Evaluation of ELISA and qPCR assays for detection of Mycoplasma bovis in milk from ruminants

Abstract: Mycoplasma bovis (M. bovis) infections are difficult to diagnose and treat, causing severe economic losses to farms with infected cows. Eurofins Milk Testing Sweden AB screens cow’s milk using qPCR (quantitative Polymerase Chain Reaction) to detect M. bovis infections. Five performance characteristics were chosen to evaluate if ELISA (Enzyme-Linked Immunosorbent Assay) could replace the current screening method; limit of detection (LOD), repeatability and intermediate precision, selectivity and matrix effects and PCR inhibition. The comparison between the two methods was complicated by the fact that qPCR test for the presence of M. bovis DNA (qPCR), while the ELISA test for the presence of anti-M. bovis antibodies. In addition, the analysis methods were compared regarding health and environmental aspects. Based on the two assays performance characteristics the qPCR outperformed the ELISA regarding the LOD and the matrix effects and PCR inhibition, while the ELISA surpassed the qPCR with respect to the repeatability and intermediate precision. Regarding the selectivity, both methods performed equally well. The qPCR includes extra steps compared to the ELISA and hence needs additional solutions, this contributes to that the qPCR is considered to be less health and environmentally friendly. The ELISA is also cheaper and more user friendly than the qPCR. Comparing of testing for antibodies and testing for DNA is complex, there is no guarantee that either antibodies or M. bovis cells are present in the milk at the time of testing. Based on the results in this degree project it is suitable to use the two methods in parallel; ELISA as the primary screening method and then to verify positive results with qPCR. Except for when a bacteriological culture is to be verified, then qPCR is suitable since the presence of M. bovis cells is to be tested.