Antibacterial and proteomic profiling of Morus alba extract against methicillin-resistant Staphylococcus aureus

Antimicrobial resistance, particularly from methicillin-resistant Staphylococcus aureus (MRSA), is a growing global health threat. Alternative therapies derived from medicinal plants are gaining attention for their potential to combat resistant pathogens. This study aimed to evaluate the antibacterial activity of Morus alba (white mulberry) extracts and investigate their action mechanisms using proteomic analysis.

Leaf and stem samples of M. alba were extracted using both decoction and maceration techniques with water and ethanol as solvents. The antibacterial activity against MRSA was assessed through minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and time-kill assays. Chemical profiling of the most active extract was performed using liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (LC-QTOF-MS/MS). Proteomic analysis was conducted to explore changes in bacterial protein expression after treatment.

The ethanol extract of M. alba stem exhibited the strongest antibacterial activity, with MIC values ranging from 0.3125 to 20 mg/mL and MBC values from 0.6250 to 40 mg/mL. A time-kill assay demonstrated that bacterial counts fell below the detection limit within 4 hours at four times the MIC concentration, based on three independent replicates. LC-QTOF-MS/MS profiling identified betulinic acid as the most abundant compound in the extract. Proteomic analysis revealed significant changes in MRSA protein expression, including upregulation of GlmU, N-acetylneuraminate lyase, and ribonuclease E, and downregulation of ribose-phosphate pyrophosphokinase and SecA. Pathway enrichment analysis suggested that the observed protein expression changes are consistent with enhanced N-acetylneuraminate catabolism and RNA polymerase activity, and suppression of protein export and fatty acid biosynthesis.

These findings highlight the strong anti-MRSA potential of M. alba stem extract and provide mechanistic insights into its antibacterial action. The extract disrupts critical metabolic and regulatory pathways in MRSA, supporting its potential development as a novel antimicrobial agent.