ModuloStat: An Internet of Things’ Path to Continuous Cultures in Mini-Bioreactors

In continuous culture, a population of microorganisms is propagated in a stable environment over many generations. This is particularly relevant for experimental evolution and metabolic studies. However, continuous culture protocols are difficult to implement, so they are not commonly used in microbiology laboratories. Here, we present the ModuloStat, a modular, open-source framework that facilitates continuous culture in mini-bioreactors. The ModuloStat system is grounded on digital fabrication tools easily accessible in FabLabs and programmable electronics. Maintaining a culture is divided into tasks assigned to dedicated printed circuit boards with a microcontroller connected to a Wi-Fi network. According to Internet of Things principles, each board operates a set of sensors and actuators autonomously and can receive and send information. The boards are stacked to implement complex behaviors and can be modified to accommodate new features. A thermoregulated box holds the components and can be placed on a laboratory bench or transported under a sterile hood for inoculation. Sterility is ensured by autoclaving, after assembly, all components that will come into contact with the culture medium. <i>In-situ</i> optical density monitoring combined with modularity and computer control enables many cultivation modes. Additionally, we present the construction of the <i>Bacillus subtilis</i> strain ZB designed for bioreactor culture that exhibits a zero-biofilm phenotype. To demonstrate the system's versatility, we performed several experimental cultures with this model organism, including chemostat, turbidostat, medium swap, and a cascade of bioreactors.