Cell-on-hydrogel platform made of agar and alginate for rapid, low-cost, multidimensional test of antimicrobial susceptibility

Abstract

Antimicrobial resistance (AMR) is a rapidly increasing threat to the effective treatment of infectious diseases worldwide. The two major remedies include: (1) using narrow-spectrum antibiotics based on rapid diagnosis; and (2) developing new antibiotics. A key part of both remedies is the antimicrobial susceptibility test (AST). However, the current standard ASTs that monitor colony formation are costly and time-consuming and the new strategies proposed are not yet practical to be implemented. Herein, we report a strategy to fabricate whole-hydrogel microfluidic chips using alginate-doped agar. This agar-based microfabrication makes it possible to prepare inexpensive hydrogel devices, and allows a seamless link between microfluidics and conventional agar-based cell culture. Different from common microfluidic systems, in our system the cells are cultured on top of the device, similar to normal agar plate culture; on the other hand, the microfluidic channels inside the hydrogel allow precise generation of linear gradient of drugs, thus giving a better performance than the conventional disk diffusion method. Cells in this system are not exposed to any shear flow, which allows the reliable tracking of individual cells and AST results to be obtained within 2-3 hours. Furthermore, our system could test the synergistic effect of drugs through two-dimensional gradient generation. Finally, the platform could be directly implemented to new drug discovery and other applications wherein a fast, cost-efficient method for studying the response of microorganisms upon drug administration is desirable.

A Multiplexed, Gradient‐Based, Full‐Hydrogel Microfluidic Platform for Rapid, High‐Throughput Antimicrobial Susceptibility Testing

“…Our device generates linear gradients of antibiotic by means of diffusion, a process verified in our previous work by an “indirect” approach with fluorescence dyes as substitutes to quantify the gradient. Subsequently we used the verified profile for calculating the concentration of molecules across the gradient in actual tests, a method that has been previously used in similar studies .…”

“…As shown in Figure , the design used in our previous work was restricted to a two‐layer device having channels on a single plane only. As such, the actual area of a 2D test, where the two gradients were combined, was limited to part of the whole concentration range.…”

“…By combining the advantages from both the emerging technology of microfluidics and the well‐trusted conventional AST techniques, this platform has great potential to become a much‐improved yet very reliable method for rapid and low‐cost susceptibility testing. The fabrication and operation principles of this platform were based on one of our previously reported hydrogel AST devices, although here we addressed certain shortcomings of our previous work with several innovative designs and techniques being developed. A key innovation is a two‐layer overlaying channel design, which allows multiple test areas to be integrated on a single device and better coverage of concentration in 2D gradients.…”

Modern Tools for Rapid Diagnostics of Antimicrobial Resistance

“…Due to the chip's multidimensionality, several antibiotics, nutrients or immunologic substances could be tested simultaneously. This system is currently under commercialization (Sun et al, 2016 ; Liu et al, 2017a , b ). Weibull et al have presented a nanowell AST device capable of real-time optical reading and growth data analysis.…”

Microfluidics as an Emerging Platform for Tackling Antimicrobial Resistance (AMR): A Review

“…Rapid and multiplexed AST systems were also developed to study bacterial interactions and drugs responses in parallel on a single device while utilising reduced amounts of reagents and samples, and performing analysis in shorter times [20][21]. Sun et al [22][23] developed an agar-based cell-on-hydrogel platform to study bacterial interactions with different concentrations of drugs. The authors generated twodimensional gradients in a stacking device and obtained AST results within 2-3 hours.…”