Establishment and comparison of real-time PCR assays to detect Anaplasma msp-2 gene with general TaqMan probs and TaqMan-MGB probe

Objective In order to develop a highly specific and sensitive real-time PCR assay to detect and indentify Anaplasma,the agent of Anaplasmoses. Methods A specific msp-2 outer membrane protein gene of A.phagocytophilum was selected for designing general TaqMan probe and TaqMan MGB probe based on the conserved sequences of msp-2 genes of 39 isolates all over the world. The evaluation of assay included the specificity,lowest detection load (LOD) and stability. Results Similar specificity and reproducibility for the two real-time PCR methods with general TaqMan probe and TaqMan MGB probe were demonstrated. Besides A.phagocytophilum,17 members of Rickettsia and other 8 common pathogens in clinical hospitals were not amplified. The LOD by PCR with TaqMan MGB probe was 10 times than that by PCR with general TaqMan. Conclusion Two highly specific and sensitive real-time PCR assays were developed to test A.phagocytophilum infection during the early phase of illness,but the assay with TaqMan MGB probe is more sensitive than that with TaqMan probe.