DefinePK

Abstract

Current diagnostic methods for clinical and subclinical mastitis typically involve a bacterial culture phase, which can be time-consuming and requires specialized laboratory facilities. To overcome these limitations, our study optimized a loop-mediated isothermal amplification (LAMP) assay for the detection of pathogens causing mastitis, including Escherichia coli (E. coli), Salmonella species, and Staphylococcus aureus (S. aureus), in milk samples collected from ruminants. Detection of S. aureus, Strep. agalactiae, and E. coli in milk samples by multiplex PCR was 97(22.6%), 54(12. 6%), and 69(16.1%), respectively. Results showed that the LAMP reaction was completed within 30min at 65°C. The positive reaction showed a green color, and the negative reaction remained unchanged under UV light. The sensitivity of multiplex PCR assay showed a high sensitivity for most pathogens, ranging from 86.9 to 98.1%, nearly similar to the LAMP assay, which ranged from 89.7 to 97.03%. The specificity of multiplex PCR and LAMP assays ranged from 92.7 to 98.7%. In conclusion, the LAMP assay is a promising method for fast on-site assays that are simple, high sensitivity, high specificity, and rapid for detecting mastitis-causing bacteria.