Introduction

Efflux pump in bacteria is a significant cause of antibiotic resistance, making the antibiotic efflux pumps as the therapeutic targets. Currently there are 4 multidrug efflux efflux pumps, out of which the AcrAB-TolC system is the most prevalent and constitutively expressed in Escherichia coli. The use of efflux pump inhibitors to improve the susceptibility of antibiotics against infections in clinical isolates has been investigated. However, their use in clinical practice is limited. A recent clinical case report has demonstrated the use of EDTA in ceftriaxone plus sulbactam combination.

Aim

The efficacy of EDTA alone and in combination with various drugs on AcrAB-TolC efflux pump has been evaluated.

Methods

Study Design

• In-vitro study  

Treatment Strategy

• A total of 23 clinical isolates of E coli collected from patients with urinary tract infections, bacteremia and pneumonia were included
• The antimicrobial agents used in the study were as follows
  • A novel antibiotic adjuvant entity (AAE) with ceftriaxone sodium and Sulbactam plus 10 mM EDTA referred to as CSE1034
  • Ceftriaxone
  • Amoxicillin plus clavulanate
  • Piperacillin plus tazobactam
  • Meropenem
  • Cefoperazone plus sulbactam
• The effect of EDTA on 4,6-diamidino-2-phenylyindole (DAPI) efflux activity was measured.
• The minimum inhibitory concentration (MIC) of each antimicrobial drug with and without EDTA was assessed
• Using reverse transcriptase polymerase chain reaction (RT-PCR), the effect of EDTA and half with half of MIC on mRNA expressions of AcrAB-TolC efflux pump was estimated

Endpoints

• Reduction in MIC
• mRNA expression
• mRNA levels

Results

• Fifteen clinical isolates were found to be positive for AcrAB-TolC efflux pump
• The MIC values reduced by 32 and 8-fold with 10 mM EDTA combined with ceftriaxone plus sulbactam and meropenem respectively
• The other drugs demonstrated a 2-fold reduction in the MIC
• A concentration-dependent inhibition of DAPI efflux was demonstrated with a significant inhibition at 10 mM
• Further increase in the concentration of EDTA did not produce any more inhibition
• The expression of AcrA, AcrB and TolC mRNA was decreased by 2.3, 2.2 and 2.4 folds by 10 mM EDTA
• The CSE 1034 group had significant reduction in mRNA levels of AcrAB-TolC as compared to meropenem; p<0.0001
• No significant changes were observed with the other drugs; p>0.05

Conclusion

• Ceftriaxone plus sulbactam plus EDTA significantly reduced the efflux pump expression and can be considered as an alternative in treating the E coli infections with efflux over expressed strains.
• EDTA disodium increases the susceptibility of E coli efflux over expressed strains by inhibiting efflux pumps.

J Microb Biochem Technol. 2012;4(6). Doi:10.4712/1948-5948.1000082.