Water kefir as a source of bacteria that inhibit American foulbrood disease in honey bees.

RODRÍGUEZ, MA., ; BRENDAN AD., ; FERNÁNDEZ LA., ; DÍAZ ML., ; GALLO CA., ; CORONA M., ; EVANS J., ; REYNALDI FJ., ; THOMPSON GJ

London, Ontario

Jornada; 53rd Ontario ecology, ethology & evolution Colloquim; 2023

Western Ontario University

We studied water kefir as an unexplored and innovative source of potential probiotics to be incorporated into honey bee diet. We described the bacteria diversity by a high-throughput sequencing technique and isolated bacteria from water kefir in order to evaluate the antimicrobial activity in vitro of soluble substances released by those bacteria against Ascosphaera apis (deadly fungal agent of Chalkbrood disease). In vivo assays were done in order to to evaluate the effect of bacteria supernatants on bee larvae. Briefly, diversity was calculated for each data set (three grain and three beverage samples) by Shannon and Simpson index. The inferred OTUs were used to interrogate the SILVA reference database to determine the bacterial relative abundance. Bacteria were isolated on MRS medium and incubated at 37°C under anaerobic conditions for five days. Isolates were identified by phenotypic characterization, centrifuged (10000rpm, 10min) and supernatants were conserved. In vitro studies of antimicrobial activity were done in plates: circles of mycelium from A. apis growing on YGPSA medium were placed in the centre of plates with wells containing the bacteria supernatants (80µl). In vivo assays were done in microplates: bee larvae were reared at laboratory (35°C, 90%RH) and the survival, weight and size of larvae was measured. Bees were fed seven days with a control diet (sugar solution with royal jelly) and a diet of royal jelly containing the supernatants. Results revealed that the estimation of species richness and the microbial α-diversity was higher in kefir beverage samples compared to grains. The most represented genus were Liquorilactobacillus, Leuconostoc and Oenococcus in grains and Anaerocolumna and Ralstonia in beverage. Isolates which supernatants have shown the greatest antagonism against A. apis were identified as Lentilactobacillus hilgardii and Lentilactobacillus buchneri. The variables weight, size and mortality of larvae did not show statistically differences (p>0,1) between control and treatment. Thus, beneficial bacteria were isolated from kefir and could be used as potential probiotics to feed bees.