Streptococcus mutans (S. mutans) is a gram-positive facultatively anaerobic bacterium and the most common pathogen associated with tooth caries. The organism is acid tolerant and can undergo physiological adaptation to function effectively in acid environments such as carious dental plaque. Some cannabinoids have been found to have potent anti-microbial activity against gram-positive bacteria. One of these is the non-psychoactive, minor phytocannabinoid Cannabigerol (CBG). Here we show that CBG exhibits anti-bacterial activities against S. mutans. CBG halts the proliferation of planktonic growing S. mutans, which is affected by the initial cell density. High-resolution scanning electron microscopy showed that the CBG-treated bacteria become swollen with altered membrane structures. Transmission electron microscopy provided data showing that CBG treatment leads to intracellular accumulation of membrane structures. Nile red, DiOC2(3) and laurdan staining demonstrated that CBG alters the membrane properties, induces membrane hyperpolarization, and decreases the membrane fluidity. CBG-treated bacteria showed increased propidium iodide uptake and reduced calcein AM staining, suggesting that CBG increases the membrane permeability and reduces the metabolic activity. Furthermore, CBG prevented the drop in pH caused by the bacteria. In summary, we present here data showing the mechanisms by which CBG exerts its anti-bacterial effect against S. mutans.
Streptococcus mutans is a common cariogenic bacterium in the oral cavity involved in plaque formation. Previous studies showed that Cannabigerol (CBG) has bacteriostatic and bacteriocidic activity against S. mutans. The aim of the present study was to study its effect on S. mutans biofilm formation and dispersion. S. mutans was cultivated in the presence of CBG, and the resulting biofilms were examined by CV staining, MTT assay, qPCR, biofilm tracer, optical profilometry, and SEM. Gene expression was determined by real-time qPCR, extracellular polysaccharide (EPS) production was determined by Congo Red, and reactive oxygen species (ROS) were determined using DCFH-DA. CBG prevented the biofilm formation of S. mutans shown by reduced biofilm biomass, decreased biofilm thickness, less EPS production, reduced DNA content, diminished metabolic activity, and increased ROS levels. CBG altered the biofilm roughness profile, resulting in a smoother biofilm surface. When treating preformed biofilms, CBG reduced the metabolic activity of S. mutans with a transient effect on the biomass. CBG reduced the expression of various genes involved in essential metabolic pathways related to the cariogenic properties of S. mutans biofilms. Our data show that CBG has anti-biofilm activities against S. mutans and might be a potential drug for preventive treatment of dental caries.
Cannabigerol (CBG) is a non-psychoactive cannabinoid naturally present in trace amounts in the Cannabis plant. So far, CBG has been shown to exert diverse activities in eukaryotes. However, much less is known about its effects on prokaryotes. In this study, we investigated the potential role of CBG as an anti-biofilm and anti-quorum sensing agent against Vibrio harveyi. Quorum sensing (QS) is a cell-to-cell communication system among bacteria that involves small signaling molecules called autoinducers, enabling bacteria to sense the surrounding environment. The autoinducers cause alterations in gene expression and induce bioluminescence, pigment production, motility and biofilm formation. The effect of CBG was tested on V. harveyi grown under planktonic and biofilm conditions. CBG reduced the QS-regulated bioluminescence and biofilm formation of V. harveyi at concentrations not affecting the planktonic bacterial growth. CBG also reduced the motility of V. harveyi in a dose-dependent manner. We further observed that CBG increased LuxO expression and activity, with a concomitant 80% downregulation of the LuxR gene. Exogenous addition of autoinducers could not overcome the QS-inhibitory effect of CBG, suggesting that CBG interferes with the transmission of the autoinducer signals. In conclusion, our study shows that CBG is a potential anti-biofilm agent via inhibition of the QS cascade.
Background Streptococcus mutans is a common cariogenic bacterium in the oral cavity involved in plaque formation. Casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) has been introduced into tooth mousse to encourage remineralization of dental enamel. The aim of this research was to study the effect of tooth mousse containing CPP-ACP (GC Tooth Mousse®) or CPP-ACP with 0.2% fluoride (CPP-ACPF; GC Tooth Mousse Plus®; GCP) on S. mutans planktonic growth and biofilm formation. Methods S. mutans was cultivated in the presence of different dilutions of the tooth mousse containing CPP-ACP or CPP-ACPF, and the planktonic growth was determined by ATP viability assay and counting colony-forming units (CFUs). The resulting biofilms were examined by crystal violet staining, MTT metabolic assay, confocal laser scanning microscopy (CLSM), and scanning electron microscope (SEM). Results The CPP-ACP tooth mousse (GC) at a dilution of 5–50 mg/ml (0.5–5%) did not inhibit planktonic growth, and even increased the ATP content and the number of viable bacteria after a 24 h incubation. The same was observed for the CPP-ACPF tooth mousse (GCP), except for the higher concentrations (25 and 50 mg/ml) that led to a drop in the bacterial count. Importantly, both compounds significantly decreased S. mutans biofilm formation at dilutions as low as 1.5–3 mg/ml. 12.5 mg/ml GC and 6.25 mg/ml GCP inhibited biofilm formation by 90% after 4 h. After 24 h, the MBIC90 was 6.25 mg/ml for both. CLSM images confirmed the strong inhibitory effect GC and GCP had on biofilm formation when using 5 mg/ml tooth mousse. SEM images of those bacteria that managed to form biofilm in the presence of 5 mg/ml tooth mousse, showed alterations in the bacterial morphology, where the streptococci appear 25–30% shorter on the average than the control bacteria. Conclusion Our data show that the tooth mousse containing CPP-ACP reduces biofilm formation of the cariogenic bacterium S. mutans without killing the bacteria. The use of natural substances which inhibit biofilm development without killing the bacteria, has therapeutic benefits, especially in orthodontic pediatric patients.
Biofilms are complex bacterial structures in which bacterial cells thrive as a community. Many bacterial species, including pathogens, form biofilms of high complexity and adaptability to a wide range of environmental conditions. One example of these is Streptococcus mutans, a gram-positive bacterium that has been associated with caries. Cannabigerol, a non-psychoactive cannabinoid, has been shown to affect S. mutans biofilms. In order to better characterize the effect of cannabigerol on biofilms of S. mutans, this paper provides a series of computational assays for biofilm analysis, applied on confocal images of S. mutans biofilms treated with cannabigerol. Confocal images are ubiquitous in biofilm analysis—they are often used to visualize the complex structure and molecular composition of biofilm macrocolonies. In this article, we demonstrate how confocal imaging data can be used to reveal more comprehensive insights into biofilm structure and measure specific anti-biofilm effects. This is accomplished by a series of computational assays, each focusing on a different aspect of biofilm structure.
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