Abstract This project aims to develop WormInvestigator™, a novel, highly innovative system for performing automated, high-throughput and longitudinal studies of the behavior of C. elegans worms freely moving and socially interacting on agar plates (hereafter: "freely moving worms") across multiple time points over extended times (e.g., multiple days) with repeated measures designs. Work in Phase I will focus on demonstrating feasibility of our novel, patent pending, WormRecognizer™ technology – the ability to perform automatic, image-based identification of individual C. elegans worms within a group of freely moving worms ("digital tagging of freely moving worms"). Work in Phase II will focus on creating the full functionality of WormInvestigator for the commercial release. The innovation inherent in WormRecognizer will serve as the basis for enabling a game- changing innovation in the field – the ability to perform high throughput longitudinal, repeated measures design analyses of locomotion and other behavior of freely moving C. elegans worms from discrete, non-continuous video sequences. Compared to study designs that have independent groups repeated measures designs offer more statistical power and the possibility to track an effect over time. Specifically, repeated measures designs for analyzing locomotion and other behavior of freely moving worms will allow researchers to definitively assess the likelihood that a particular behavior is associated with a prior behavior, which is impossible without repeated measures designs or impractical continuous imaging and tracking under constant illumination. WormRecognizer will leverage the Deep Convolutional Neural Network (CNN) architecture to perform automatic identification of the tracks of the same worm in videos of groups of freely moving worms recorded at different time points; encouraging pilot data were generated during preparation of this application. C. elegans is increasingly used as a model organism in research focusing on brain mechanisms underlying complex behaviors and pathological alterations thereof, including research into neurodevelopment, Alzheimer's disease, autism, schizophrenia and traumatic brain injury. Thus, WormInvestigator will enable significant advancements in various mental neuroscience applications that use C. elegans as a model organism. Specifically, the fact that C. elegans express many of the neurotransmitters and associated receptors that are found in higher eukaryotes, including humans, makes C. elegans highly attractive for the (high throughput) screening of next generation therapeutics for mental diseases such as Alzheimer's disease, as well as for disorders that rely on neurotransmitter release modulation such as next generation treatments for schizophrenia. We will perform extensive feasibility studies, product validation and usability studies of WormInvestigator in close collaboration with expert neuroscientists. Market research performed during preparation of this application indicated that WormInvestigator will expand the use of C. elegans as a model organism to many laboratories that do not currently use them. A competing technology is not available. We anticipate the global market size for WormInvestigator to be more than 300 systems.

Narrative Performing longitudinal studies and repeated measures design analyses in both short-term and long-term experimental assays focusing on the analysis of locomotion and other behavior of multiple C. elegans worms holds the promise of profound progress in next-generation neuroscience studies such as understanding neurodevelopmental, neuropsychiatric and neurodegenerative disorders, as well as aging research, drug discovery and toxicology. Our proposed product will be a transformative technology, using new analytical methods based on artificial intelligence algorithms that, for the first time, will enable researchers to perform these data-rich longitudinal studies and, thus, repeated measures design analyses in assays using C. elegans as a model organism. This system will allow researchers to make new discoveries based on new studies that are currently not feasible, ultimately providing the basis for developing novel treatments to prevent and fight complex brain diseases.