Honeybees (Apis mellifera) are constantly exposed to a wide variety of environmental stressors such as parasites and pesticides. Among them, Nosema ceranae and neurotoxic insecticides might act in combination and lead to a higher honeybee mortality. We investigated the molecular response of honeybees exposed to N. ceranae, to insecticides (fipronil or imidacloprid), and to a combination of both stressors. Midgut transcriptional changes induced by these stressors were measured in two independent experiments combining a global RNA-Seq transcriptomic approach with the screening of the expression of selected genes by quantitative RT-PCR. Although N. ceranae-insecticide combinations induced a significant increase in honeybee mortality, we observed that they did not lead to a synergistic effect. According to gene expression profiles, chronic exposure to insecticides had no significant impact on detoxifying genes but repressed the expression of immunity-related genes. Honeybees treated with N. ceranae, alone or in combination with an insecticide, showed a strong alteration of midgut immunity together with modifications affecting cuticle coatings and trehalose metabolism. An increasing impact of treatments on gene expression profiles with time was identified suggesting an absence of stress recovery which could be linked to the higher mortality rates observed.