# Acoustic Detection of Vaccine Reactions in Hens for Assessing Anti-Inflammatory Product Efficacy
## 4 Discussion  
This work aims to use PLF technology to characterize the vocalization of hens on a commercial farm before, after, and during their reactions to the ILT vaccine, and to identify and analyze the most important features of automatic detection. A second objective is to quantify the effectiveness of a treatment that reduces the aforementioned vaccination reaction in the animal. Figure 8: Percentage of vaccine reaction detection with Gaussian NB classifier for test and evaluation datasets. Figure 7: Normalized confusion matrix of test classification results with CR dataset. The centroid and vocalNum are stress-related acoustic features. In Figure 4, the centroid feature showed an increase in value on the day of vaccination and throughout the next four days. Furthermore, the maximum peak values of vocalNum were found in the first three days of the vaccination reaction. Higher values in the reaction period suggest the presence of stress caused by manual vaccination with eye drops, and the presence of vaccination side effects. An analysis during daylight, night-time, and throughout the day showed that the night-time interval presented a lower data deviation (Table 1), due to less interference from the surrounding machinery, which allowed for clearer recordings of vocalizations. Furthermore, mean differences between CR and TX groups were found at night, resulting in smaller \(p\)-values (Table 2). The minimum \(p\)-values of the entire analysis were found in the reaction phase block, as follows: 2.41 \(\times\) 10\({}^{-14}\) centroid at night and 1.42 \(\times\) 10\({}^{-6}\) vocalNum at night. These important statistical differences could be due to the effect of the administration of the product to TX, as this was the only difference between the two groups. The higher mean value differences obtained in the reaction block for the centroid feature suggested the possibility of automating vaccine reaction detection. Of all the algorithms tested, Gaussian Naive Bayes performed best, with an F1 score and accuracy above 0.80 (Table 3). The performance of this model is acceptable for vaccine reaction detection. Analyzing the permutation importance of the Gaussian NB algorithm, the two most important features, according to the algorithm, were the centroid, at a value of 0.14, and MFCC4, with a value of 0.10. According to previous publications by several authors of this paper, these features are also used in poultry to indicate food intake, namely the centroid feature [16] and MFCC4, with high concentrations of \(CO_{2}\)[17]. This is in contrast with to the vocalNum parameter, which is also recognized as identifying stress, but only detected a few peak vaccine reaction side effects in hens during the initial three days of administration, making it a less effective feature. The Gaussian NB classification model was fitted with training data and tested with the CR dataset. Table 5a and Figure 7 show that the non-reaction class was better classified, with an F1 score of 0.83, than the reaction class, which had an F1 score of 0.76. The superior classification of the non-reaction class may be attributed to the ease of labeling periods without reactions, as reaction intervals could vary in intensity levels depending on animal welfare, leading to potential misclassifications. When testing the treatment dataset with the trained model of the control group (see Table 5b), it was observed that the TX data was predominantly classified as non-reactive (with an F1 score of 0.99) over the 12 days of the study. The algorithm detected the effectiveness of the product in alleviating respiratory problems, as it classified samples after vaccination (from day 0 onwards) as non-reactive. By calculating the daily detection rate of the vaccine reaction, Figure 8 for the test data shows that the maximum vaccine detection above 75% was found on day 3 after vaccination. Furthermore, on day 5, which was classified as non-reactive, 30% of reactions were still detected, as some side effects of the vaccine were still present, although very reduced.