Validation of a commercial adenosine triphosphate luminometry swab and the Petrifilm to assess the cleanliness of automatic milk feeders for preweaning calves.

The main objective of this observational study was to assess the sensitivity and specificity of a commercial ATP luminometry swab (AquaSnap Total; Hygiena, CA, USA) and an on-farm bacteriological culture (Petrifilm; 3M aerobic colony count; MN, USA) in identifying bacterial presence in automated milk feeders (AMFs) for calves. The secondary purpose of this study was to examine the association between season, AMF design (Förster-Technik base models, Urban models with non-return valves, and Holm & Laue models with non-return valves and a peristaltic pump), and the type of equipment piece (automatically washed by the system, tube-like parts, other small parts, and nipples) and bacterial presence. Twenty-four dairy farms from Centre-du-Québec (Canada) using AMFs participated in the project. Only one farm used an optional hygiene feature. Each farm was visited 6 times from March 2024 to February 2025. Sterile physiological water was used to rinse each piece of equipment, and the residual liquid was poured in a sterile collection tube. All samples were analyzed with the luminometer (LUM, measured in relative light units; RLU), which detects ATP, and with the Petrifilm (BCT, measured in cfu/mL). We used BCT thresholds of > 10,000, > 20,000, > 50,000, and > 100,000 cfu/mL and selected corresponding thresholds for LUM that maximized sensitivity and specificity. Sensitivity and specificity, with their 95% Bayesian credible intervals (BCIs), were calculated for both tests with Bayesian latent class models assuming conditional dependence between the tests. Risk factors for BCT > 20,000 and > 100,000 cfu/mL were assessed through odds ratios (ORs) with 95% BCIs obtained with Bayesian logistic regression models. A total of 980 samples were collected in winter (n = 183; 18.7%), spring (n = 265; 27.0%), summer (n = 271; 27.7%), and fall (n = 261; 26.6%) from the milk containers (n = 211; 21.6%), nipples (n = 255; 26.0%), tube-like parts (n = 245; 25.0%), and other small parts (n = 269; 27.4%). The median results were 40,000 cfu/mL for BCT (interquartile range = 3,775-340,000) and 334 RLU for LUM (interquartile range = 75-2,184). The correlation between LUM and BCT was strong (rS = 0.83, 95% CI = 0.81-0.86). The LUM thresholds of > 200, > 250, > 350, and > 450 RLU for LUM corresponded to the > 10,000, > 20,000, > 50,000, and > 100,000 cfu/mL thresholds, respectively. The BCT and LUM methods had similar sensitivity and specificity at these thresholds. For example, the sensitivity of BCT and LUM were 0.86 (95% BCI = 0.82-0.90) and 0.81 (95% BCI = 0.75-0.87), and their specificity were 0.80 (95% BCI = 0.74-0.86) and 0.80 (95% BCI = 0.73-0.87) for the > 250 RLU and > 20,000 cfu/mL thresholds, respectively. Samples taken in spring (OR = 1.86; 95% BCI = 1.15-2.74), summer (OR = 1.97; 95% BCI = 1.21-2.87), and fall (OR = 1.69; 95% BCI = 1.09-2.51) had higher odds of being > 450 RLU, as measured by LUM, than the ones collected in winter. Samples taken from designs with non-return valves and a peristaltic pump had lower odds of being > 450 RLU than samples from base models (OR = 0.26; 95% BCI = 0.07-0.59). Manually washed parts had higher odds of being > 450 RLU (nipples: OR = 7.18; 95% BCI = 4.29-10.7; tube-like parts: OR = 3.70; 95% BCI = 2.26-5.46; other small parts: OR = 2.66; 95% BCI = 1.53-3.99) than automatically washed parts (milk container). These results can help investigate hygiene problems of feeding equipment in dairy calf barns.