81 Detection of AmpC beta-lactamases in E. coli using different screening agars Introduction Over the past decades, the importance of resistant Enterobacteriales increased substantially due to the emergence of various resistance traits that inactivate most beta-lactam antibiotics. In Escherichia coli, resistance to 2nd and 3rd generation cephalosporins is most frequently due to the occurrence of plasmid-encoded ESBL, as well as both plasmid-encoded (pampC) and chromosomally-encoded (campC) AmpC genes. As the rapid and accurate detection of resistant bacteria is crucial for treatment and control, selective media for detection of extended-spectrum beta-lactamase-producing Enterobacteriales (ESBL-E) have been developed. A considerable number of studies were published on the performance of ESBL-E screening agars (Glupczynski et al. 2007; Réglier-Poupet et al. 2008; Randall et al. 2009; Al Naiemi et al. 2009; Overdevest et al. 2011). So far, few studies have focused on screening media for AmpC beta-lactamaseproducing Enterobacteriales (AmpC-E). The basis of most agars that are selective for ESBL is a adding a 3rd generation cephalosporin, e.g. cefotaxime or ceftazidime. Some ESBL-E screening agars use AmpC inhibitors, such as cloxacillin to suppress Enterobacteriales that intrinsically produce AmpC beta-lactamase on high level (e.g. Enterobacter spp) (Randall et al. 2009; Al Naiemi et al. 2009; Overdevest et al. 2011). Overgrowth of such species with intrinsic AmpC beta-lactamase hyperproduction makes it more difficult to screen for E. coli with acquired beta-lactamases, which may lead to a lower sensitivity of the ESBL-E screening agars (Randall et al. 2009). E. coli isolates produce chromosomally encoded AmpC (cAmpC) constitutively, but only on a low level. However, they may acquire alterations in the promoter/attenuator region leading to hyperproduction of cAmpC betalactamase (Tracz et al. 2005; 2007). Growth of both cAmpC and pAmpC producing Enterobacteriales is inhibited by cloxacillin. Cloxacillin containing agars may improve the yield of ESBL producing E. coli but they lead to a lower yield of both pAmpC and cAmpC beta-lactamase producing E. coli. One option to solve this problem could be to include an additional agar specifically developed for AmpC producing E. coli during screening. According to the EUCAST guidelines combined resistance to 3rd generation cephalosporins and cefoxitin, a cephamycin antibiotic, may be used as phenotypic criteria for AmpC screening (Martinez and Simonsen 2017). Cefoxitin is highly active against ESBL producing bacteria, but has little activity against most AmpC producing bacteria (Jacoby 2009; Birnbaum et al. 1978). This makes cefoxitin useful in screening strategies for AmpC (Ingram et al. 2011; Polsfuss et al. 2011; Peter-Getzlaff et al. 2011). 5
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