A 3D-printed capillary tube holder for high-throughput chemotaxis assays

Bacterial chemotaxis is an important behavior to study to understand spatial segregation of species in mixed communities and the assembly of host microbiomes. This is particularly relevant in the rhizosphere, where chemoattraction toward root exudates is an important determinant of plant colonization. However, current methods to screen chemoeffectors are limited in their throughput, creating a barrier to generating comprehensive data sets describing chemotactic profiles for species of interest. Here, we describe a novel 3D-printed capillary tube holder approach, which facilitates up to 384 simultaneous capillary tube chemotaxis assays. We optimized and benchmarked our assay using <i>Escherichia coli</i> K12 and <i>Bacillus subtilis</i> 3610 with known chemoattractants: serine and aspartate. We then tested the threshold concentration of these chemoattractants using our assay and found that we could detect chemoattraction toward concentrations spanning multiple orders of magnitude. In this paper, we describe our high-throughput chemotaxis assay in detail and provide the necessary files for 3D printing the capillary tube holder.IMPORTANCEChemotaxis is an important behavior to study to understand how microbial communities assemble and respond to their environment. Identifying chemoattractants may uncover key targets for microbiome engineering. However, the generation of large data sets describing chemotactic profiles has been limited by a lack of high-throughput tools to quantitatively screen chemotaxis. We designed a 3D-printed assay allowing for up to 384 simultaneous capillary tube chemotaxis assays and validated our method with two different bacterial species. The throughput of our approach is greatly increased by the ability to use lag time as a proxy for cell count. Our approach is easy to use and low cost, effectively lowering the barrier to expanding more comprehensive data sets describing the chemotactic profiles of different bacterial species.