*2.2. Production of N. ceranae Spores*

Honey bee colonies infected with *N. ceranae* were identified in a apiary nearby Modena (Italy), in the city of Savignano sul Panaro (44°29- 03.4-- N 11°03- 28.1-- E). Spores were collected from diseased hives by capturing flying foragers honey bees over the colony entrance. Obtained foragers were sacrificed and the midgut and rectum extracted and broken up in distilled water. Spores were divided in multiple stocks and conserved in a 10% glycerol PBS solution at −80 °C to guarantee the same *N. ceranae* strain availability for all the assays. When an assay was established, spores were retrieved from cryostat and about thirty-five newly emerged bees were caged and infected by feeding bees with the prepared solution to allow *N. ceranae* proliferation and sporulation, in order to obtain fresh and highly infective spores. A sample of the spores obtained was characterized and confirmed as *Nosema ceranae* according to [44] and the same stock was used for all infections. Fresh spores were extracted from infected and caged honey bees, counted with Neubauer chamber to estimate the load per ml, diluted in a sugar syrup solution to the final concentration of 10<sup>4</sup> spore/mL and used as a fresh and standardized *N. ceranae* propagules for the ongoing assay.

#### *2.3. Oral Infection with N. ceranae Spores*

Newly emerged honey bees (*Apis mellifera ligustica*) were obtained from brood frames with worker bees ready to emerge. Brood frames were picked from three different colonies for every assay, in order to homogenize the genetic variability [45]. Frames were were maintained under controlled conditions (32 °C; 60% RH) until honey bees emerged. Then, newly emerged bees were kept in ventilated cages for 2 days until individual inoculation with control treatment (syrup) or 5 × 104 *N. ceranae* freshly prepared spores in syrup according to Porrini et al. [46]. The temperature and relative humidity were maintained during the experiments at 29 °C and 60% RH, respectively. Before starting the administration of the ingredients, a sample of newly emerged individuals was taken to test the potential basal infection with *N. ceranae*.

#### *2.4. Cultivation of Saccharomyces sp.*

*Saccharomyces* sp. was isolated from forest soil collected at the Bologna Apennines (Italy) 250 m a.s.l. (data not shown) and it was grown on PDB culture broth and incubated at 120 rpm for 5 days at 35 °C. Then, the obtained culture was mixed in sugar syrup 1:1 (*w*:*v*) in equal amounts in order to obtain 1011 CFUmL.
