Abstract
We report a microfluidic device for automated sorting and cultivation of chemotactic microbes from pure cultures or mixtures. The device consists of two parts: in the first part, a concentration gradient of the chemoeffector was built across the channel for inducing chemotaxis of motile cells; in the second part, chemotactic cells from the sample were separated, and mixed with culture media to form nanoliter droplets for encapsulation, cultivation, enumeration, and recovery of single cells. Chemotactic responses were assessed by imaging and statistical analysis of droplets based on Poisson distribution. An automated procedure was developed for rapid enumeration of droplets with cell growth, following with scale-up cultivation on agar plates. The performance of the device was evaluated by the chemotaxis assays of Escherichia coli (E. coli) RP437 and E. coli RP1616. Moreover, enrichment and isolation of non-labelled Comamonas testosteroni CNB-1 from its 1:10 mixture with E. coli RP437 was demonstrated. The enrichment factor reached 36.7 for CNB-1, based on its distinctive chemotaxis toward 4-hydroxybenzoic acid. We believe that this device can be widely used in chemotaxis studies without necessarily relying on fluorescent labelling, and isolation of functional microbial species from various environments.
Highlights
A recent study has reported the use of phase contrast microscopy[14] in visualization of chemotaxis in flow-free gradients, it is still challenging to evaluate chemotaxis without labelling of cells
We introduce a microfluidic system which interfaces parallel-flow based chemotactic sorting with droplet microfluidics
Chemotactic response was assessed by counting of droplets containing cell growth, following with statistical analysis based on Poisson distribution
Summary
A recent study has reported the use of phase contrast microscopy[14] in visualization of chemotaxis in flow-free gradients, it is still challenging to evaluate chemotaxis without labelling of cells. Single-cell isolation in droplets has been validated that it can improve recovery of slow-growing species[32,33]. We introduce a microfluidic system which interfaces parallel-flow based chemotactic sorting with droplet microfluidics. Steady-state gradient for chemotactic sorting and high throughput droplet encapsulation for single cell cultivation are incorporated in a seamless and automated manner. The use of droplets allows straightforward enumeration of non-labelled cells for quantitative evaluation of chemotaxis, and enable enrichment and rapid recovery of species for further studies
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