Pronounced Antibacterial Effects of Rhodomyrtone, a Novel Antibiotic Candidate, on Streptococcus pneumoniae as Revealed by Proteomic and Metabolomic Analysis

Abstract

The emergence of antibiotic-resistant pathogenic bacteria is a healthcare problem worldwide. We evaluated the antimicrobial activity of rhodomyrtone, an acylphloroglucinol present in Rhodomyrtus tomentosa leaves, against the human Gram- positive pathogens, Streptococcus pneumoniae. The compound exhibited pronounced anti-pneumococcal activity against a broad collection of clinical isolates. We studied the effects at the molecular level by a combination of proteomics and metabolomics. The integration of proteomic and metabolomic analyses revealed alterations in enzymes and metabolites involved in different metabolic pathways including amino acid biosynthesis, nucleic acid biosynthesis, glucid, and lipid metabolism. Notably, the levels of two enzymes (glycosyltransferase and UTP-glucose-1-phosphate uridylyltransferase) and three metabolites (UDP-glucose, UDP-glucuronic acid and UDP-N-acetyl-D-galactosamine) participating in the synthesis of the pneumococcal capsule clearly diminished in cells exposed to rhodomyrtone. Rhodomyrtone-treated pneumococcal cells significantly possessed less amount of capsule, as measured by a colorimetric assay and visualized by electron microscopy. These findings reveal the utility of combining proteomic and metabolomic analyses to provide insight into phenotypic features of S. pneumoniae treated with this potential novel antibiotic. The ability of Rhodomyrtus tomentosa leaf extract and rhodomyrtone to prevent biofilm formation and eradicate mature biofilms was assessed. The extract and rhodomyrtone at 1/8 × MIC significantly inhibited biofilm formation in all clinical isolates (P < 0.05). The viability of 8-day biofilm-grown cells significantly decreased following the treatment with the extract and rhodomyrtone at 16 × MIC. 40-90% reduction in the bacterial adhesion and invasion to A549 human alveolar epithelial cells was observed after challenging with the extract and rhodomyrtone, compared with the control within 60 min. Increase in 90-99% phagocytosis of the bacterial cells by RAW264.7 macrophage cell line was detected following the treatment with the extract and rhodomyrtone at 1/2 × MIC, compared with the control. The results suggested potential medicinal benefits of the extract and rhodomyrtone for the treatment of pneumococcal infections. In conclusion, rhodomyrtone is a promising alternative antibacterial agent for the treatment of the infections caused by the Gram-positive pathogens including S. pneumoniae.