UC San Diego

The ocean makes up over 97% of the Earth’s biosphere and much of it is located within deep-sea habitats. Life at these depths must possess adaptations for growth at high hydrostatic pressures. Microorganism that grow preferentially at elevated pressures are termed “piezophiles.” Although prior studies have examined the piezophilic adaptations of Gram-negative microbes, little attention has been given to Gram-positive bacteria. In this project, I examined one of the few Gram-positive piezophiles, Carnobacterium sp. AT7. Two lineages of AT7 were subjected to progressively increasing hydrostatic pressures over the course of 130 generations. From these adaptive laboratory evolution (ALE) lineages, I obtained mutant strains which possessed improved rates of growth at high pressure and increased upper pressure limits. During these experiments, it was discovered that incubation of AT7 under stressful, high hydrostatic pressure for approximately 200 hours resulted in the acquisition of greatly improved high pressure growth in a single incubation cycle. Interestingly, cells derived from these incubations did not display stable high pressure growth and are presumed to have gained this ability through a physiological rather than genetic mechanism. Because it is well known that high pressure growth requires the production of sufficient proportions of membrane unsaturated fatty acids, it was investigated whether the long-term incubation cultures display increased unsaturated fatty acid fractions. Surprisingly, the Carnobacterium high pressure growth mutants do not produce increased proportions of unsaturated fatty acids. It was likewise discovered that these ALE-derived mutants do not possess alterations in cell size or shape. This thesis provides data and strains with which to begin to shed light on the processes employed by Gram-positive bacteria to adapt to elevated high pressure conditions.