RESUMO
Microorganism sensing of and responding to ambient chemical gradients regulates a myriad of microbial processes that are fundamental to ecosystem function and human health and disease. The development of efficient, high-throughput screening tools for microbial chemotaxis is essential to disentangling the roles of diverse chemical compounds and concentrations that control cell nutrient uptake, chemorepulsion from toxins, and microbial pathogenesis. Here, we present a novel microfluidic multiplexed chemotaxis device (MCD) which uses serial dilution to simultaneously perform six parallel bacterial chemotaxis assays that span five orders of magnitude in chemostimulant concentration on a single chip. We first validated the dilution and gradient generation performance of the MCD, and then compared the measured chemotactic response of an established bacterial chemotaxis system (Vibrio alginolyticus) to a standard microfluidic assay. Next, the MCD's versatility was assessed by quantifying the chemotactic responses of different bacteria (Psuedoalteromonas haloplanktis, Escherichia coli) to different chemoattractants and chemorepellents. The MCD vastly accelerates the chemotactic screening process, which is critical to deciphering the complex sea of chemical stimuli underlying microbial responses.
Many microorganisms such as bacteria swim to explore their fluid habitats, which range from the human digestive system to the oceans. They can detect minute traces of food, toxins and other chemicals in their environment, and through a process called chemotaxis respond by swimming towards or away from them. Chemical concentrations naturally decrease with distance away from their source, forming gradients. By sensing these chemical gradients, and adjusting their swimming direction accordingly, cells can locate nutrients and other resources in harsh environments as well as avoid toxins and potential predators. Over the past 20 years, laboratory devices that manipulate minute volumes of fluid known as microfluidics devices have been indispensable for studying chemotaxis. They enable researchers to generate gradients of chemicals in carefully designed networks of microscopic channels, controlling the conditions that swimming cells are exposed to and mimicking their natural habitats. However, large-scale studies of chemotaxis have been limited by the sheer range of chemicals that are present at different levels in natural environments. Conventional microfluidic devices often compromise between distinguishing how individual cells behave, precise control over the chemical gradient, or the ability to execute multiple assays at the same time. Here, Stehnach et al. designed a microfluidic device called the Multiplexed Chemotaxis Device. The device generates five streams of precise dilutions of a chemical and then uses these streams alongside a control stream lacking the chemical to measure chemotaxis in six different conditions at the same time. The device was tested using a well-studied bacterium, Vibrio alginolyticus, which is commonly found in marine environments. The device reliably examined the chemotaxis response of the population to various chemicals, was able to carry out multiple assays more rapidly than conventional devices, and can be easily applied to study the response of individual bacteria under the same conditions. The Multiplexed Chemotaxis Device is relatively easy to manufacture using standard methods and therefore has the potential to be used for large-scale chemotaxis studies. In the future, it may be useful for screening new drugs to treat bacterial infections and to help identify food sources for communities of microbes living in marine environments.
