Electrochemical detection of serum antibodies against Mycobacterium avium subspecies paratuberculosis

Authors: HATATE, KAORU - University Of Tennessee; RICE, J. HUNTER - University Of Tennessee; PARKER, KARSTEN - University Of Tennessee; WU, JAYNE - University Of Tennessee; Turner, Amy; Stabel, Judith; EDA, SHIGETOSHI - University Of Tennessee

Submitted to: Frontiers in Veterinary Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/3/2021
Publication Date: 3/9/2021
Citation: Hatate, K., Rice, J., Parker, K., Wu, J., Turner, A.M., Stabel, J.R., Eda, S. 2021. Electrochemical detection of serum antibodies against Mycobacterium avium subspecies paratuberculosis. Frontiers in Veterinary Science. 8. https://doi.org/10.3389/fvets.2021.642833.
DOI: https://doi.org/10.3389/fvets.2021.642833

Interpretive Summary: Johne's disease is a chronic, debilitating intestinal disorder in cattle characterized by diarrhea, reduced feed intake, weight loss and death. Cattle usually become infected as young calves by ingesting feces containing the causative bacteria. During the subclinical phase of infection the animal may be shedding the organism in its feces without showing any clinical signs of disease. In addition infected cattle can shed the organism into the milk, thereby presenting a potential mode of infection for humans. Infection with Mycobacterium avium subsp. paratuberculosis is difficult to diagnose and therefore to control. Development of accurate and sensitive diagnostic tests is critical for control of this disease. In the present study, the sensitivity and specificity of a a novel diagnostic test to measure serum antibodies was developed using electrochemical detection. The device was more sensitive than current commercial methods for antibody detection. In addition, the detection device is portable so can easily be used cow-side, allowing for rapid and inexpensive diagnosis of infection.

Technical Abstract: Mycobacterium avium subsp. paratuberculosis causes a chronic inflammatory intestinal disease, called Johne’s disease in many ruminants. In the dairy industry, Johne’s disease is responsible for significant economic losses due to decreased milk production and premature culling of infected animals. Test-and-cull strategy in conjunction with risk management is currently recommended for Johne’s disease control in dairy herds. However, current diagnostic tests are labor-intensive, time-consuming and/or too difficult to operate on site. In this study, we developed a new method for the detection of anti-M. paratuberculosis antibodies from sera of M. paratuberculosis-infected animals. M. paratuberculosis antigen-coated magnetic beads were sequentially reacted with bovine serum followed by a horseradish peroxidase-labeled secondary antibody. The reaction of horseradish peroxidase with its substrate was then quantitatively measured electrochemically using a redox-active probe, ferrocyanide. After optimization of electrochemical conditions and concentration of the redox-active probe, we showed that the new electrochemical detection method could distinguish samples of M. paratuberculosis-infected cattle from those of uninfected cattle with greater separation between the two groups of samples when compared with a conventional colorimetric testing method. Since electrochemical detection can be conducted with an inexpensive, battery-operated portable device, this new method may form a basis for the development of an on-site diagnostic system for Johne’s disease.