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Artesunate enhances the antibacterial effect of {beta}-lactam antibiotics against Escherichia coli by increasing antibiotic accumulation via inhibition of the multidrug efflux pump system AcrAB-TolC.

OBJECTIVES
Occasionally, we found that artesunate enhanced the antibacterial effects of antibiotics in vitro. Therefore, the enhancement of various β-lactam antibiotics by artesunate against Escherichia coli and the possible mechanism were investigated in the present study.


METHODS
Antibacterial effects were observed using the serial 2-fold dilution method and dynamic bacterial growth. Daunomycin accumulation within E. coli was observed using fluorospectrophotometry and laser confocal scanning microscopy. AcrAB-TolC, AmpC and TEM-1 mRNA expression was observed using a PCR method. Antisense oligonucleotides (as-ODNs) targeting AcrB were designed and used to block AcrB gene expression within E. coli ATCC 35218.


RESULTS
Although artesunate itself had no antibacterial ability, artesunate significantly increased the antibacterial effect of β-lactam antibiotics against E. coli ATCC 35218 and an E. coli clinical strain. Artesunate increased daunomycin accumulation within E. coli ATCC 35218 in a dose-dependent manner and reduced the mRNA expression of AcrAB-TolC, an important multidrug efflux system for Gram-negative bacteria. The bacterial number was significantly reduced by as-ODN targeting AcrB, but did not further decrease after additional artesunate treatment. In contrast, artesunate lost its enhancement of β-lactam antibiotics against E. coli AG100A, a strain lacking the gene encoding AcrAB, and artesunate did not increase daunomycin accumulation within E. coli AG100A. After the transformation of pET28a-AcrB into E. coli AG100A, artesunate regained enhancement of β-lactam antibiotics. Furthermore, artesunate did not inhibit the expression of AmpC and TEM-1 mRNA.


CONCLUSIONS
Artesunate enhances the antibacterial effect of various β-lactam antibiotics against E. coli, which might be associated with the suppression of a major multidrug resistance system, AcrAB-TolC.