Previously, a recombinant vaccine preparations, based on the i-antigen GDCI3, developed strong specific and non-specific immune responses in vaccinated fish and enhanced their anti-parasite ability, but GDCI3 was cloned and high-level expressed in under a cadmium-inducible metallothionein gene promoter (Jiang et al. Luo et al. 2007), which suggests that vaccination may be a safe and effective method to prevent disease in the future. The external surface including the ciliary membrane of ciliates is usually covered with a variable surface protein known as the immobilization antigen (IAG) (Lee and Kim 2008). IAGs from were firstly extracted with Triton 5′-GTP trisodium salt hydrate X-114 answer according to the method explained by Hatanaka et al. (2007, 2008). Structurally, the so-called IAGs are usually surface antigens, which generally contain a transmission peptide at the N-termini and a glycosylphosphtidylinositol (GPI) modification at the C-termini (Mo et al. 2019). Since surface antigens locate on the surface of parasites and interact directly with their hosts, they are usually considered to be the main dominant antigens and potential vaccine candidates (Mo et al. 2019; Pagheh et al. 2020; Wang and Dickerson 2002). So far, a variety of surface antigens have been 5′-GTP trisodium salt hydrate recognized from (Adura 2016; Bai et al. 2008; Hatanaka et al. 2007, 2008; Huang et al. 2012; Mo et al. 2016)IAGs expressed by are immuno-dominant and have become candidate proteins for vaccine development (Jose Priya et al. 2012; Mo et al. 2019; von Gersdorff Jorgensen et al. 2017). Recently, the orange-spotted grouper (play an important role in acquired protective immunity in fish. However, due to the potential risks to the body, DNA vaccine has not been widely used in aquatic animal disease prevention (Duan et al. 2021; Kayansamruaj et al. 2017). Subunit vaccine, higher security, based on the protective immunogens of could show a similar protective effect. Previously, a recombinant vaccine preparations, based on the i-antigen GDCI3, developed strong specific and nonspecific immune responses in vaccinated fish and enhanced their anti-parasite ability, but GDCI3 was cloned and high-level expressed in under a cadmium-inducible metallothionein gene promoter (Jiang et al. 2021; Mo et al. 2019), which might TFRC cause severe environmental problem in practical use (Bisharyan et al. 2003). Thus, it is of great value to find safe and effective vaccines for the prevention and control of contamination. In our previous study, a gene encoding a surface antigen of was cloned and characterized (GenBank accession number: JF812643). Its coding protein was designated as CiSA-32.6 (Huang et al. 2012). CiSA-32.6 protein contains many antigenic determinants. Herein, a recombinant CiSA32.6t protein (rCiSA32.6t) was prepared through prokaryotic expression system, and the protective effect against challenge was evaluated in large yellow croaker 5′-GTP trisodium salt hydrate (used in the experiment were originally isolated from your gills of the infected during an outbreak of white spot disease in Xiapu, Fujian Province, in 2011 (Sun et al. 2011). And then the parasites were cultured and passaged by using marbled rockfish as hosts in our laboratory (Sun et al. 2011). The healthy (20C30?g/fish) were purchased from Ningde Fufa Fishery Organization Limited, Ningde, Fujian, China. First, fish were cautiously checked to ensure that there was no contamination. During the 2?weeks prior to the experiment, the fish were cultured in 1000-L round plastic tanks and were fed twice daily with commercial pellet feed (~?1.5% of body weight) until eating and other activities were normal. Any feces were siphoned off before feeding. During the study, sand-filtered seawater was used as the culture water, with a pH of 7.9C8.3,?