SNP E4-205 C/T in C-type lectin of Portunus trituberculatus is association with susceptibility/resistance to Vibrio alginolyticus challenge

Abstract

C-type lectins play an important role in innate immunity of invertebrates, especially in defending them against bacterial pathogens. In this study, the full-length genomic DNA of C-type lectin (designated as CTL gene) was isolated from the swimming crab Portunus trituberculatus . The genomic DNA of CTL consists of 1473 bp, containing four exons and three introns. Polymorphisms of the CTL gene were identified to explore their association with susceptibility/resistance to Vibrio alginolyticus infection. Four sites of single nucleotide polymorphisms (SNPs) and one site of insertion/deletion (ins/del) polymorphism were identified in CTL . Three of these SNPs were synonymous and one was non-synonymous. The distribution of these polymorphisms in the susceptible and resistant stocks was identified, according to the survival time after V. alginolyticus challenge. The non-synonymous SNP E4-205 C/T, a dimorphism caused from C to T transition that resulted in a Threonine to Isoleucine substitution at position 152 in the peptide of CTL protein, showed significant difference between the two stocks according to Chi-squared test ( P < 0.05). The SNP site has three genotypes that are C/T, C/C and T/T. The T/T genotype frequency was 6.1% in susceptible stock, and 21.2% in resistant stock, with a significant difference between them ( P = 0.022). Therefore, the T/T genotype was suggested to be associated with increased resistance to V. alginolyticus ( P < 0.05). The presumption was confirmed by another challenge experiment, in which high resolution melting (HRM) analysis was used to analyze the three genotypes distribution. The cumulative mortality of genotype T/T (57.1%) was significantly lower than C/C (82%) and T/C (81.4%) ( P = 0.001). These results suggested that the E4-205 genotype T/T is a potential marker for selection of swimming crabs resistant to V. Alginolyticus challenge. • The full-length genomic DNA of C-type lectin was acquired from Portunus trituberculatus and designated as CTL gene. • The polymorphisms were identified in CTL and the SNP E4-205 C/T showed significant difference between the resistant and susceptible crabs stocks. • The high resolution melting (HRM) analysis method was established to analyze the three genotypes of E4-205 SNP. • The T/T genotype of E4-205 SNP could be a potential gene marker for future selection of swimming crabs with enhanced resistance to V. alginolyticus .