College of Animal science and Animal medicine, Inner Mongolia Agriculture University, Huhhot 010018, China.
J Microbiol Methods. 2010 Dec;83(3):326-9. doi: 10.1016/j.mimet.2010.09.013. Epub 2010 Sep 28.
This study characterized the dynamic distribution of bacteria in the blood of pigs infected with Streptococcus suis serotype 2 using specific primers and a TaqMan probe designed to amplify the highly conserved S. suis serotype 2 glutamate dehydrogenase (GDH) gene sequences. Gene copy numbers were used to determine the concentration of bacteria in the blood of infected pigs over time using established TaqMan real-time quantitative PCR methodologies (RT-qPCR). The results showed that the detection limit of the RT-qPCR was 10 GDH gene copies. The advantages of utilizing this approach are the high levels of specificity, sensitivity and reproducibility. Bacteria were detected in the blood of infected pigs after 24 h post infection and S. suis GDH gene copies in the experimental group were highest (10(4.15)) on day 7 post infection. Data presented in this report demonstrate that the TaqMan RT-qPCR detection method can be used to characterize the dynamic changes occurring during S. suis serotype 2 blood infections in Bama minipigs thereby facilitating research associated with defining pathogenic mechanisms associated with this organism.
本研究采用特异性引物和 TaqMan 探针,对猪链球菌 2 型谷氨酸脱氢酶(GDH)基因序列进行扩增,对感染猪链球菌 2 型的猪血液中细菌的动态分布进行了特征描述。采用建立的 TaqMan 实时荧光定量 PCR(RT-qPCR)方法,根据细菌基因拷贝数的变化,可确定感染猪血液中细菌浓度随时间的变化。结果表明,RT-qPCR 的检测限为 10 个 GDH 基因拷贝。该方法的优点是特异性、灵敏度和重现性高。感染后 24 h 即可在感染猪血液中检测到细菌,感染后第 7 天实验组猪链球菌 GDH 基因拷贝数最高(10(4.15))。本研究结果表明,TaqMan RT-qPCR 检测方法可用于描述巴马小型猪猪链球菌 2 型菌血症过程中的动态变化,从而为研究该病原体相关的致病机制提供便利。