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Apis cerana cerana is an excellent subspecies of Apis cerana, playing a vital role in pollination for wild flowers and crops as well as ecological balance. Nosema ceranae, an emergent fungal parasite infecting various bee species, originates from eastern honeybee. In this article, midguts of N. ceranae-inoculated A. c. cerana workers at 7 days post inoculation (dpi) and 10 dpi (AcT1 and AcT2) and un-inoculated workers’ midguts (AcCK1, AcCK2) were subjected to Nanopore-based genome-wide DNA methylation sequencing. Totally, 1773258, 2151476, 1927874 and 2109961 clean reads were generated from AcCK1, AcCK2, AcT1, and AcT2 groups, with the N50 lengths of 7548, 7936, 7678, and 7291 and the average quality value of 8.97, 8.95, 9.24, and 8.98, respectively. Among these, 93.85%, 94.49%, 88.69%, and 81.27% clean reads could be mapped to the reference genome of A. c. cerana. In the aforementioned four groups, 2149685, 2614513, 1637018 and 2726985 CHG sites were identified; the numbers of CHH sites were 9581990, 11801082, 7178559, and 12342423, whereas those of CpG sites were 14325356, 15703508, 14856284 and 13956849, respectively. Additionally, there were 36114, 118867, 30249, and 82984 6mA methylation sites respectively discovered. These data can be used for identifying differential 5mC methylation and 6mA methylation engaged in response of eastern honeybee workers to N. ceranae infestation, and for investigating the 5mC or 6mA methylation-mediated mechanism underlying host response.Value of the dataThe data provide enrichment for information about 5mC and 6mA methylation in eastern honeybees.Our data contributes to clarification of the epigenetic mechanism underlying eastern honeybee worker’s response to microsporidian infestation.This presented data offer novel insights into interaction between Apis cerana and Nosema ceranae.