Abstract
Two diterpenoids, salvipisone (Salv) and aethiopinone (Aeth), isolated from hairy
roots of Salvia sclarea, were tested with respect to their activity against methicillin-resistant Staphylococcus aureus (MRSA) and S. epidermidis (MRSE) strains, cultured as planktonic cells or as adherent biofilms. The standard
CLSI method, MTT reduction assay or confocal laser scanning microscopy (CLSM) were
used for this purpose and also applied for testing the susceptibility to oxacillin,
vancomycin, linezolid and their potential synergy with diterpenoids (evaluated as
a fractional inhibitory concentration (FIC) index). Salv and Aeth were shown to be
bactericidal or bacteriostatic against S. aureus and S. epidermidis planktonic cultures. Both diterpenoids, at the concentrations of œ MIC, showed synergy
with antibiotics representing the β-lactam, glycopeptide and oxazolidinone groups.
None of the antibiotics used at a high concentration killed the staphylococcal biofilms.
On the contrary, Salv and Aeth decreased the number of live biofilm cells by 45.7
- 77.1 % and slightly reduced the biofilm inhibitory concentration of oxacillin. Diterpenoids
also changed the parameters of biofilm morphology, as shown by the CLSM image processing
package (PHLIP). It was concluded that salvipisone and aethiopinone (relatively highly
lipophilic, log P respectively = 3.4; 4.8) synergized the action of β-lactam antibiotics towards MRSA
and MRSE probably by alteration of cell surface hydrophobicity and cell wall/membrane
permeability, but not by changing penicillin-binding protein, PBP2a expression and
penicillinase production or by direct binding to the cell wall peptidoglycan and teichoic
acids.
Abbreviations
Aeth:aethiopinone
ATCC:American Type Culture Collection
CAM:calcein-AM
CLSI:Clinical and Laboratory Standards Institute
CLSM:confocal laser scanning microscopy
CSH:cell surface hydrophobicity
DMSO:dimethyl sulfoxide
FIC:fractional inhibitory concentration
MHB:Mueller-Hinton broth
MIC:minimal inhibitory concentration
MRSA/MSSA:methicillin resistant/susceptible Staphylococcus aureus
MRSE/MSSE:methicillin resistant/susceptible S. epidermidis
MTT:3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide
OD:optical density
PBP:penicillin binding protein
PG:peptidoglycan
PI:propidium iodide
Salv:salvipisone
SAT:salt aggregation test
TA:teichoic acids
TSB:tryptic soy broth
Key words
Salvia sclarea
- Lamiaceae - hairy roots - salvipisone - aethiopinone - biofilm -
Staphylococcus
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Prof. Dr. hab. Barbara Rozalska
Department of Immunology and Infectious Biology
Institute of Microbiology and Immunology
University of Łodz
Banacha 12/16
90-237 Łodz
Poland
Phone: +48-42-635-4472
Fax: +48-42-665-5818
Email: rozab@biol.uni.lodz.pl