微滴数字聚合酶链反应检测婴儿唾液中变形链球菌和远缘链球菌的研究

目的 使用微滴数字PCR检测婴儿口腔内变形链球菌和远缘链球菌,评价其可行性.方法 以变形链球菌c型和远缘链球菌ATCC 27607标准菌株基因设计引物,对123名5~8个月婴儿唾液样本分别进行微滴数字PCR、巢氏PCR和常规PCR反应,检测婴儿唾液中变形链球菌和远缘链球菌.结果 微滴数字PCR定性、定量检测变形链球菌和远缘链球菌标准菌,1×10-4 ng/μl靶细菌的拷贝数达到4.8×104和3.6×104,灵敏度可达0.01%;巢氏PCR检测灵敏度为0.01%;常规PCR检测灵敏度为0.1%.用微滴数字PCR、巢氏PCR和常规PCR检测婴儿唾液中变形链球菌检出率分别为78.9%、33.3%和6.5%;远缘链球菌检出率分别为11.4%、4.1%和0,微滴数字PCR检出率明显高于常规PCR和巢氏PCR(P<0.05).结论 微滴数字PCR能早期检测婴儿口腔中变形链球菌和远缘链球菌,该方法具有较高的灵敏性和特异性,有广泛的临床应用前景.

The use of droplet digital polymerase chain reaction for detection of Streptococcus mutans and Streptococcus sobrinus in infants saliva

Objective To evaluate the feasibility of using droplet digital polymerase chain reaction for the detection of Streptococcus mutans in infants saliva. Methods The primers were designed based on the gene of S. mutans c type and S. sobrinus ATCC27607 standard strains. The saliva of 123 infants aged from 5-8 months was examined for S. mutans and S. sobrinus by droplet digital PCR, nest PCR and conventional PCR reaction. Results The copy number of 1 × 10-4 ng/μl target bacteria reached 4. 8 × 104 and 3. 6 × 104 with a sensitivity of 0. 01% by ddPCR quantitative detection of S. mutans and S. sobrinus. The sensitivity of nest PCR was 0. 01% and of the conventional PCR 0. 1%. The detection rate of S. mutans by dd PCR, nest PCR and conventional PCR was 78. 9%, 33. 3% and 6. 5%, respectively, and the detection rate of S. sobrinus was 11. 4%, 4. 1% and 0% respectively. The droplet digital PCR detection rate was significantly higher than the conventional PCR and nest PCR (P <0. 05). Conclusion Droplet digital PCR can detect S. mutans and S. sobrinus in infants saliva with high sensitivity and specificity.