Effects of Hydroxymethylfurfural (HMF) on learning behavior and head learning-related genes expression of Apis mellifera ligustica workers

Hydroxymethylfurfural (HMF) is a carbohydrate substance easily produced after heating and denaturation. It emerges as a degradation product of specific sugars that may produced by bee feed if beekeeping processes lack standardization. While the toxicological impacts of Hydroxymethylfurfural on mammals are well-documented, studies on its neurotoxic effects on honey bees are still limited, particularly with respect to its potential impact on learning and cognitive function. In this study, we investigated whether HMF affected learning and cognition in Apis mellifera ligustica worker bees by administering a defined concentration of HMF and assessing their associative learning and cognitive ability using odor associative learning, with the aim of exploring whether HMF had an impact on the learning and cognitive abilities of bees and verifying it at the molecular level. We further examined the molecular basis of any observed effects to provide a scientific basis for risk assessment related to HMF exposure in practical beekeeping and colony management. Six-day-old worker bees were collected from colonies and reared in cages under controlled temperature and humidity conditions for 10 days. During this period, the treatment group was allowed ad libitum access to a 30% (w/v) sucrose solution containing 1, 500 ng/μL HMF, while the control group received a 30% sucrose solution without HMF. On day 16, the proboscis extension response (PER) assay was used to assess olfactory associative learning. Additionally, quantitative real-time PCR (qRT-PCR) was conducted to evaluate differential expression of learning-related genes in the bee head. There was no significant difference in mortality between the treatment group and the control group during the 10-day feeding period (P > 0.05). The PER experiment results revealed that the proboscis extension rate of the treatment group was significantly lower than that of the control group (P < 0.05). In comparison with the control group, the expression of head learning-related genes TpnC Ⅲa, OPN4, Obp 4 and PRKACB in the treatment group down-regulated significantly, while pkc up-regulated significantly. These findings indicate that HMF at 1 500 ng/μL does not induce acute mortality in honey bees but impairs their learning performance and alters the expression of head genes associated with learning significantly.