Rhamnus prinoides Plant Extracts and Pure Compounds Inhibit Microbial Growth and Biofilm Formation

The increased prevalence of antibiotic resistance threatens to render all of our current antibiotics ineffective in the fight against microbial infections. Biofilms, or microbial communities attached to biotic or abiotic surfaces, have enhanced antibiotic resistance and are associated with chronic infections including periodontitis, endocarditis and osteomyelitis. The “biofilm lifestyle” confers survival advantages against both physical and chemical threats, making biofilm eradication a major challenge. A need exists for anti-biofilm treatments that are “anti-pathogenic”, meaning they act against microbial virulence in a non-biocidal way, leading to reduced drug resistance. A potential source of anti-biofilm, anti-pathogenic agents is plants used in traditional medicine for treating biofilm-associated conditions. My dissertation describes the anti-pathogenic, anti-biofilm activity of Rhamnus prinoides (gesho) extracts and specific chemicals derived from them. Rhamnus prinoides, an evergreen shrub native to east Africa, is used in the fermented beverages te’j and tella and to treat a variety of illnesses including atopic dermatitis. Gesho has antibacterial and antiplasmodial activity but little is known about its effect against microbial biofilms. Preliminary work determined that gesho leaf ethanol extracts inhibited Gram positive bacterial biofilm formation up to 99 percent without inhibiting microbial growth, suggesting an anti-pathogenic mechanism of activity. Fractionation, chemical analysis and activity screens identified ethyl 4-ethoxybenzoic acid (EEB) as a novel gesho-derived compound with anti-pathogenic anti-biofilm activity. Structure-activity analysis of EEB-related compounds identified 4-ethoxybenzoic acid (4EB) as a more potent anti-pathogenic compound against S. aureus biofilms. 4EB inhibited 70 percent S. aureus biofilm formation with minimal impact on planktonic cell viability. 4EB decreased the fraction of hydrophobic S. aureus cells in culture, potentially reducing surface colonization. Additionally, treatments of existing biofilms with a combination of 4EB and vancomycin synergistically decreased the viability of biofilm dwelling cells up to 85 percent when compared to vancomycin alone. Work with gesho stem extracts measured more than 90 percent reduction of dual-species biofilms comprised of the oral pathogens Streptococcus mutans and Candida albicans. Reduced biofilm formation correlated with inhibition of extracellular polysaccharide production. Overall, gesho extracts and gesho-derived compounds have potential for use in topical and oral hygiene products, for wound treatments and other anti-biofilm applications