High-pressure continuous culturing: life at the extreme.

Microorganisms adapted to high hydrostatic pressures at depth in the oceans and within the subsurface of Earth's crust represent a phylogenetically diverse community thriving under extreme pressure, temperature, and nutrient availability conditions. To better understand the microbial function, physiological responses, and metabolic strategies at in-situ conditions requires high-pressure (HP) continuous culturing techniques that, although commonly used in bioengineering and biotechnology applications, remain relatively rare in the study of the Earth's microbiomes. Here, we focus on recent developments in the design of HP chemostats, with particular emphasis on adaptations for delivery and sampling of dissolved gases. We present protocols for sterilization, inoculation, agitation, and sampling strategies that minimize cell lysis, applicable to a wide range of chemostat designs.