A high concentration of hydroxymethylfurfural (HMF) (e.g., 15 mg HMF per kg honey) indicates quality deterioration for a wide range of foods. In honey bee colonies, HMF in stored honey can negatively affect bee health and survival. Therefore, in the laboratory, we experimentally determined the effects of HMF on the longevity and midgut integrity of worker Apis mellifera carnica by feeding bees standard diets containing five concentrations of HMF (100, 500, 1000, and 1500 ppm). Simultaneously, we also examined HMF’s effect on Nosema ceranae spore counts within infected honey bees. We performed an immunohistochemical analysis of the honey bee midgut to determine possible changes at the cellular level. No correlation was established between HMF concentration and N. ceranae spore counts. Negative effects of HMF on bees were not observed in the first 15 days of exposure. However, after 15 to 30 days of exposure, HMF caused midgut cells to die and an increased mortality of honey bee workers across treatment groups.