College of Animal Science and Technology, Northwest A&F University, Shaanxi Key Laboratory of Molecular Biology for Agriculture, Yangling 712100, China.
Fish Shellfish Immunol. 2011 Jan;30(1):45-50. doi: 10.1016/j.fsi.2010.09.004. Epub 2010 Sep 30.
Toll-like receptors (TLRs) have emerged as crucial sensors of invading microbes through recognition of pathogen-associated molecular patterns (PAMPs) in viruses, bacteria, fungi and protozoa. The polymorphisms in TLRs are closely associated with the resistance to pathogen infections. TLR3 involved in the recognition of double stranded RNA in humans, mice, pigs and fishes. In present study, the nucleotide sequence polymorphisms of TLR3 gene in grass carp (Ctenopharyngodon idella) (CiTLR3) were investigated to explore their association with susceptibility/resistance to grass carp reovirus (GCRV). Twelve single nucleotide polymorphisms (SNPs) and an ins/del mutation were detected in the complete sequence of CiTLR3. Ten of them were sited in the non-coding region. The two SNPs in exon were synonymous mutation. The ins/del mutation was coincidental at the start codon. To investigate the association between the polymorphism and the susceptibility/resistance to GCRV, we selected eight SNPs in the non-coding region and analyzed the genotype and allele distribution in susceptible and resistant groups with PCR-RFLP. The statistical results indicated that only -764 G/T was significantly associated with the resistance of grass carp to GCRV both in genotype (P = 0.040) and allele (P = 0.025). Linkage disequilibrium analysis revealed -543 A/G, -488 G/T, 4116 G/T and 4731 C/T were linkage disequilibrium, and haplotype analysis revealed that haplotype GTTT frequency in susceptible group was significantly higher than that in the resistant group (OR = 2.01, 95% CI 0.996-4.043, P = 0.049). To further confirm the correlation, an additional infection experiment was carried out. The mortality in the -764 GG genotype individuals was significantly lower than GT genotype (OR = 0.208, 95% CI 0.067-0.643, P = 0.011) and TT genotype (OR = 0.183, 95% CI 0.052-0.648, P = 0.015). All the results indicated that haplotype GTTT and genotype -764 TT and -764 GT individuals were susceptible to GCRV while -764 GG was resistant, which could be the optional markers for selective breeding for the GCRV-resistant grass carp in future.
Toll 样受体 (TLRs) 通过识别病毒、细菌、真菌和原生动物中的病原体相关分子模式 (PAMPs)，已成为入侵微生物的关键传感器。TLRs 的多态性与对病原体感染的抗性密切相关。TLR3 参与人类、小鼠、猪和鱼类中双链 RNA 的识别。本研究旨在探讨草鱼 (Ctenopharyngodon idella) TLR3 (CiTLR3) 基因的核苷酸序列多态性与草鱼呼肠孤病毒 (GCRV) 易感性/抗性的关系。在 CiTLR3 的完整序列中检测到 12 个单核苷酸多态性 (SNP) 和一个插入/缺失突变。其中 10 个位于非编码区。外显子中的两个 SNP 为同义突变。插入/缺失突变与起始密码子巧合。为了研究多态性与 GCRV 易感性/抗性的关系，我们选择了非编码区的 8 个 SNP，并通过 PCR-RFLP 分析了易感组和抗性组的基因型和等位基因分布。统计结果表明，-764 G/T 与草鱼对 GCRV 的抗性显著相关，基因型 (P = 0.040) 和等位基因 (P = 0.025) 均有显著差异。连锁不平衡分析显示-543 A/G、-488 G/T、4116 G/T 和 4731 C/T 存在连锁不平衡，单倍型分析显示易感组 GTTT 单倍型频率明显高于抗性组 (OR = 2.01，95%CI 0.996-4.043，P = 0.049)。为了进一步证实相关性，进行了额外的感染实验。-764 GG 基因型个体的死亡率明显低于 GT 基因型 (OR = 0.208，95%CI 0.067-0.643，P = 0.011) 和 TT 基因型 (OR = 0.183，95%CI 0.052-0.648，P = 0.015)。所有结果表明，GTTT 单倍型和基因型-764 TT 和 -764 GT 个体易感染 GCRV，而-764 GG 个体则具有抗性，这可能是未来草鱼 GCRV 抗性选择性育种的可选标记。
