Establishment of a visual recombinase polymerase amplification assays for Mycoplasma bovis
The aim was to establish an efficient and rapid visual recombinase polymerase amplification(LFD-RPA)clinical diagnostic method for Mycoplasma bovis.With the uvre gene sequence of Mycoplasma bovis as the target gene,specific primers and probes are designed and validated by sensitivity,specificity,repeatability and clinical sample detection test through screening of primers and probes and optimizing the reaction conditions.The results showed that the optimal primers for LFD-RPA of Mycoplasma bovis established in this test were F2/R2,and the opti-mal reaction condition was 39 ℃ for 25 min;the detection sensitivity was up to 2.08 copies·μL-1,100 times of common PCR;there was no cross reaction with Mycoplasma synoviae,Salmonella,Mycoplasma ovipneumoniae,Pas-teurella multocida,Staphylococcus aureus,Streptococcus agalactiae and Clostridium perfringens;the repeatability was stable;50 nasal swab samples were detected,the positive rate was 26％,and the coincidence rate with domestic in-dustry standard PCR detection methods was 89.6％.In this study,the LFD-RPA detection method for Mycoplasma bovis was successfully established,which has the advantages of easy operation,rapidity,high efficiency,and sensi-tivity,and provides technical support for the rapid clinical diagnosis of Mycoplasma bovis.
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