Evaluation of analytical-sensitivity and -specificity of a commercially available one-step real-time polymerase chain reaction assay kit for the detection of burkholderia mallei

Abstract

Burkholderia mallei (B. mallei) is the etiological agent of glanders, a highly infectious and zoonotic disease of solipeds. Prompt and accurate detection of B. mallei and diagnosis of glanders is important for both, humans and animals. The present study was designed to evaluate the analytical sensitivity and specificity of a commercially available one-step real-time (RT) PCR assay kit (Genekam Biotechnology AG, Germany) for the detection of B. mallei. Furthermore, the comparative evaluation of the analytical sensitivity of an already published fliP RT-PCR protocol and this kit was made using a real-time PCR (RT-PCR) platform: the Mx3000P TM (Stratagene®, Canada). Diagnostic parameters were assessed with a panel of 20 B. mallei, 20 B. pseudomallei, 15 B. mallei related and 10 clinically relevant non-Burkholderia species. Using the one-step RT-PCR on the Mx3000PTM platform, the limit of detection (LOD) was determined as 10 fg. Applying a modified fliP RT-PCR using the Mx3000P platform resulted in a LOD of 100 fg. Authors conclude that the one-step RT-PCR kit is specific for DNA of B. mallei strains. The one-step RT-PCR assay kit is a robust, rapid, reliable, specific and sensitive tool with a fast turnaround time for the specific identification and detection of B. mallei from culture material.

Keywords

• burkholderia mallei
• analytical specificity
• analytical sensitivity
• one-step rt-pcr kit