Apis mellifera is an important pollinator for agriculture, with a large distribution across the globe, generalist feeding behaviour, and high efficiency as a pollinator. The recent decline of Apis mellifera is affected by deforestation, climate change, spillover of pathogens, and a high demand for honey. The gut microbiota of honey bees can be a key to improving the health of the colonies and stopping the decline. This project aims to determine whether the gut microbiota of honey bees changes over winter and analyze the similarities and differences in the gut microbiota of the Apis mellifera mellifera subspecies by comparing samples collected in early autumn and spring. The study will specifically focus on the impact of the honey bee's diet, which varies between honey and sugar-water intake. This was accomplished by sequencing the 16S rRNA gene using nanopore sequencing technology. As a result, it was determined that the gut microbiota changes during the winter season, and Bartonella may play a crucial role in honey bees’ well-being in winter. The honey-fed group contained the most diverse bacterial composition, including species that were not determined in the sugar-fed and reference groups. Thus, it is believed, the diet affected the gut microbiota composition. The honey-fed group also contained a novel species Novisyntrophococcus fermenticellae, which was not previously found in honey bees.