# Reporting Key Features in Cold-Adapted Bacteria
## 4 Envelopes and Cold Adaptation  
The composition and structure of envelopes are recognized as important for cold adaptation as they constitute a dynamic interface between cell and environment, enabling them to cope with environmental challenges. Among the strategies used by cold-adapted microorganisms, a general well-known mechanism consists in the modification of cell membrane lipid composition, favoring shorter chains and decreasing lipid saturation [4] in order to maintain membrane fluidity while avoiding stiffness at low temperatures. Although the modifications of the internal membrane are well studied (Table 1), little is known about the changes that occur in other envelope components. In Gram negative bacteria, the envelope consists of an inner and an outer membrane separated by the periplasmic space containing a thin peptidoglycan layer. The outer membrane is formed by phospholipids, proteins and lipopolysaccharide-LPS contains the lipid A, anchored to the membrane, an intermediate oligosaccharide component (core) and an external O-polysaccharide. In _P. haloplanktis_, the presence of a higher percentage of cell envelope genes in the genome has been suggested as a specialized adaptation to cope with cold [56]. Similar observations were reported for _P. arcticus_, in which genes involved in membrane lipid and cell wall synthesis were identified as relevant for growth under subzero temperatures [57]. Proteomic analysis of _S. alaskensis_ showed that several proteins related to cell wall, membrane, exopolysaccharide biosynthesis and envelope biogenesis had a higher abundance at 10 \({}^{\circ}\)C [18]. In the Antarctic bacterium _Pseudomonas syringae_ Lz4w, it was observed that low temperatures caused changes in the composition and fluidity of LPSas higher polymyxin B sensitivity was detected along with an increased amount of hydroxy fatty acids [58]. In another Antarctic _Pseudomonas--P. extremaustralis--_the relevance of the core integrity of the LPS to cope with cold was evidenced by analyzing a _wapH_ mutant strain, encoding a core LPS glycosyltransferasee [59]. The _wapH_ strain was impaired to grow in the cold. The _wapH_ deficiency provoked a strong aggregative phenotype and modifications of envelope nanomechanical properties, such as lower flexibility and higher turgor pressure, cell permeability and surface area to volume ratio (S/V). Changes in these characteristics were also observed when _P. extremaustralis_ was grown at different temperatures. These results indicate that LPS appears as a novel essential feature for active growth under cold conditions [59]. In Gram positive bacteria where the envelope is composed of the cell wall and the inner membrane, the importance of these components has also been recognized. The increase in cell wall biosynthesis transcripts was observed in _E. sibiricum_ at \(-2.5\)\({}^{\circ}\)C, suggesting that the thickening of the cell wall may protect the cell against disruption by ice formation and/or osmotic pressure, which can be generated at subzero temperatures [10]. Cell wall, membrane and envelope-component synthesis were also shown to be fundamental to supporting cold active growth in _Planococcus halocyophilus_ Or1. This bacterium possesses an unusual cell envelope characterized by encrustations surrounding cells during subzero growth at \(-15\)\({}^{\circ}\)C [13]. Transcriptomic analysis showed the relevance of genes encoding peptidoglycan synthetase, along with those involved in the synthesis of precursors [13]. Further microscopic characterization of the cell envelope showed increasing hydrophobicity and distinct extracellular encrustations of 20% calcium carbonate, 50% peptidoglycan and 29% choline closely associated with the cell wall. Along with this, genome analysis allowed the identification of several copies of genes encoding carbonic anhydrase, responsible for mineralization of calcium carbonate.