22Decrease in the frequency of arginine and increase in lysine are the trends that have 23 been identified in the genomes of cold adapted bacteria. However, some cold adapted 24 taxa show only limited or no detectable changes in the frequencies of amino acid 25 composition. Here, we examined Arthrobacter spp. genomes from a wide range of 26 environments on whether the genomic adaptations can be conclusively identified 27 across genomes of taxa from polar and alpine regions. Phylogenetic analysis with a 28 concatenated alignment of 119 orthologous proteins revealed a monophyletic 29 clustering of seven polar and alpine isolated strains. Significant changes in amino acid 30 composition related to cold adaptation were exclusive to seven of the twenty-nine 31 strains from polar and alpine regions. Analysis of significant indicator genes and cold 32 shock genes also revealed that clear differences could only be detected in the same 33 seven strains. These unique characteristics may result from a vast exchange of 34 genome content at the node leading to the monophyletic cold adapted Arthrobacter 35 cluster predicted by the birth-and-death model. We then experimentally validated that 36 strains with significant changes in amino acid composition have a better capacity to 37 grow at low temperature than the mesophilic strains. 38 Importance 39 Acquisition of novel traits through horizontal gene transfer at the early divergence of 40 the monophyletic cluster may accelerate their adaptation to low temperature. Our 41 study reached a clear relationship between adaptation to cold and genomic features 42 and would advanced in understanding the ambiguous results produced by the previous 43 studies on genomic adaption to cold temperature. 44 regions 46 47 65 validated by analysis the structural, kinetic and microcalorimetric of cold adapted 66 enzymes, e.g. aminopeptidase, β-lactamases, and dienelactone hydrolase (GaDlh) 67 (12-14).68 69 Comparative genomic analyses to examine differences in amino acid composition 70 toward cold adaptation came through the study of two methanogenic Archaea, 71 Methanogenium frigidum and Methanococcoides burtonii from Ace Lake, Antarctica 72 (15). Proteins from these cold-adapted Archaea were characterized by a higher 73 proportion of non-charged polar amino acids, such as glutamine and threonine, and a 74 lower proportion of hydrophobic amino acids, particularly leucine (15). The amino 75 acid shifts toward increased enzyme flexibility, which confers catalytic efficiency and 76 contribute to cold adaptation can be identified in genomes of Psychromonas 77 ingrahamii 37, Exiguobacterium sibiricum 255-15, Psychrobacter arcticus 273-4, 78 Shewanella spp. and Glaciecola spp. (16-19). However, these changes were limited in 79 Colwellia psychrerythraea 34H, Planococcus halocryophilus Or1, Rhodococcus sp. 80 JG3, Arthrobacter spp., Actinotalea sp. KRMCY2, Polaromonas sp. Eur3 1.2.1, 81 Paenisporosarcina sp. Eur1 9.01.10, Methylobacterium sp. AL-11 and Kocuria sp. 82 KROCY2 (13, 20-24). Even, no changes ...