Evaluation of phenotypic tests for the detection of AmpC beta-lactamase in clinical isolates of Escherichia coli.

Background: AmpC beta lactamases are cephalosporinases that confer resistance to a wide range of beta lactam drugs thereby causing serious therapeautic problem. As there are no CLSI guidelines for detection of AmpC mediated resistance in Gram negative clinical isolates and it may pose a problem due to misleading results, especially so in phenotypic tests. Although cefoxitin resistance is used as a screening test, it does not reliably indicate AmpC production. Materials and Methods: We planned a study to determine the occurrence of AmpC beta lactamase in hospital and community, clinical isolates of Escherichia coli and simultaneously evaluate different phenotypic methods for detection of AmpC beta lactamases. Results: It was observed that 82.76% isolates were ESBL positive and 59% were cefoxitin screen positive. Using phenotypic confi rmatory tests the occurrence of Amp C beta lactamases was found to be 40% and 39% by inhibitor based method using boronic acid (IBM) and modifi ed three dimensional test (M3D) respectively. Conclusion: Both the test showed concordant result. Co-production was observed in 84.62% isolates Screening of ESBL and Amp C can be done in routine clinical microbiology laboratory using aztreonam and IBM respectively as it is a simple, rapid and technically less demanding procedure which can be used in all clinical laboratories.

An Increase in the Clinical Isolation of Acquired AmpC beta-Lactamase-Producing Klebsiella pneumoniae in Korea from 2007 to 2010

We investigated the occurrence and genetic basis of AmpC beta-lactamase (AmpC)-mediated antibiotic resistance, by examining Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis isolates at a university hospital, from 2007 to 2010. The ampC genes were detected by multiplex AmpC PCR, and AmpC-positive strains were subjected to DNA sequencing. Extended-spectrum beta-lactamase (ESBL) production was assessed using the ESBL disk test based on the utilization of boronic acid. Carbapenem-resistant isolates were further investigated by the modified Hodge test, a carbapenemase inhibition test and SDS-PAGE experiments. AmpC expression was detected in 1.6% of E. coli (39 DHA-1, 45 CMY-2, and 1 CMY-1) isolates, 7.2% of K. pneumoniae (39 DHA-1, 45 CMY-2, and 1 CMY-1) isolates, and 2.5% of P. mirabilis (8 CMY-2 and 1 CMY-1) isolates. Of the 198 acquired AmpC producers, 58 isolates (29.3%) also produced an ESBL enzyme. Among the acquired AmpC-producing K. pneumoniae isolates, the minimum inhibitory concentration (MIC) MIC50/MIC90 values for cefoxitin, cefotaxime, cefepime, imipenem, and meropenem were >32/>32, 16/>32, 1/16, 0.25/0.5, and or =2 microg/mL for 2 K. pneumoniae isolates, both of which carried the blaDHA-1 gene with a loss of OmpK36 expression, but were negative for carbapenemase production. The acquisition of AmpC-mediated resistance in K. pneumoniae isolates increased, as did the proportion of AmpC and ESBL co-producers among the hospital isolates. The accurate identification of isolates producing AmpCs and ESBLs may aid in infection control and will assist physicians in selecting an appropriate antibiotic regimen.

Celulitis facia odontogénica severa infrecuent causada por citrobacter freunddii productora de ampc en un paciente con diabetes mellitus 2. reporte de caso / Uncmmon severe oodontogenic facial cellulitis caused by ampcproducing citrobacter freundii in a patient with type 2 diabetes mellitus. case report / Celulite facial odontogênica sevea infrequente causada por citrobacter freundii produtora de ampc em um paciente com diabetes mellitus 2. relatório e caso

El manejo de la celulitis facial odontogénica no deja de ser un tema controversial en el campo de la cirugía oral y maxilofacial; los principios quirúrgicos y terapéuticos han sido sometidos a modificacio nes basadas en los hallazgos clínicos, imagenológicos y microbiológicos a través del tiempo. En pacientes con diabetes mellitus 2 se incrementa el riesgo a sufrir infecciones bacterianas oportunistas con tiempos de hospitalización más prolongados que la población no diabética. La literatura es clara estableciendo las diferencias clínicas y microbiológicas de la celulitis facial odontogénica en este grupo de pacientes, sin embargo, no existe un protocolo médico quirúrgico destinado a ellos. El microorganismo comúnmente aislado es Klebsiella pneumoniae, mientras Citrobacter freundii es inusual en las infecciones odontogénicas, su capacidad para producir betalactamasas de amplio espectro (AmpC) le permite bloquear la acción de los antibióticos de uso empírico en Cirugía Oral y Maxilofacial. A continuación, presentamos el caso de una paciente de 61 años con diabetes Mellitus tipo 2 y celulitis facial odontogénica por Citrobacter freundii productora de AmpC.

The management of odontogenic facial cellulitis is still a controversial issue in the field of Oral and Maxillofacial Surgery. Surgical and therapeutic principles have undergone modifications based on clinical findings, imaging and microbiological over time. In patients with type 2 Diabetes Mellitus the risk of opportunistic bacterial infections is increased thus suffering longer hospitalization periods than the nondiabetic population. The literature is clear by setting the clinical and microbiological differences of odontogenic facial cellulitis in this group of patients, but there is no surgical medical protocol for them. Klebsiella pneumoniae is the most common microorganism isolated while Citrobacter freundii is unusual in relation to oral infections; their ability to produce ESBLs (AmpC) allows them to block the action of empirical antibiotics used in Maxillofacial Surgery. We present the case of a 61 year old patient with type 2 Diabetes Mellitus and odontogenic facial cellulitis caused by AmpCproducing Citrobacter freundii.

O tratamento da celulite facial odontogênica não deixa de ser um tema controverso no campo da Cirurgia Oral e Maxilofacial; os princípios cirúrgicos e terapêuticos foram submetidos a modificações baseadas nos descobrimentos clínicos, imagenológicos e microbiológicos através do tempo. Em pacientes com Diabetes Mellitus 2 aumenta o risco de sofrer infecções bacterianas oportunistas com tempos de hospitalização mais prolongados que na população não diabética. A literatura é clara estabelecendo as diferenças clínicas e microbiológicas da Celulite Facial Odontogênica neste grupo de pacientes; porém, não existe um protocolo médico cirúrgico destinado a eles. O microrganismo comunmente isolado é o Klebsiella pneumoniae, enquanto que o Citrobacter freundii é inusual nas infecções odontogênicas, sua capacidade para produzir betalactamases de amplo espectro (AmpC) lhe permite bloquear a ação dos antibióticos de uso empírico em Cirurgia Oral e Maxilofacial. A seguir apresentamos o caso de uma paciente de 61 anos com Diabetes Mellitus tipo 2 e celulite facial odontogênica por Citrobacter freundii produtora de AmpC.

Evaluation of the MicroScan NegCombo Panel Type 44 for Detection of Extended-Spectrum beta-Lactamase among Clinical Isolates of Escherichia coli, Klebsiella species, and Proteus mirabilis / 대한진단검사의학회지

BACKGROUND: Accurate and rapid detection of extended-spectrum beta-lactamases (ESBLs) is important in guiding proper antimicrobial therapy for infected patients. We evaluated the performance of MicroScan NegCombo Type 44 panel (Dade Behring, USA), which was developed to confirm ESBL-producing Enterobacteriaceae using ceftazidime/clavulanate and cefotaxime/clavulanate. METHODS: From August 30 to September 20, 2007, 206 non-duplicate clinical isolates, including 106 Escherichia coli, 81 Klebsiella pneumoniae, 11 Klebsiella oxytoca, and 8 Proteus mirabilis were subcultured and tested with Type 32 and Type 44 panels. The results were compared with those of the CLSI phenotypic confirmatory test (CLSI-PCT) and disk approximation test (DAT). Isolates not susceptible to cefotetan or flagged as "Possible ESBL, unable to interpret confirm test (Possible ESBL)" on Type 44 panel were tested with boronic acid disks to confirm AmpC beta-lactamases (AmpC) production. RESULTS: Of the 206 isolates tested, 44 (21.4%) produced ESBL by CLSI-PCT or DAT, including 27 E. coli, 14 K. pneumoniae, 2 K. oxytoca, and 1 P. mirabilis. Thirty-eight isolates flagged as "Confirmed ESBL" on Type 44 panel were all confirmed as ESBL-producers. Of 14 K. pneumoniae flagged as "Possible ESBL", 6 were confirmed as ESBL and AmpC co-producers and 8 as AmpC-producers. CONCLUSIONS: Type 44 panel showed an excellent performance in detecting ESBL-producing E. coli, Klebsiella spp., and P. mirabilis. When flagged as "Confirmed ESBL", no other confirmatory test was necessary to report as ESBL; however, "Possible ESBL" required a differential test for AmpC production.

New Method for Detecting Plasmid-mediated AmpC Beta-lactamases / 中华医院感染学杂志

OBJECTIVE To establish a convenient method for detecting plasmid-mediated AmpC beta-lactamases in Enterobacteriaceae lacking chromosomal AmpC beta-lactamases and to investigqte their genotype.METHODS Fifteen isolates of Klebsiella pneumoniae were collected and the diameters of inhibitory zone of cefoxitin in combination with or without cloxacillin were measured separately by standard disc diffusion test.The M3D assay was used as control method.And the activity of AmpC beta-lactamases of all isolates was simultaneously assayed.Multiplex PCR was performed to determine the genotype of AmpC beta-lactamases.RESULTS Among 15 isolates,8 isolates were identified to have plasmid-mediated AmpC beta-lactamases.The sensitivity and specificity of AmpC beta-lactamases phenotypic confirmatory test were 100%.Eight of 15 isolates were identified to be DHA-1 beta-lactamases by multiplex PCR.CONCLUSIONS The new AmpC beta-lactamases phenotypic confirmatory test is a reliable method for detecting plasmidmediated AmpC beta-lactamases in Enterobacteriaceae lacking chromosomal AmpC beta-lactamases.DHA-1 beta-lactamases are the main genotypes of plasmid-mediated AmpC beta-lactamases in Hubei Province of China.

Plasmid-encoded AmpC beta-lactamases: how far have we gone 10 years after the discovery?

The dogma that ampC genes are located exclusively on the chromosome was dominant until about 10 years ago. Since 1989 over 15 different plasmid-encoded AmpC beta-lactamases have been reported from several countries. Most of these enzymes evolved in two clusters. The major cluster includes several enzymes with a high similarity to CMY-2, which is the closest related chromosomal AmpC enzyme of Citrobacter freundii. A second cluster centers around CMY-1. It is less homogeneous and not closely related chromosomal AmpC enzymes. Molecular diversification by amino acid substitutions does not usually translate into a change in the resistance phenotype. At this time, CMY-2 appears to be the most prevalent and widely distributed. Further global increase of prevalence and diversity of plasmidic AmpC beta-lactamases have to be anticipated in the next millenium.