Antagonistic effect of Bacillus subtilis isolated and identified from different honey species against Klebsiella pneumoniae bee pathogens

Abstract

The search for alternative methods for treating and preventing bee dysbacteriosis is a priority for beekeeping as a branch of veterinary medicine. Lime honey, buckwheat honey, flower honey, forest honey, and acacia honey were tested to evaluate their antagonistic effect against a test culture of enterobacteria of bees of Klebsiella pneumoniae species. The study was conducted in several stages: 1. Determine the activity of honey microflora against a pure culture of enterobacteria of bees of Klebsiella pneumoniae bee pathogens; 2. The identification and isolation of Bacillus subtilis – bacteria-antagonists against Klebsiella pneumoniae bee pathogens; 3. Determine the antagonistic effect of pure culture of Bacillus subtilis against a pure culture of enterobacteria of Klebsiella pneumoniae bee pathogens. The antagonistic action of honey microorganisms and the determination of the most effective honey species were established by the diffusion method in agar wells. Staining of typical colonies from different types of honey revealed bacillary colonies of Gram-positive bacilli with endospores. Isolation of clean culture was conducted by a method of Gold. The cultural, tinctorial, and morphological signs of bacteria have been consistently determined in all investigated kinds of honey and coincided with characteristics of the Bacellaceae family. Specific belonging of bacteria-competitors was identified by biochemical typing. After determining their physiological properties in reactions and tests (activity of catalase, oxidase, urease, the ability to grow at different temperatures and to ferment carbohydrate substrate), the distinguished stamms of microorganisms from the investigated kinds of honey belong to the type of Bacillus subtilis. The repeated estimation of antagonistic action of pure cultures of Bacillus subtilis (isolated from each type of honey) against a pure culture of enterobacteria of bees of Klebsiella pneumoniae species confirmed their high activity. This type of microorganisms can represent the alternative component in probiotics at the therapy of dysbiosis of bees.

Keywords

• bees, honey, identification, bacillus subtilis, antagonism; klebsiella pneumoniae