Major histocompatibility complex (MHC) class Ⅱ B molecules play an important role in the adaptive immune response in fish. Previous study has reported that two highly polymorphic class ⅡB genes, Cyse-DAB and Cyse-DBB exist in half-smooth tongue sole (Cynoglossus semilaevis). In this study, the polymorphism within exon 2 of the class Ⅱ B genes following bacterial challenge was evaluated. Two hundred C. semilaevis individuals were injected intraperitoneally with Vibrio anguillarum. Muscle tissue from the first 20 dead and 20 of the survivors was collected for genotyping. Sixty alleles from the 40 individuals were isolated, of which 32 belonged to Cyse-DAB and 28 belonged to Cyse-DBB. The rate of dN (non-synonymous substitution) was higher than that of dS (synonymous substitution) in the PBRs (peptide binding residues) of both class Ⅱ B genes. Conversely, the rate of dS was higher than dN in the non-PBRs and the complete exon 2 sequence. Thus, the results suggest that positive selection has occurred in the PBRs and purifying selection in the non-PBRs and exon 2. Thirteen class Ⅱ B alleles were used to study the association between alleles and resistance to infection. Though not significant, alleles Cyse-DAB*0601, Cyse-DAB*0706, and Cyse-DBB*0101, Cyse-DBB*1301 were only found in surviving individuals and may represent alleles that have resistance against V. anguillarum infection. Alleles Cyse-DAB*0701 and Cyse-DAB*1301 were significantly more prevalent in dead individuals than in surviving ones and may represent alleles that are associated with increased susceptibility to V. anguillarum infection. Major histocompatibility complex (MHC) class II B molecules play an important role in the adaptive immune response in fish. Previous study has reported that two highly polymorphic class IIB genes, Cyse-DAB and Cyse- DBB exist in half-smooth tongue sole (Cynoglossus semilaevis). In this study, the polymorphism within exon 2 of the class II B genes following bacterial challenge was evaluated. Two hundred C. semilaevis individuals were injected intraperitoneally with Vibrio anguillarum. Muscle tissue from the first 20 dead and 20 of the survivors was Sixty alleles from the 40 individuals were isolated, of which 32 belonged to Cyse-DAB and 28 belonged to Cyse-DBB. The rate of dN (non-synonymous substitution) was higher than that of dS (synonymous substitution) in the PBRs (peptide binding residues) of both class II B genes. Conversely, the rate of dS was higher than dN in the non-PBRs and the complete exon 2 sequences. Thus, the results suggest that positive sel ection has occurred in the PBRs and purifying selection in the non-PBRs and exon 2. Thirteen class II B alleles were used to study the association between alleles and resistance to infection. Though not significant, alleles Cyse-DAB * 0601, Cyse-DAB * 0706, and Cyse-DBB * 0101, Cyse-DBB * 1301 were only found in surviving individuals and may represent alleles that have resistance against V. anguillarum infection. Alleles Cyse-DAB * 0701 and Cyse-DAB * 1301 were significantly more prevalent in dead individuals than in surviving ones and may represent alleles that are associated with increased susceptibility to V. anguillarum infection.