Antimicrobial activities of Bacillus velezensis strains isolated from stingless bee products against methicillin-resistant Staphylococcus aureus

Abstract

Infections caused by methicillin-resistant Staphylococcus aureus (MRSA) have reached epidemic proportions globally. Therefore, there is an urgent need for a continuous supply of antibiotics to combat the problem. In this study, bacteria initially identified as species belonging to the Bacillus amyloliquefaciens operational group were re-identified based on the housekeeping gene, gyrB. Cell-free supernatants (CFS) from the strains were used for antimicrobial tests using the agar well diffusion assay against MRSA and various types of pathogenic bacteria. The minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and physicochemical characteristics of the CFS were determined. Based on gyrB sequence analysis, five strains (PD9, B7, PU1, BP1 and L9) were identified as Bacillus velezensis. The CFS of all B. velezensis strains showed broad inhibitory activities against Gram-negative and -positive as well as MRSA strains. Strain PD9 against MRSA ATCC 33742 was chosen for further analysis as it showed the biggest zone of inhibition (21.0 ± 0.4 mm). The MIC and MBC values obtained were 125 μl/ml. The crude antimicrobial extract showed bactericidal activity and was stable at various temperatures (40–80°C), pH (4–12), surfactants (Tween 20, Tween 80, SDS and Triton X-100) and metal ions (MgCI2, NaCI2, ZnNO3 and CuSO4) when tested. However, the crude extract was not stable when treated with proteinase K. All these properties resembled the characteristics of peptides. The antimicrobial compound from the selected strain was purified by using solvent extraction method and silica gel column chromatography. The purified compound was subjected to High Performance Liquid Chromatography which resulted in a single peak of the anti-MRSA compound being detected. The molecular weight of the anti-MRSA compound was determined by using SDS-PAGE and zymogram. The size of the purified antimicrobial peptide was approximately ~ 5 kDa. The antimicrobial peptide produced from B. velezensis strain PD9 is a promising alternative to combat the spread of MRSA infections in the future.

Antimicrobial activity of supernatants produced by bacteria isolated from Brazilian stingless bee’s larval food

“…In the study by Nishio et al [ 41 ], the antibacterial activity of honey produced by stingless bees Scaptotrigona postica and Scaptotrigona bipunctata against methicillin-resistant Staphylococcus aureus (MRSA) and sensitive S. aureus strains was verified. A recent study demonstrated broad inhibiting activity against MRSA strains by the supernatant of Bacillus velezensis isolated from stingless bees [ 44 ]. However, contrary to our study, most authors also report relevant antimicrobial action on Sensitive S. aureus (RRR).…”

Scale-up production of and dietary supplementation with the recombinant antimicrobial peptide tilapia piscidin 4 to improve growth performance in Gallus gallus domesticus

“…In addition, the antibacterial activity of AMPs can be enhanced by modifying the structural properties of natural peptides [ 36 – 38 ]. In a similar report, the crude antimicrobial extract of Bacillus velezensis strains isolated from stingless bee products showed bactericidal activity against methicillin-resistant Staphylococcus aureus and was stable at various temperatures (40–80°C) [ 39 ]. Furthermore, Plantaricin LD1 produced by Lactobacillus plantarum LD1 showed stability at high temperatures (100°C for 20 min and 121°C for 15 min under 15 psi) and inhibited bacteria such as Staphylococcus aureus , urogenic Escherichia coli , Pseudomonas aeruginosa , Salmonella typhi , Shigella flexneri and Vibrio sp [ 40 ].…”

Complete Genome Sequence of Bacillus velezensis GMEKP1, Isolated from a Natural Bamboo Hive of Stingless Bees

“…The isolation procedure used the spread plate method of each serial dilution on International Streptomyces Project-2 (ISP-2) medium, with incubation at 30°C for 3 days under aerobic conditions. In another previous study, B. velezensis isolated from stingless bee products of Heterotrigona itama exhibited antimicrobial activity against methicillin-resistant Staphylococcus aureus ( 3 ). In this study, we present the annotated genome sequence of B. velezensis GMEKP1 and report the potential active compounds encoded by its biosynthetic gene clusters (BGCs).…”