Abstract. Isnansetyo A, Istiqomah I, Anshary H, Sriwulan S, Yudiati E, Subagiyo S, Arif A, Kartikasari DW. 2022. Identification and antibiotic-resistant properties of Vibrio owensii and V. alginolyticus isolated from the Spermonde Islands, Indonesia. Biodiversitas 23: 5995-6005. The Spermonde Islands are located in Makassar Strait, southern Sulawesi, Indonesia, and have attracted the attention of researchers for decades because of their mega biodiversity and abundance of bioresources. However, no study has evaluated the potential mariculture diseases in this area. The present study assessed the potential bacterial fish diseases based on the current status of mariculture in the Spermonde Islands. The samples were collected from three marine aquaculture sites at Barrang Caddi and the Samaloa Islands. Vibrio was isolated on Thiosulfate-citrate-bile-sucrose (TCBS) agar medium. The most common bacterial diseases were observed by bacterial isolation, morphological and biochemical observations, and molecular identification based on 16S rDNA sequence. The microbial, physical, and chemical water qualities at Barrang Caddi and Samaloa Island were suitable for marine aquaculture. Although cultured species in this area did not exhibit gross disease signs, potential opportunistic pathogens of Vibrio were found. The 16S rDNA sequences analysis indicated that the bacterial isolates closed to Vibrio owensii, V. alginolyticus, and V. neocaledonius. Multi-drug resistant V. owensii was identified as a potential pathogen in marine aquaculture. This is the first report on V. owensii isolated from Indonesian marine waters, particularly from the Spermonde Islands. V. alginolyticus was another potential opportunistic pathogen in this area. These results gave the alarm to develop countermeasure methods for potential diseases to minimize the possible outbreak of vibriosis.