In vitro evaluation of metal chelators as potential
metallo--lactamase inhibitors
Rafiatu Azumaha, b, Jyotibon Duttaa, Anou M. Somboroa, b, Melissa Ramtahala, b, Louis Choncod, Raveen Parboosingd, Linda
A. Besterc, Hendrik G. Krugera, Tricia Naickera, Sabiha Y. Essackb*, Thavendran Govendera*
aCatalysis
and Peptide Research Unit, bAntimicrobial Research Unit, cBiomedical Resource Unit, University of KwaZulu-Natal, Durban, South
Africa dDepartment of Virology, National Health Laboratory Service, Durban, South Africa
Introduction & Purpose:
The indiscriminate use of antibiotics has resulted in
bacterial resistance, which is a major concern to public
health. Enterobacteriaceae, especially, carbapenemresistant Enterobacteriaceae (CRE) are increasingly
emerging as common pathogens worldwide with
mortality rates of >50% reported in some cases (1).
Most antibiotics in clinical use are now ineffective
against CREs with polymixins being the sole remaining
therapeutic options in many environments (2,3). In the
absence of new antibiotics, this study evaluated metal
chelators as potential class B, metallo-β-lactamase
(MBL) inhibitors.
Methods :
The minimum-inhibitory concentration (MIC) of
meropenem was ascertained alone and in combination
with various concentrations of macrocyclic (NODAGA
peptide derivatives) and acyclic (TPEN and DPA)
metal chelators against MBL producing CREs using
the broth microdilution method. The combined effect of
meropenem and the various inhibitors was determined
for 12 bacterial isolates producing the MBL enzyme
using the checkerboard method.
Results:
L-1
L-1
TPEN at 4 mg
and 8 mg
showed the best
activity by decreasing meropenem MICs to 0.5 mg L-1
and 0.06 mg L-1 respectively for some isolates
producing the NDM -1 & 4 and VIM -1 enzymes. DPA
at 8 and 16 mg L-1 was also able to decrease
meropenem MICs to 1 and 0.125 mg L-1 respectively
for these CREs. NODAGA peptide derivatives showed
the least inhibition with concentrations as high as 32
mg L-1 required for meropenem MICs to be decreased
to 0.06 mg L-1 against an NDM-1 producing isolate.
Table 1. Comparison of different metal chelators in
combination with meropenem as potential metallo-βlactamase inhibitors against NDM-1-producing E. coli.
MIC (mg L-1)
Drug / Compound
Drug / Compound alone
meropenem /Compound
Meropenem
>16
-
NOTA
>64
0.12a/4b
NODAGA-4
>64
4/161 or 0.06/322
NODAGA-8
>64
4/16 or 0.06/32
DPA
>64
4/8 or 1/16
TPEN
32
1/4 or 0.06/8
References :
Table 2. Comparison of MICs of meropenem (MEM),
inhibitors (TPEN and DPA) and their combinations
against other reference MBL-producing CRE strains
-1
MIC (mg L )
CRE strain
1.
Escherichia coli NDM - 1
MEM
alone
>16
TPEN
alone
32
DPA
alone
>64
MEM/TPEN
1a/4b or 0.06/8
MEM/DP
A
4/81 or 1/162
2.
Citrobacter freundii NDM - 1
8
32
>64
0.5/4 or 0.06/8
2/4 or 0.5/8
3.
Providencia rettgeri NDM - 1
>16
>64
>64
2/4 or 1/16
4/8 or 2/64
4.
Providencia stuartii NDM - 1
>16
>64
>64
4/4 or 2/8
8/8 or 2/16
5.
Enterobacter cloacae NDM - 1
>16
32
>64
0.5/4 or 0.12/8
1/8 or 0.5/16
6.
Escherichia coli NDM - 4
>16
64
>64
1/4 or 0.25/8
4/16 or 1/32
7.
Escherichia coli VIM - 1
>16
>64
>64
4/4 or 2/8
4/8 or 1/16
8.
Enterobacter cloacae VIM - 1
8
>64
>64
1/4 or 0.12/8
2/8 or 0.5/16
9.
Klebsiella pneumoniae VIM -1
>16
32
>64
0.5/4 or 0.12/8
1/8 or 0.12/8
10.
Escherichia coli IMP - 8
16
32
>64
1/8 or 0.06/16
8/32 or 8/64
11.
Enterobacter cloacae IMP - 8
>16
64
>64
12.
Klebsiella pneumoniae IMP - 8
8
>64
>64
13.
Escherichia coli
0. 03
32
>64
0.5/8
or 16/32
or
0.06/16
16/64
2/16 or 0.5/32
8/32 or 8/64
-
-
ATCC 25922 (Control Strain)
a
Conclusion :
MICs of meropenem against some MBL
producing
CREs
were
decreased
to
concentrations as low as 0.06 mg L-1 in the
presence of some metal chelators.
TPEN, DPA and NODAGA peptide derivatives
were able to inhibit the MBLs in decreasing order
of activity, rendering CREs susceptible to
meropenem.
and b represents the MIC of meropenem and inhibitor
respectively. 1 and 2 represent MICs of meropenem at different
concentrations of the inhibitors in both tables.
1. Falagas, M.E., Lourida, P., Poulikakos, P., Rafailidis,
P.I. and Tansarli, G.S. (2013) Antibiotic treatment of
infections
due
to
carbapenem-resistant
Enterobacteriaceae: systematic evaluation of the
available evidence. Antimicrob Agents Chemother,
AAC. 01222-01213.
2. Livermore, D.M., Warner, M., Mushtaq, S., Doumith, M.,
Zhang, J. and Woodford, N. (2011) What remains
against carbapenem-resistant Enterobacteriaceae?
Evaluation of chloramphenicol, ciprofloxacin, colistin,
fosfomycin, minocycline, nitrofurantoin, temocillin and
tigecycline. Int J Antimicrob Ag 37, 415-419.
3. Bush, K. (2013b) Carbapenemases: Partners in crime.
J Glob Antimicrob Resist 1, 7-16.
Acknowledgments:
The authors are grateful to the College of Health
Sciences, University of Kwa-Zulu Natal & the South
African National Research Foundation for funding the
study.
Disclosure:
Professor Essack is a member of the Global
Respiratory Infection Partnership sponsored by Reckitt
& Benckiser