# Adaptation of the Marine Bacterium _Shewanella baltica_ to Low Temperature Stress
## 2 Results and Discussion
### Global Changes in the Transcriptome of S. baltica in Response to Cold Stress
(**B**) Heatmap showing row Z-score transformed, row scaled expression for corresponding genes. Column 0 stands for control sample for 0 min time point; replicates for 90 and 180 min time points are labeled R\({}_{1}\) and R\({}_{2}\). (**C**) Direction of changes in respect to control (time 0) for corresponding RNA-seq and RT-qPCR assays. Asterisk (*) marks the expression change categorized as up although in fact it is upregulated only after 90 min while downregulated after 180 min. Changes in the Metabolic Pathways of S. baltica in the Adaptation and Survival under Cold Stress Conditions  
For analysis of metabolic pathways which could be enriched upon cold stress, the KEGGprofile package version 1.30.0 was employed [33]. The three-letter identifier for _S. baltica_ OS185 used in our analysis was \"sbm\". Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment in genes up- and downregulated upon cold stress was considered significant for \(p\) value \(<\) 0.01. For numbers of enriched genes, gene names and statistics obtained in the pathway enrichment analysis see the Supplementary Table S3. Fold change values of differentially expressed genes are available in Supplementary Table S1. Enrichment analysis showed that only few pathways were upregulated in the cold stress exposed _S. baltica_ (Supplementary Figure S1 and Table S3). After 90 min, there was a significant upregulation of a few metabolic pathways, including glycerolipid metabolism (KEGG: 00561), riboflavin metabolism (KEGG: 00740), pentose phosphate pathway (KEGG: 00030), and aminoacyl-tRNA biosynthesis (KEGG: 00970). Interestingly, flagellar assembly (KEGG: 02040) was also detected as a significantly upregulated cellular process. Additionally, after 180 min the sulfur metabolism pathway became upregulated (KEGG: 00920). However, much more pathways were downregulated upon cold stress (Supplementary Figure S1 and Table S3). After 90 min, there was a significant downregulation of glycolysis/glucoseogenesis (KEGG: 00010), fatty acid degradation (KEGG: 00071), pyruvate metabolism (KEGG: 00620), propanoate metabolism (KEGG: 00640), C5-branched dibasic acid metabolism (KEGG: 00660), and pantotheanate and CoA biosynthesis (KEGG: 00770). Significantly downregulated pathways were assigned to amino acid metabolism, including pathways of valine, leucine and isoleucine degradation and biosynthesis (KEGG: 00280 and 00290), histidine metabolism (KEGG: 00340), and phenylalanine, tyrosine and tryptophan biosynthesis (KEGG: 00400). Downregulation was also observed for a bacterial secretion system (KEGG: 03070). After 180 min, additional pathways became downregulated, including pathways of amino acid metabolism--arginine and proline metabolism (KEGG: 00330) and beta-alanine metabolism (KEGG: 00410), as well as of butanoate metabolism (KEGG: 00650), and nicotinate and nicotinamide metabolism (KEGG: 00760). Many pathways that were affected by gene expression changes in _S. baltica_ in our study, were also reported as affected by cold stress in _S. oneidensis_[23]. For example, downregulation of genes in _S. oneidensis_ was dominant among amino acids biosynthesis or energy metabolism pathways. Genes whose products are involved in the fatty acid and phospholipid metabolism or transport/binding proteins were affected as well. The same pathways or similar processes were affected in our study of _S. baltica_. To conclude, _S. baltica_ shows a wide range of transcriptional changes in response to the cold stress challenge. Comparison of these changes with changes in other _Shewanella_ species may reveal a more general picture of the transcriptomic cold stress response of these bacteria. In subsequent sections, we will discuss a possible role of selected gene expression changes in potential adaptation and survival strategies of _S. baltica_ undergoing cold stress.