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Objectives: Antimicrobial resistance genes (ARGs) can be transferred by means of mobile genetic elements, which play a critical role in the dissemination of resistance in the bacterial community. ARG transmission within mobile genetic elements has been reported in plasmids and transposons but less frequently in bacteriophages. Here, the bacteriophage fraction of seven human faecal samples was purified and deep-sequenced to detect the presence of ARGs in the phage particles.Methods: Seven faecal samples (five from healthy individuals and two from a patient before and after receiving ciprofloxacin treatment) were used to extract phage DNA, which was purified and then sequenced in a MiSeq (Illumina). Generated reads were checked for quality and assembled, and then the generated contigs analysed with Kraken, PHASTER, VirSorter and Prokka. Some genes were also validated by quantitative PCR.Results: Analysis of the purified phage DNA by Kraken identified from 4 to 266 viruses in the samples. The viral fraction corresponded mainly to the order Caudovirales, including phages from the Siphoviridae and Myoviridae families. Bacterial genes associated with antimicrobial resistance were detected in the viral DNA, as confirmed by quantitative PCR. Higher densities of ARG-carrying phage particles were observed in the post- versus pre-ciprofloxacin treatment sample.Conclusions: The finding of ARGs in phage particles supports the description of phages as mobile elements contributing to the dissemination of bacterial antibiotic resistance and suggests ciprofloxacin treatment may play a role in the release of ARG-carrying particles, thereby increasing resistance. [ABSTRACT FROM AUTHOR]

Objectives To describe the prevalence and risk factors for infection due to AmpC β-lactamase-producing Escherichia coli (AmpC-EC). Methods For the prevalence study, all clinical isolates of E. coli with reduced susceptibility to third-generation cephalosporins were prospectively included from June 2010 to November 2011. For risk factor analysis, a case–control study was conducted. Cases were patients with an infection due to AmpC-EC. Controls were patients infected with cephalosporin-susceptible E. coli, matched 1 : 2. Detection of blaAmpC genes was done with a multiplex AmpC–PCR, and hyperproduction of E. coli chromosomal blaAmpC by quantitative RT–PCR. Alteration of the blaAmpC promoter was studied by PCR and sequencing. Results We identified 243 (1.1%) AmpC-EC strains out of 21 563 clinical isolates. Three cases with strains carrying ESBLs, 18 strains that were considered due to colonization and 8 cases lost to clinical follow-up were excluded. Finally, 214 cases were included in the analysis. Ninety-one cases (42.5%) and 269 (62.8%) controls were strictly community acquired (P < 0.001). Thirty-five (16.3%) cases and 186 controls (43.5%) did not have any identifiable risk factor (P < 0.001). Among cases, 158 (73.8%) were found to harbour an acquired AmpC (73.4% CMY-2). Previous use of fluoroquinolones [OR 2.6 (95% CI 1.12–3.36); P = 0.008] was independently associated with AmpC-EC in the multivariate analysis. Conclusions Prevalence of AmpC in E. coli remains low in our area. Plasmid acquisition (CMY type) represents the main mechanism of AmpC production. A high proportion of community-acquired isolates and patients with no identifiable risk factors were found. Previous use of fluoroquinolones was identified as a risk factor. [ABSTRACT FROM PUBLISHER]

Objectives To gain insights into ampC transmission between bacterial strains. Methods We examined the genetic context of 117 acquired ampC genes from 27119 Enterobacteriaceae collected between 1999 and 2007. Plasmid analysis was carried out by PCR-based replicon or relaxase typing, S1-PFGE and Southern hybridization. I-CeuI/PFGE was used for isolates not characterized by plasmid analysis. PCR reactions were used to map the genetic organization of the ampC genes. Results Among the isolates studied, 81.2% of ampC genes were located on plasmids of known Inc/MOB groups, 7.7% were chromosomally located and 11.1% were not determined. A/C, I1 and K were the most commonly found replicons in plasmids carrying blaCMY-2, while L/M replicons were associated with blaDHA-1. blaACC-1 was linked to I1 and MOBF11 plasmids; blaCMY-27 was associated with IncF and MOBP12 plasmids; the plasmid carrying blaCMY-25 could not be typed, and blaCMY-40 was chromosomally located. All 87 isolates carrying blaCMY-2, blaCMY-4, blaCMY-25, blaCMY-27, blaCMY-40 or blaACC-1 displayed the transposon-like structures ISEcp1/ΔISEcp1-blaCMY-blc-sugE or ΔISEcp1-blaACC-1-gdha. The most prevalent structure in blaDHA-1 (93.3% of cases) was identical to that described in the Klebsiella pneumoniae pTN60013 plasmid. Remarkably, in three isolates containing chromosomal blaCMY-2, this gene was mobilized by conjugation. Conclusions Although plasmids are the main cause of the rapid dissemination of ampC genes among bacteria, we need to be aware that other mobile genetic elements such as integrative and conjugative elements (ICEs) can be involved in the mobilization of these genes. [ABSTRACT FROM PUBLISHER]

Objectives To characterize the vectors involved in the dissemination of blaCMY-2 genes in clinical isolates of Proteus mirabilis collected between 1999 and 2007. Methods Plasmid analysis of 19 P. mirabilis carrying ampC genes was performed by PCR-based replicon typing, S1-PFGE and Southern hybridization with ampC and replicon probes. Isolates that could not be characterized were examined for the presence of SXT/R391-like elements. To demonstrate the involvement of these elements in the dissemination of blaCMY-2, we performed a PCR amplification of the integrase (int) and toxin/antitoxin (TA) genes from SXT/R391-like integrative conjugative elements (ICEs). Later on, I-Ceu-I PFGE gels and hybridization with blaCMY-2, int and prfC probes were performed. The genetic organization of blaCMY-2 was also studied. Results ampC genes were located on large conjugative plasmids in 11 of the 19 (58%) P. mirabilis studied. However, in eight of these isolates a plasmid was not involved in the mobilization of ampC genes. I-Ceu-I PFGE and hybridization analyses revealed that blaCMY-2 were chromosomally located in these eight P. mirabilis isolates. The genetic organization of blaCMY-2 and hybridization analyses revealed that blaCMY-2 was carried by an ICE almost identical to ICEPmiJpan1 in seven out of these eight isolates. Conclusions The prevalence of ICEs carrying blaCMY-2 was surprisingly high [37% (7 out of 19)]. This is the first study giving prevalence data on ICEs carrying blaCMY-2 genes. These results suggest the need to study these mobile genetic elements in the context of dissemination of acquired AmpC β-lactamases and also of other β-lactamases, such as extended-spectrum β-lactamases and carbapenemases. [ABSTRACT FROM PUBLISHER]

Objectives: We describe 12 VIM-1-producing strains (7 Enterobacter cloacae, 2 Klebsiella pneumoniae and 3 clonal Klebsiella oxytoca strains) detected among clinically relevant Enterobacteriaceae isolates from routine cultures at the Hospital del Mar (Barcelona, Spain) from December 2006 to May 2007. Methods: Susceptibility to carbapenems was evaluated with the MicroScan system. b-Lactamases were identified by PCR and sequencing. Clonal relationships between the isolates were analysed by PFGE. Transferability of the enzymes was tested by conjugation. Plasmid characterization was performed by PCR-based replicon typing and PFGE with S1 nuclease digestion of whole genomic DNA. The PFGE gels were then transferred and hybridized. Results: The disc diffusion method correctly identified five of the seven E. cloacae isolates as intermediate or resistant strains. All isolates produced the VIM-1 enzyme. Three E. cloacae and three K. oxytoca strains were also CTX-M-9-producing strains, and one E. cloacae was also a CTX-M-3-producing strain. The plasmids carrying the blaVIM gene, of unknown incompatibility group, had a size of ∼75 kb (eight strains) or 40 kb (three strains) and also contained the qnrS and the aac(6′)-Ib-cr genes. In the remaining strain the blaVIM-1 gene was found in an HI2 plasmid of 290 kb together with blaCTX-M-9, qnrA, qnrS and the aac(6′)-Ib-cr genes. Conclusions: The results showed a linkage between the blaVIM-1 and the qnrS and the aac(6′)-Ib-cr genes, and between the blaCTX-M-9 and the qnrA genes. [ABSTRACT FROM AUTHOR]

The article discusses a study on the prevalence of carbapenemsase-producing Enterobacteriaceae (CPE) carriers admitted to intensive care units (ICUs) in 11 acute-care Spanish hospitals from 7 different geographical areas. It advises for the implementation of guidelines for the management of the infection control measures to reduce MDR Gram-negative bacteria recommended by ESCMID.

The aim of this study was to evaluate the incidence of decreased susceptibility to broad-spectrum cephalosporins in Enterobacteriaceae that lack inducible chromosomal bla genes, and to determine the enzymes responsible for resistance. From all clinically relevant Enterobacteriaceae strains isolated between 1994 and 1996, 88 of 7054 Escherichia coli, seven of 581 Klebsiella pneumoniae and 23 of 166 Klebsiella oxytoca strains were studied because of their decreased susceptibilities to broad-spectrum cephalosporins (as reflected in intermediate susceptibilities and/or positive synergy tests and/or irregular crenellated inhibition zones). The most frequent mechanism implicated in decreased susceptibility to broad-spectrum cephalosporins displayed by E. coli and K. oxytoca was hyperproduction of chromosomal beta-lactamase, followed by plasmid-mediated SHV-1 hyperproduction in E. coli. In our hospital, the incidence of plasmid-mediated extended-spectrum beta-lactamases (ESBLs) between 1994 and 1996 was low. ESBLs were found in only 10 (0.14%) E. coli strains (six CTX-M-9, two TEM-12 and two SHV-2), in one (0.17%) K. pneumoniae strain (SHV-2) and in no K. oxytoca strains. The relatively wide variety of beta-lactamases that were detected among these common bacteria isolated from a single medical centre, including non-TEM- and non-SHV-derived ESBLs, appears epidemiologically remarkable. [ABSTRACT FROM AUTHOR]

CMY-2 plasmid-mediated AmpC beta-lactamase (CMY-2) was detected in 21 isolates from two hospitals located in different geographical regions of Spain between October 1999 and December 2000. The isolates comprised two Salmonella enterica serovars (Mikawasima and Montevideo), 16 Escherichia coli, one Klebsiella pneumoniae, one Klebsiella oxytoca and one Proteus mirabilis. In addition to the expected resistance to beta-lactams, including extended-spectrum cephalosporins and cefoxitin, all isolates showed a broad spectrum of associated resistance. All were resistant to sulfamethoxazole, chloramphenicol, tetracycline and streptomycin, and all but two were also resistant to gentamicin. Five isolates were studied in detail and all transferred CMY-2 and other resistance determinants by conjugation. Genomic DNA restriction pattern analysis of the E. coli isolates excluded the dissemination of a single clone. To the best of our knowledge this is the first time that CMY-2 has been detected in P. mirabilis, K. oxytoca and S. enterica serovars Mikawasima and Montevideo. It is also the first time that CMY-2 has been described in Spain. [ABSTRACT FROM AUTHOR]

The article presents a study that deals with genetic clones of CTX-M-15-producing Klebsiella pneumoniae in the Barcelona metropolitan area in Spain. The authors assessed the clonality and persistence of CTX-M-15-producing K. pneumoniae isolates. A retrospective study of extended-spectrum ß-lactamases-producing K. pneumoniae clinical samples was conducted. They were collected from samples stored from 2005 through 2008 at Hospital Sant Pau and Hospital Val d'Hebron. Also included in the study were all the ESBL-producing K. pneumoniae isolates encountered at Hospital General de l'Hospitalet during the outbreak period in July 2008. Based on their findings, the authors highlighted the dissemination and persistence of three K. pneumoniae clonal strains producing CTX-M-15.

A correction to the article "Detection of Three Stable Genetic Clones of CTX-M-15-Producing Klebsiella pneumoniae in the Barcelona Metropolitan Area, Spain" that was published in a 2009 issue of the journal is presented.