Cannabis sativa CBD Extract Exhibits Synergy with Broad-Spectrum Antibiotics against Salmonella enterica subsp. Enterica serovar typhimurium

Abstract

New generation antibiotics are needed to combat the development of resistance to antimicrobials. One of the most promising new classes of antibiotics is cannabidiol (CBD). It is a non-toxic and low-resistance chemical that can be used to treat bacterial infections. The antibacterial activity of Cannabis sativa L. byproducts, specifically CBD, has been of growing interest in the field of novel therapeutics. As research continues to define and characterize the antibacterial activity that CBD possesses against a wide variety of bacterial species, it is important to examine potential interactions between CBD and common therapeutics such as broad-spectrum antibiotics. In this study it is demonstrated that CBD-antibiotic (combination of CBD and antibiotic) co-therapy can effectively fight Salmonella typhimurium (S. typhimurium) via membrane integrity disruption. This research serves to examine the potential synergy between CBD and three broad-spectrum antibiotics (ampicillin, kanamycin, and polymyxin B) for potential CBD-antibiotic co-therapy. In this study, it is revealed that S. typhimurium growth is inhibited at very low dosages of CBD-antibiotic. This interesting finding demonstrates that CBD and CBD-antibiotic co-therapies are viable novel alternatives to combating S. typhimurium.

Keywords: Salmonella; cannabidiol; cell-membrane integrity; co-therapy; novel antibacterial agents.

Figure 1

Synergistic analysis of ampicillin and CBD co-treatment of S. typhimurium. (A) Synergistic activity was observed at 0.5 μg/mL ampicillin and 1 μg/mL CBD co-treatment. Data presented as average OD⁶⁰⁰ ± SEM (B) Immunofluorescence staining analysis positive (2% SDS) and negative (dH₂O) controls. Immunofluorescence staining of S. typhimurium treated with ampicillin at 0.5 and 5 μg/mL as well as the observed synergistic combination of 0.5 μg/mL ampicillin and 1 μg/mL CBD.

Figure 1

Synergistic analysis of ampicillin and CBD co-treatment of S. typhimurium. (A) Synergistic activity was observed at 0.5 μg/mL ampicillin and 1 μg/mL CBD co-treatment. Data presented as average OD⁶⁰⁰ ± SEM (B) Immunofluorescence staining analysis positive (2% SDS) and negative (dH₂O) controls. Immunofluorescence staining of S. typhimurium treated with ampicillin at 0.5 and 5 μg/mL as well as the observed synergistic combination of 0.5 μg/mL ampicillin and 1 μg/mL CBD.

Figure 2

Synergistic analysis of kanamycin and CBD co-treatment of S. typhimurium. (A) Potential synergistic activity was observed at 5 μg/mL kanamycin and 1 μg/mL CBD co-treatment. Data presented as average OD⁶⁰⁰ ± SEM. (B) Immunofluorescence staining analysis positive (2% SDS) and negative (dH₂O) controls. Immunofluorescence staining of S. typhimurium treated with kanamycin at 0.5 and 5 μg/mL as well as the observed synergistic combination of 5 μg/mL kanamycin and 1 μg/mL CBD.

Figure 2

Synergistic analysis of kanamycin and CBD co-treatment of S. typhimurium. (A) Potential synergistic activity was observed at 5 μg/mL kanamycin and 1 μg/mL CBD co-treatment. Data presented as average OD⁶⁰⁰ ± SEM. (B) Immunofluorescence staining analysis positive (2% SDS) and negative (dH₂O) controls. Immunofluorescence staining of S. typhimurium treated with kanamycin at 0.5 and 5 μg/mL as well as the observed synergistic combination of 5 μg/mL kanamycin and 1 μg/mL CBD.

Figure 3

Synergistic analysis of polymyxin B and CBD co-treatment of S. typhimurium. (A) Potential synergistic activity was observed at 0.5 μg/mL polymyxin B and 1 μg/mL CBD co-treatment. Data presented as average OD⁶⁰⁰ ± SEM. (B) Immunofluorescence staining analysis positive (2% SDS) and negative (dH₂O) controls. Immunofluorescence staining of S. typhimurium treated with polymyxin B at 0.5 and 5 μg/mL as well as the observed synergistic combination of 0.5 μg/mL polymyxin B and 1 μg/mL CBD.

Figure 3

Synergistic analysis of polymyxin B and CBD co-treatment of S. typhimurium. (A) Potential synergistic activity was observed at 0.5 μg/mL polymyxin B and 1 μg/mL CBD co-treatment. Data presented as average OD⁶⁰⁰ ± SEM. (B) Immunofluorescence staining analysis positive (2% SDS) and negative (dH₂O) controls. Immunofluorescence staining of S. typhimurium treated with polymyxin B at 0.5 and 5 μg/mL as well as the observed synergistic combination of 0.5 μg/mL polymyxin B and 1 μg/mL CBD.

Figure 4

Percent Inhibition of both mono-treatment at MIC and co-treatment at synergistic concentrations as determined through checkerboard screening (SEM # = 0.059, ## = 0.018, ### = 0.006, #### = 0.062).

Figure 5

Synergistic activity of CBD and Antibiotics (Ampicillin (AMP), Kanamycin (KAN), Polymyxin-B (POLY-B)) against S. typhimurium. Isobologram showing the synergy (FICI < 0.5), partial synergy (0.5 ≤ FICI ≥ 0.75), additive (0.76 ≤ FICI ≥ 1), indifference (1 ≤ FICI ≥ 4), or antagonism (FICI > 4) effects of CBD paired with broad-spectrum antibiotics against S. typhimurium. Representative data for determination of FICI is included.

Figure 6

Comparative efficacy of CBD-antibiotic co-treatment and MIC antibiotic mono-treatment against S. typhimurium. Three co-treatments examined include ampicillin (0.5 μg/mL)-CBD (1 μg/mL) (A), kanamycin (5 μg/mL)-CBD (1 μg/mL) (B), and polymyxin B (0.5 μg/mL)-CBD (1 μg/mL) (C).

Figure 6

Comparative efficacy of CBD-antibiotic co-treatment and MIC antibiotic mono-treatment against S. typhimurium. Three co-treatments examined include ampicillin (0.5 μg/mL)-CBD (1 μg/mL) (A), kanamycin (5 μg/mL)-CBD (1 μg/mL) (B), and polymyxin B (0.5 μg/mL)-CBD (1 μg/mL) (C).