This study developed a new method for forensic saliva identification using three oral bacteria, Streptococcus salivarius, Streptococcus sanguinis, and Neisseria subflava, combined with a real-time polymerase chain reaction (RT-PCR) system we called OB mRT-PCR. Analytical sensitivity results showed that the target bacteria were amplified at 10²-10⁷ copies/reaction, and analytical specificity was assessed using 24 other viruses, bacteria, and protozoa. To evaluate the OB mRT-PCR kit for forensic applications, saliva from 140 Korean individuals was tested, and at least two target bacteria were detected in all the samples. Additional studies on non-saliva samples demonstrated the specificity of the kit. Comparison of the kit with two conventional saliva test methods, the SALIgAE and RSID-Saliva assays, indicated that it was more sensitive and applicable to saliva samples in long-term storage (up to 14 weeks). Additionally, through amplification of mock forensic items and old DNA samples (isolated without lysis of the bacterial cells, regardless of their Gram-positivity), we found that the kit was applicable to not only saliva swabs, but also DNA samples. We suggest that this simple RT-PCR-based experimental method is feasible for rapid on-site analysis, and we expect this kit to be useful for saliva detection in old forensic DNA samples.

中文翻译：

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基于实时PCR的口腔细菌快速检测可用于唾液的法医鉴定。

这项研究开发了一种使用三种口腔细菌，唾液链球菌，血链球菌和亚氏奈瑟氏菌的法医唾液鉴定的新方法，结合了我们称为OB mRT-PCR的实时聚合酶链反应（RT-PCR）系统。分析灵敏度结果表明目标细菌在10 ² -10 ⁷处被扩增复制/反应，以及使用其他24种病毒，细菌和原生动物评估分析特异性。为了评估OB mRT-PCR试剂盒的法医应用，对140名韩国人的唾液进行了测试，并且在所有样品中至少检测到两种靶细菌。对非唾液样品的其他研究证明了该试剂盒的特异性。将该试剂盒与两种常规唾液测试方法（SALIgAE和RSID-Saliva分析）进行比较，表明该试剂盒更灵敏，适用于长期保存（长达14周）的唾液样品。此外，通过扩增模拟取证物品和旧的DNA样本（无论细菌细胞是否革兰氏阳性，都无需裂解即可分离），我们发现该试剂盒不仅适用于唾液拭子，还适用于DNA样本。