Workers of Apis mellifera Reared in Small-Cell Combs Show Higher Activity of the Proteolytic System in Hemolymph than Workers Reared in Standard-Cell Combs in Laboratory Cage Tests

Simple Summary Honeybees live in two different environments that are affected by multiple factors, e.g., pathogens or weather conditions. Thanks to the construction of the nests and behavioral cooperation honeybees became independent from the influence of external conditions. Moreover, a single bee has barriers that maintain homeostasis in its body and protect it from harmful factors, such as pesticides, air pollutants, or pathogens, through a system of proteases and their inhibitors. In feral bee colonies, bee comb cells vary in their width, while in Europe, colonies are kept only on standard-cell combs (5.50 mm cell width) or small-cell combs (4.90 mm cell width). In apiary conditions, the activities of proteases and their inhibitors in the 1-day-old workers were always higher in workers reared in standard-cell combs than in workers reared in small-cell combs, and opposite results were observed in groups of older workers (aged 7, 14, and 21 d). However, evaluating the influence of the nest factor is complex and difficult in field tests. Therefore, we assessed the proteolytic system activity in the hemolymph of colonies kept simultaneously on standard and small-cell combs in laboratory cage tests. The use of two types of combs in bee colony nests is an innovative approach to the use of small-cell combs in beekeeping. Irrespective of the age of the bees, the workers reared in the small-cell combs had higher protein concentrations. In turn, the activities of proteases and their inhibitors in the hemolymph of 1-day-old workers were higher in the group reared in the standard-cell combs. In older workers (aged 7, 14, and 21 days), higher protease and inhibitor activities were detected in the group reared in small-cell combs. The comparison of the results of apiary studies with those of laboratory cage tests shows that functions performed in the nest have a greater influence than age on the activity of proteases and their inhibitors in the hemolymph of worker bees. Abstract The aim of the study was to compare the activities of proteases and their inhibitors in the hemolymph of honeybee workers reared in small-cell combs (SMC) and standard-cell combs (STC) in laboratory cage tests. The analyses conducted in laboratory conditions facilitated assessment of the impact of the comb cell width (small vs. standard) along with eliminating the influence of many environmental factors on the results. The width of the comb cells in which the workers were reared had a significant effect on the protein concentrations and proteolytic system activities in the hemolymph. Irrespective of the age of the workers, higher protein concentrations were found in the hemolymph of the SMC workers. In turn, the activities of proteases and their inhibitors in the hemolymph of 1-day-old bees were higher in the STC workers. In older bees, aged 7–21 days, activity was higher in the SMC workers. The role of the considerable cell width variability in natural combs that were built without the use of an artificially produced wax foundation is worth investigating. It is highly probable that the impact of the comb cell width on the features of workers reared in these combs modifies the age polyethism in the worker caste as well. The investigation results of one-season studies of honeybees could be seriously affected by random factors. To reduce the risk of these effects, it is advisable to continue experiments over a few consecutive years.