Inverse Relationship Between Halophilic Growth and Cell Integrity Under Extremely Chaotropic Conditions

Concentrated magnesium chloride brines are extreme environments that are inhospitable to life on Earth. The ionic strength of these brines significantly depresses water activity and concomitantly exerts significant chaotropic stress. Although these brines are largely considered sterile, the well-known preservative effects of magnesium chloride on certain biomolecules, such as DNA, confound life detection approaches and efforts to constrain precisely the habitable window of life on Earth. While the ability of these brines to preserve genetic material is well documented, the preservation of whole cells, which are generally thought to be preserved in magnesium chloride brines, is poorly described. This work explores the effects of long-term exposure of highly chaotropic magnesium chloride on viability, cell integrity, and DNA preservation in the model organisms <i>Escherichia coli</i>, <i>Salinibacter ruber</i>, <i>Halobacterium salinarum</i>, and <i>Haloquadratum walsbyi</i>. The selected halophiles are relevant for this study as they are abundant and globally distributed in brine environments, while <i>E. coli</i> was chosen to represent infall or transport of non-adapted cells. We observed unexpected resilience in <i>E. coli</i>, which survived in 4 M magnesium chloride for longer than the tested halophiles, and nonviable cells maintained structural whole-cell integrity for over 3 years. Whole <i>S. ruber</i> cells were also preserved in 4 M magnesium chloride, while the tested haloarchaea lost viability and completely degraded within hours of exposure. DNA from all tested strains was recovered from incubations after upwards of 3 years of exposure; it showed some signs of degradation but was nonetheless still amplifiable via polymerase chain reaction. Our work demonstrates that the preservation of whole cells in magnesium chloride brines is not universal. Considering the potential abundance of chaotropic brine environments within our solar system, understanding the limits of life and the preservation of biosignatures in these brines is critical to inform future life detection missions on Earth and beyond.