Antibacterial potential components of Bacillus species and antibiotics residues in branded and unbranded honey samples from Nigeria

Abstract

Honey is a sweet viscous liquid produced by honey bee, Apis mellifera from the nectar of plants. Honey is a natural product that has been used from ancient times till now as food and for medicinal purpose. This study was carried out to determine the mode of action of Bacillus species and antibiotics residues in branded and unbranded honey samples from Nigeria. Bacilli spp. count was carried out by initially heating diluted honey samples in water bath set at 80°C for 15 min, while total bacterial count was carried out using the pour plate method. Antibacterial activity of identified Bacillus spp on Micrococcus was determined using well-in agar method while the mode of action was carried out by reporter assay method. Detection of tetracycline, gentamycin and streptomycin was analyzed using high performance liquid chromatography (HPLC) column oven L-2300 and Column intensil ODS-3C18 (250 x 4.6 mm). Honey samples (2 g) were extracted for HPLC by deprotenizing using acetonitrile and methanol with flow rate of 1 mL/min and RID detector was used to detect antibiotic residue. Bacilli from honey were characterized physiologically, morphologically and biochemically, they were tentatively identified as Bacilli licheniformis, Bacilli subtilis, Bacilli coagulans, Bacilli cirulans, Bacilli pumilis and Bacilli badius. The most prevalent Bacillus spp. were B. licheniformis and B. subtilis. Total bacteria count for branded honey ranged from 2.2 x 10² to 5.5 x 10³ cfu/g, while Bacilli count ranged from nil to 6.2 x 10² cfu/g. For unbranded honey samples, total bacteria count ranged from 7.0 x 10³ to 3.5 x 10² cfu/g, while Bacilli count ranged from 5 x 101 to 1.6 x 10³ cfu/g. Four of the isolates representing branded (SF2 and RW2) and unbranded honey samples (EH2 and TC2) exhibited antibacterial activity against Micrococcus; one isolate (SF2) showed cell wall causing antibacterial activity. Tetracycline was detected more in the unbranded honey samples while gentamycin and streptomycin were detected in just two unbranded honey samples, indicating that tetracycline is used frequently for the treatment of bee diseases that is why it is detected as residue in the finished honey product.

Key words: Antibiotics, Bacillus, health benefit honey, high performance liquid chromatography (HPLC), residues.