Rapid efficacy testing of cattle footbath disinfectants by flow cytometry: a laboratory-validated screening method complementary to EN 1656

Introduction Disinfectants are essential for preventing infectious diseases in livestock, particularly in cattle footbath applications. Conventional culture-based methods for assessing bactericidal efficacy are reliable but time-consuming, leaving the need unmet for rapid laboratory-based screening approaches. Material and Methods Thirteen commercially available cattle footbath disinfectants were evaluated using the EN 1656 dilution–neutralisation method and flow cytometry (FCM) with propidium iodide (PI) staining. A binary logistic regression model was developed to relate the proportion of PI + cells to bactericidal efficacy. Results Disinfectants containing iodine or iodide, organic acids, hydrogen peroxide, quaternary ammonium compounds, glutaraldehyde or alcohol showed high bactericidal efficacy, whereas those with zinc sulphate were ineffective. The predictive model demonstrated good performance (AUC = 0.787) and identified an optimal threshold of approximately 80% PI + cells, yielding 85.47% sensitivity and 76.92% specificity. Agreement between FCM and the EN 1656 method was high, with 7.69% inconsistent classifications. Conclusion Flow cytometry represents a rapid and efficient laboratory-based screening method with the potential to complement culture-based approaches for evaluating bovine footbath disinfectant efficacy.