challenge test was performed by immersing the surviving larvae from the growth experiment in a Streptococcus agalactiae suspension. Pathogenic S. agalactiae (isolate no. NK1) was kindly provided by the Agency for Marine Affairs and Fisheries Research and Development of the Ministry of Fisheries and Marine Affairs, Indonesia. About 10 μL of stock culture was inoculated into Brain Heart Infusion broth (Criterion C5141, Hardy Diagnostics, California, USA) and incubated for 24 h at 30 °C under constant agitation (140 rpm). After culture, the cells were washed twice in sterile saline. Twelve units of 2 L plastic tanks were filled with 1.5 L of chlorinated and aerated freshwater and added with S. agalactiae culture at a density of 107 CFU/mL. From three out of four growth test tanks per treatment, ten healthy fish were selected and transferred to an immersion tank. After 5 days, the survival of the larvae was determined.