干血斑和血清样本用于检测乙型肝炎病毒DNA及耐药突变的比较

Comparison of dried blood spot samples and serum samples for HBV DNA quantification determination and drug resistance mutation

目的：验证干血斑（DBS）样本替代静脉血血清样本用于检测乙型肝炎病毒（HBV）基因诊断的可行性。方法选择慢性乙型肝炎患者100例，血清HBV的病毒载量〉1×10^2拷贝/mL，抽取静脉血同时制备DBS样本和血清样本，进行HBV DNA定量检测，比较DBS与血清样本检测结果的相关性。选取73例基因型主要为B、C型的患者，进行DBS和血清样本基因型检测的比较。同时随机选取56例乙型肝炎患者的DBS和血清样本，采用实验室自建的套式聚合酶链反应（PCR）扩增HBV的逆转录酶（RT）区段，通过geno2pheno数据库进行结果分析，比较二者耐药性的检测结果。结果 DBS样本HBV DNA的检测范围为2-8 log10拷贝/mL，与血清样本HBV DNA的检测结果高度相关（r^2=0.82，P〈0.05），DBS样本HBV DNA定量值比血清样本低1个数量级。2种样本HBV耐药性检测结果的总符合率为95.83%；DBS和血清样本耐药状态的检出率分别为82.14%和85.71%（P〉0.05）。2种样本HBV基因分型结果的符合率为100.00%。结论 DBS样本可用于HBV DNA的定量及耐药性的检测。

Objective To assess the feasibility of dried blood spot（DBS） sample as an alternative to serum sample for hepatitis B virus（HBV）determination. Methods DBS samples and serum samples were prepared through extracting venous blood,and HBV DNA was determined for 100 patients with chronic hepatitis B,whose HBV virus load 〉1×10^2 copies/mL. DBS and serum sample results were compared by correlation analysis. HBV genotypes were analyzed in 73 patients of genotype B and genotype C. A total of 56 DBS and serum samples from patients with hepatitis B were collected randomly. Using an in-house nest polymerase chain reaction（PCR） for reverse transcriptase segment,the data were analyzed using geno2pheno. Results HBV DNA of DBS samples was determined in a range of 2-8 log10 copies/mL. The correlation coefficient（r^2） for HBV DNA between DBS samples and serum samples was 0.82（P〈0.05）. The quantification of HBV DNA determination in DBS samples was 1 log10 lower than that in serum samples. The coincidence rate of the 2 kinds of samples was 95.83%. For DBS and serum samples,the determination rates for drug resistance were 82.14%and 85.71%（P〉0.05）. The coincidence rate of DBS and serum samples for HBV genotype was 100.00%. Conclusions DBS samples can be used in HBV DNAquantification and drug resistance determinations.