set at 50 mg/kg BW. OTC levels in the kidney and spleen were increased 2-fold when 50 mg/kg BW of OTC and 1250 mg/kg BW of citric acid were administered for 5 consecutive days. This coadministration improved the clinical efficacy of oral OTC treatment against nocardiosis by significantly reducing the mortality rate compared to that of the group that received OTC alone and the untreated control group. These results suggest that coadministration of 1250 mg/kg BW of citric acid enhanced OTC absorption and therefore improved the clinical efficacy of oral OTC treatment against nocardiosis.