[Objectives]  To investigate the expression of the p38 MAPK gene, which plays an important role in the insect cold response mechanism, in different overwintering stages of the western honeybee Apis mellifera. [Methods] The amino acid sequence of the Apis mellifera p38 MAPK gene was analyzed and its mRNA expression in different overwinter stages of Apis mellifera ligustica, Apis mellifera mellifera, Apis mellifera caucasica and Apis mellifera carnica detected using RT-qPCR. [Results]  The p38 MAPK gene of A. mellifera includes a 1 083 bp open reading frame, encodes a polypeptide with 360 amino acids and contains a putative dual phosphorylation motif TGY and a phosphorylated activating ring sequence. P38 MAPK was expressed in all honeybee tissues. Highest and lowest expression occurred in the thorax and abdomen, respectively. Expression varied in different overwintering stages and subspecies. Expression was higher outdoors than indoors in all four subspecies P<0.05). In winter, p38 MAPK mRNA expression first increased then decreased in A. mellifera ligustica and A. mellifera mellifera which have different overwintering strategies. Expression increased in both outdoor and indoor overwintering colonies of A. mellifera caucasica. Unlike these three subspecies, expression remained constant in indoor overwintering colonies of A. mellifera carnica and tended to decrease after an initial increase in outdoor overwintering colonies of this subspecies. Overwintering expression levels varied among subspecies except in January of the second year (P<0.05). [Conclusion]  The p38 MAPK gene plays an important physiological role in the overwintering processes of the western honeybee. The p38 MAPK signal pathway could be a candidate signal pathway for further study of the honeybee cold resistance mechanism.