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The objective of the study was to assess the prevalence of plasmid-mediated quinolone resistance (PMQR) genes (qnrA, qnrB, qnrC, qnrD, qnrS, aac(6')-Ib-cr, qepA, and oqxAB) in a collection of 120 extended-spectrum β-lactamases (ESBLs)-producing enterobacteria and to characterize them. Overall, PMQR determinants were detected in 72 (60%) isolates (20 Escherichia coli, 32 Klebsiella pneumoniae, and 20 Enterobacter cloacae). PMQR frequencies were as follows: qnr genes (25.8%), oqxAB (21.6%), and aac(6')-Ib-cr variant (19.2%). Four qnr alleles were identified as qnrB1 (83.8%), qnrB4 (6.4%), qnrB2 (3.2%), and qnrS1 (6.4%). qnr genes were mainly detected in E. cloacae (50%), aac(6')-Ib-cr in E. coli (47.5%), and oqxAB in K. pneumoniae (65%). Overall, blaCTX-M-15 (90.3%) was the most prevalent blaESBL type followed by blaSHV-12 (6.4%) and blaSHV-27 (2.7%). Rates of mutations in gyrA and parC genes were 75% for E. coli, 72.8% for K. pneumoniae, and 50% for E. cloacae. Isolates with mutations in their quinolone resistance-determining regions exhibited high fluoroquinolones resistance levels compared to those with wild ones. Genetic study of PMQR-harboring isolates revealed a great genomic diversity among each Enterobacteriaceae species. Our findings indicate the high prevalence of PMQR determinants among ESBL-producing Enterobacteriaceae isolates from our hospital and their diffusion in various unrelated CTX-M-15-producing clones.

Healthcare-associated infections due to cefotaxime-resistant (CTX-R) Enterobacteriaceae have become a major public health threat, especially in intensive care units (ICUs). Often acquired nosocomially, CTX-R Enterobacteriaceae can be introduced initially by patients at admission. This study aimed to determine the prevalence and genetic characteristics of CTX-R Enterobacteriaceae-intestinal carriage in ICU patients, to evaluate the rate of acquisition of these organisms during hospitalization, and to explore some of the associated risk factors for both carriage and acquisition. Between December 2014 and February 2015, the 63 patients admitted in the ICU of Charles Nicolle hospital were screened for rectal CTX-R Enterobacteriaceae colonization at admission and once weekly thereafter to identify acquisition. CTX-R Enterobacteriaceae fecal carriage rate was 20.63% (13/63) at admission. Among the 50 non-carriers, 35 were resampled during their hospitalization and the acquisition rate was 42.85% (15/35). Overall, 35 CTX-R Enterobacteriaceae isolates were collected from 28 patients (25 Klebsiella pneumoniae, seven Escherichia coli, and three Enterobacter cloacae strains). Seven patients were simultaneously colonized with two CTX-R Enterobacteriaceae isolates. CTX-M-15 was detected in most of the CTX-R Enterobacteriaceae isolates (30/35, 88.23%). Three strains co-produced CMY-4 and 22 strains were carbapenem-resistant and co-produced a carbapenemase [OXA-48 (n = 13) or NDM-1 (n = 6)]. Molecular typing of K. pneumoniae strains, revealed eight Pulsed field gel electrophoresis (PFGE) patterns and four sequence types (ST) [ST101, ST147, ST429, and ST336]. However, E. coli isolates were genetically unrelated and belonged to A (n = 2), B1 (n = 2) and B2 (n = 3) phylogenetic groups and to ST131 (two strains), ST572 (two strains), ST615 (one strain) and ST617 (one strain). Five colonized patients were infected by CTX-R Enterobacteriaceae (four with the same strain identified from their rectal swab and one with a different strain). Whether imported or acquired during the stay in the ICU, colonization by CTX-R Enterobacteriaceae is a major risk factor for the occurrence of serious nosocomial infections. Their systematic screening in fecal carriage is mandatory to prevent the spread of these multidrug resistant bacteria.

This study describes an outbreak of NDM-1-producing K. pneumoniae strains, isolated from a Tunisian hospital, caused by two clones belonging to ST147 and ST307; and highlights the role of OMPs loss, in combination with β-lactamase expression, in conferring high carbapenem resistance.

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Avian ESBL-producing Escherichia coli isolates have been increasingly reported worldwide. Animal to human dissemination, via food chain or direct contact, of these resistant bacteria has been reported. In Tunisia, little is known about avian ESBL- producing E. coli and further studies are needed. Seventeen ESBL-producing Escherichia coli isolates from poultry feces from two farms (Farm 1 and farm 2) in the North of Tunisia have been used in this study. Eleven of these isolates (from farm 1) have the same resistance profile to nalidixic acid, sulfonamides, streptomycin, tetracycline, and norfloxacine (intermediately resistant). Out of the six isolates recovered from farm 2, only one was co-resistant to tetracycline. All isolates, except one, harbored blaCTX-M-1 gene, and one strain co-harbored the blaTEM-1 gene. The genes tetA and tetB were carried, respectively, by 11 and 1 amongst the 12 tetracycline-resistant isolates. Sulfonamides resistance was encoded by sul1, sul2, and sul3 genes in 3, 17, and 5 isolates, respectively. The qnrB1 was detected in nine strains, one of which co-harbored qnrS1 gene. The search for the class 1 and 2 integrons by PCR showed that in farm 1, class 1 and 2 integrons were found in one and ten isolates, respectively. In farm 2, class 1 integron was found in only one isolate, class 2 was not detected. Only one gene cassette arrangement was demonstrated in the variable regions (VR) of the 10 int2-positive isolates: dfrA1- sat2-aadA1. The size of the VR of the class 1 integron was approximately 250 bp in one int1-positive isolate, whereas in the second isolate, no amplification was observed. All isolates of farm 1 belong to the phylogroup A (sub-group A0). However, different types of phylogroups in farm 2 were detected. Each of the phylogroups A1, B22, B23 was detected in one strain, while the D2 phylogroup was found in 3 isolates. The virulence genes iutA, fimH, and traT were detected in 3, 7, and 3 isolates, respectively. Two types of gene combination were detected: iutA+fimH+traT in 3 isolates and iutA+fimH in one isolate. The isolates recovered in farm 1 showed the same profile of PFGE macro-restriction, while isolates of farm 2 presented unrelated PFGE patterns. We conclude that these avian ESBL-producing E. coli isolates show homo- and heterogenic genetic background and that plasmids harboring ESBL genes could be involved in the dissemination of this resistance phenotype.

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