Microscopic characterization of the bacterial cell envelope of Planococcus halocryophilus Or1 during subzero growth at −15 °C

Abstract

Microbial psychrophiles continue to expand our understanding of the adaptations required to thrive in cold environments. Planococcus halocryophilus strain Or1, a gram-positive, aerobic bacterial isolate from a Canadian high Arctic permafrost active layer, divides at temperatures as low as −15 °C and high salinity of 18 % NaCl. Initial studies of P. halocryophilus Or1 identified that under subzero conditions the cell envelope changed in appearance and composition. Our goal was to further analyze these features using scanning and transmission electron microscopy (SEM, TEM), confocal laser scanning microscopy (CLSM), and scanning transmission X-ray microscopy (STXM), which showed progressive changes in cell envelope composition during growth from optimal (25 °C) down to subzero (−15 °C) temperatures. S/TEM and CLSM illustrate that growth at −15 °C coincides with increasing hydrophobicity and distinct extracellular encrustations closely associated with the cell wall. STXM analyses resolved differences in cell composition with temperature, favoring higher amounts of protein and polysaccharide at higher temperatures compared to cells grown at −15 °C that were characterized by a cell envelope comprised of 20 % calcium carbonate, 50 % peptidoglycan, and 29 % choline. Analyses of the sequenced genome found the presence of several copies of carbonic anhydrase, a protein responsible for mineralization of calcium carbonate, and transcriptomic analyses revealed increased expression of a single copy at −15 °C along with the synthesis of peptidoglycan. The unique cell features of P. halocryophilus Or1 grown at −15 °C demonstrate unusual physiology that expands our understanding of psychrophilic adaptations and provides an example of microbially mediated calcium carbonate precipitation at subzero temperatures.