Comparison of five commercial DNA extraction kits for the recovery of Francisella tularensis DNA from spiked soil samples.

摘要

Francisella tularensis is the etiologic agent of the zoonotic disease tularemia and is thought to be maintained in the environment principally by various terrestrial and aquatic vertebrate animals. The organism is known to persist in water or mud for long periods of time and Francisella-specific DNA has been identified from water and soil. To gain a better understanding of the ecology and epidemiology of F. tularensis, it will be important to further explore its distribution in the environment. Therefore, methods must be established to efficiently extract Francisella-specific DNA from the soil and be able to eliminate potential PCR inhibitors. Thus, we evaluated five commercial DNA extraction kits for their ability to recover F. tularensis-specific DNA from soil samples and eliminate potential PCR inhibitors. The kits evaluated included the Puregene DNA purification kit, QIAamp Stool Mini kit, Epicentre Biotech SoilMaster DNA extraction kit, and the UltraClean and PowerMax soil DNA isolation kits from MoBio. Soil samples were spiked with gamma-irradiated F. tularensis SHU-4 strain (corresponding to a range from 10 to 10(5)CFU). Spiked samples were extracted with each kit and evaluated using a F. tularensis-specific real-time PCR assay and an internal positive control assay that measures the presence of potential PCR inhibitors. DNA extraction using the UltraClean and PowerMax kits resulted in the most consistently positive results at the lowest limit of detection (20 and 100CFU/g soil, respectively) for all soil types tested, suggesting that these kits can provide the most sensitive methods for extracting F. tularensis from environmental soil samples. Processing time and cost were also evaluated.