The polyextremophilic bacterium Clostridium paradoxum attains piezophilic traits by modulating its energy metabolism and cell membrane composition

In polyextremophiles, i.e ., microorganisms growing preferentially under multiple extremes, synergistic effects may allow growth when application of the same extremes alone would not. High hydrostatic pressure (HP) is rarely considered in studies of polyextremophiles, and its role in potentially enhancing tolerance to other extremes remains unclear. Here, we investigated the HP-temperature response in Clostridium paradoxum a haloalkaliphilic, moderately thermophilic endospore-forming bacterium, in the range 50-70 °C and 0.1-30 MPa. At ambient pressure, growth limits were extended from previously reported 63 to 70 °C, defining C. paradoxum as an actual thermophile. Concomitant application of high HP and temperature as compared to standard conditions ( i.e. , ambient pressure, 50 °C) remarkably enhanced growth, with an optimum growth rate observed at 22 MPa and 60 °C. HP distinctively defined C. paradoxum physiology: at 22 MPa biomass production increased by 75% and the release of fermentation products per cell decreased by >50% as compared to ambient pressure. This metabolic modulation was apparently linked to an energy-preserving mechanism triggered by HP, involving a shift towards pyruvate as the preferred energy and carbon source. High HPs decreased cell damage as determined by Syto9 and propidium iodide staining, despite no organic solute was accumulated intracellularly. A distinct reduction in carbon chain length of phospholipid fatty acids (PLFAs) and an increase in the amount of branched-chain PLFAs occurred at high HP. Our results describe a multi-faceted, cause-effect relationship between HP and cell metabolism, stressing the importance of applying HP to define the boundaries for life under polyextreme conditions. Importance Hydrostatic pressure (HP) is a fundamental parameter influencing biochemical reactions and cell physiology, however, it is less frequently applied as compared to other factors such as pH, temperature and salinity when studying polyextremophilic microorganisms. In particular, how HP affects microbial tolerance to other and multiple extremes remains unclear. Here, we show that under polyextreme conditions of high pH and temperature Clostridium paradoxum demonstrates a moderately piezophilic nature as cultures grow to highest cells densities and most efficiently at a specific combination of temperature and HP. Our results highlight the importance of considering HP when exploring microbial physiology under extreme conditions and thus have implications for defining the limits for microbial life in nature and for optimizing industrial bioprocesses occurring under multiple extremes.