A-059. The Effect of Manuka Honey on the Activity of Bacteriolytic Cell Wall Enzymes in MRSA

R. E. Jenkins, N. F. Burton, R. A. Cooper;
Univ. of Wales Inst. Cardiff, Cardiff, UNITED KINGDOM.

Background: Manuka honey has been shown to affect the cell cycle of MRSA by impeding cell division, but the precise mode of action is unknown. Cell division depends on the formation of septa and cell separation is facilitated by the cleavage of peptidoglycan. This study was designed to determine whether manuka honey affected the activity of bacteriolytic enzymes involved in the hydrolysis of peptidoglycan. Methods: EMRSA-15 (NCTC 13142) was cultivated at 37°C for up to 24 hours in TSB with 10% (w/v) manuka honey, as well as 10% (w/v) artificial honey. Cells were harvested at known time intervals (0, 60, 120, 240, 720, 1080 and 1440 minutes), washed and cell free extracts were prepared with a French press (intracellular enzymes). Culture supernatants (extracellular enzymes) were also collected at each time point and concentrated using Centicon 10 (Millipore). Bacteriolytic activity of cell free extracts and culture supernatants was determined by a cell wall turbidity assay using Micrococcus lysodeiktikus cell walls and 200 ug/ml protein from each sample. It was also determined using zymography gels containing 1mg/ml M. lysodeikticus cell wall with 15 ug protein of each sample. Results: Activity of extracellular bacteriolytic enzymes was at undetectable levels in turbidity assays and zymographs. Intracellular enzymes of untreated cells and of artificial honey treated cells showed similar patterns of bacteriolytic activity in both tests. Manuka honey treated cells exhibited markedly reduced activity compared to controls in cell wall assays and zymographs. After 24 hours turbidity results from manuka honey treated cells compared to control cells were significantly different (p = 0.001). Conclusion: Activity of bacteriolytic cell wall enzymes was reduced in MRSA cells exposed to bactericidal concentrations of manuka honey.
This study was supported by the British Society for Antimicrobial Chemotherapy.