Extended spectrum AmpC and metallo-beta-lactamases in Serratia and Citrobacter spp. in a disc approximation assay

Abstract

Objectives: This study aimed to develop a novel model for detection of extended spectrum beta-lactamase (ESBL), AmpC and metallo-betalactamase (MBL) producing Serratia and Citrobacter species using cefoperazone sulbactam as well as other inducer-substrate combinations in a disc approximation assay. In the absence of molecular tools in developing countries, we attempted to standardize simple phenotypic techniques for detection of beta-lactamases to allow effective patient care in our countries. These techniques have been scarcely used in Serratia and Citrobacter spp., which are emerging as significant pathogens in our region. Methodology: Clinical isolates of Serratia and Citrobacter were tested for ESBL production. Cefoperazone (CP)/cefoperazone sulbactam (CPS), piperacillin (PIP)/piperacillin-tazobactam (TZP) and ceftazidime (CAZ)/ceftazidime-clavulanic acid (CAZ-CLAV) combinations were compared for their ability to detect ESBL producers phenotypically. Multi-drug resistant strains were further tested for detection of inducible/derepressed AmpC mutants by a disc approximation assay. Isolates were screened for MBL production by Imipenem (IMI). MBL production was confirmed using Ethylenediaminetetraacetic acid (EDTA) in a double disc synergy assay and Hodge test. Minimal inhibitory concentration (MIC) was performed for CP, CPS and IMI by agar dilution method for all isolates of Serratia and Citrobacter spp. Results: Thirty-three percent of isolates of Serratia spp. and 35.4% of Citrobacter spp. were ESBL producers. CPS was a more sensitive inducer of ESBL than TZP and CAZ/CAZ-CLAV. AmpC producers were detected in 25.6% of the isolates of Serratia spp. (40% inducible and 60% derepressed mutants) and in 35.4% of the isolates of Citrobacter spp. (33% inducible and 66% derepressed mutants). Six isolates (four class B and two class A) of Serratia and eight isolates (seven class B and one class A) of Citrobacter spp. were MBL producers. Multiple mechanisms co-existed in eight isolates of Serratia and 15 isolates of Citrobacter spp. CPS was more effective in identifying ESBLs and inducible AmpC producers as well as type 1 carbapenemases than TZP and CAZ-CLAV were able to identify inducible AmpC producers. Conclusions: The high prevalence of ESBL, AmpC, and MBL in Serratia and Citrobacter species in this study suggests that detection of these by phenotypic methods in the absence of more specific molecular tests should be actively considered in not only developing countries but also in the developed world as this approach can lead to timely and appropriate antibiotic treatment. CPS may be advised due to the triple advantage of detection of all three types of beta-lactamases.

Serratia Infections: from Military Experiments to Current Practice

“…In a study from India of isolates recovered from different types of clinical specimens from 2007 to 2008, 25.6% of Serratia isolates produced AmpCs, 40% of which were inducible, and 60% of these isolates were derepressed mutants. Besides S. marcescens, the authors did not identify the isolates to the species level (321).…”

“…A survey of S. marcescens isolates from 2006 to 2009 in Mexico revealed that 20.5% were ESBL producers, and all of the ESBLs were SHV-type enzymes (143). In India, Rizvi and others found that 33% of Serratia species recovered from various clinical specimens from 2007 to 2008 were ESBL producers; they did not determine the type of enzymes present and did not report which species of Serratia were present besides S. marcescens (321).…”

“…Another type of plasmid-encoded metallo-␤-lactamase, VIM, has been found in S. marcescens (422) and S. liquefaciens (271). A survey of Serratia species from clinical isolates from India in 2007 to 2008 found that 15.4% produced metallo-␤-lactamases, although the type of enzyme was not determined, and besides S. marcescens, the other Serratia species were not identified (321).…”

Chronic suppurative otitis media: a clinico-microbiological menace

“…An increase in a diameter of 5 mm was considered positive for ESBL detection. 7 Detection of inducible and derepressed AmpC betalactamase: Isolates resistant to ceftriaxone, cefixime, cefoperazone and cefoperazone-sulbactam were tested for AmpC production. Organism resistant to cefoperazone and cefoperazone-sulbactam combination were considered AmpC producers.…”

Incidence of Metallo Betalactamases Producing Pseudomonas aeruginosa in Burn Ward of a Tertiary Care Rural Hospital

“…There are several screening tests for detection of MBL production but no single test when used alone is specific for the same. 13 Rizvi et al 29 found DDST using EDTA better than Hodge test. Lee K et al also found that Hodge test was a simple screening method for MBL producing isolates, but occasional isolates showed false negative result.…”