Fig. 2

The tracking software successfully extracts trajectories of multiple animals at a time despite background noise. a Principal component analysis over all detected animal instances in a single frame shows that worm entities are faithfully segregated from background entities. For example, during long temporal recordings, drop condensation on the lid and trail marks contribute to the identification of approximately 2000 erroneous entities per frame. The number of genuine animal entities, however, remains constant (around 100) throughout the movie (see also Additional file 6: Figure S1). Thus, the software successfully learns the crucial features to detect animal entities and discards background noise. b Accuracy of worm detection as a function of the training size. The precision (fraction of actual worms out of total entities classified as worms) reaches nearly 90%, and recall (out of all worms genuinely found on the plate, how many are indeed classified as worms) reaches approximately 85%. Fscore is the harmonic average of the two parameters (precision and recall); as few as 50 worms are sufficient to reach these accuracies (see Methods for details how these parameters were obtained). c Implementing a variant of a Kalman predictor significantly enhances track length. Shown are the lengths (in seconds) of all tracks without implementing the predictor (left), and when using the predictor (right). The y-axis and x-axis denote the start and end time, respectively, of the period the tracks lengths were averaged on. Shown are all tracks identified throughout the movie. Significant longer tracks are extracted if implementing the predictor (80 ± 0.54 [s] compared to without the predictor 58 ± 0.3 [s], in the period of the experiment that yielded the longest tracks, P < 0.001). d There is a trade-off between the number of worms loaded on the assay plate and the average length of detected tracks. The more worms loaded the shorter are the continuously detected tracks. Up to 200 worms can be assayed simultaneously in a field of view of 4 × 4 cm if not requiring extraction of long continuous tracks