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The aim of the present study was to examine the prevalence and determine the molecular characteristics of extended-spectrum β-lactamase-producing Escherichia coli (ESBL-EC) causing bacteraemia in a Spanish Hospital over a 12-year period (2000 to 2011). As far as we know, this is the first study which has investigated and compared the serotypes, phylogroups, clonotypes, virotypes, and PFGE profiles of ST131 and non-ST131 clones of bacteraemia ESBL-EC isolates. Of the 2,427 E. coli bloodstream isolates, 96 (4.0%) were positive for ESBL production: 40 for CTX-M-15, 36 for CTX-M-14, eight for CTX-M-1, four for CTX-M-9, CTX-M-32, and SHV-12. The number of ESBL-EC increased from 1.0% during 2000 to 2005 to 5.5% during 2006–2011 (P < 0.001). The 96 ESBL-EC isolates belonged to 36 different STs. The commonest was ST131 (41 isolates), followed by ST58, ST354, ST393 and ST405 (four isolates each). Most CTX-M-15 isolates (87.5%, 35/40) were ST131, whereas the 36 CTX-M-14 isolates belonged to 23 different STs and only 3 (8.3%) of them were ST131. The 35 ST131 CTX-M-15-producing isolates belonged to the H 30Rx subclone and 29 of them showed the virotype A. A drastic change in ST131 virotypes happened in 2011 due to the emergence of the virotypes E (sat , papGII , cnf1 , hlyA , and kpsMII-K5) and F (sat , papGII , and kpsMII-K5) which displaced virotype A (afa/draBC , afa operon FM955459, sat , and kpsMII-K2). Although the 96 ESBL-EC isolates showed 21 O serogroups and 17 H flagellar antigens, 39 belonged to serotype O25b:H4 (ST131 isolates). The second most prevalent serotype (O15:H1) was found to be associated with another important high-risk clone (ST393). In conclusion, the ST131 was the most frequent sequence type, being the H 30Rx subclone responsible for the significant increase of ESBL-EC isolates since 2006. Here, we report two new virotypes (E and F) of the H 30Rx subclone emerged in 2011. Future molecular studies are needed to understand the dynamics of expansion of this successful high-risk subclone in order to prevent its spread and establish the importance of the two new virotypes. [ABSTRACT FROM AUTHOR]

The present study was carried out to evaluate the prevalence of the clonal subgroup O16:H5-ST131 and the H 30 and H 30-Rx subclones among E. coli isolates causing extraintestinal infections and to know their virulence potential. The ST131 clonal group accounted for 490 (16%) of the 2995 isolates obtained from clinical samples in five Spanish hospitals during the study period (2005–2012). Among those 490 ST131 isolates, 456 belonged to serotype O25b:H4, 27 to O16:H5 and seven were O-non-typeable:H4 (ONT:H4). All 27 O16:H5 isolates showed fimH 41, whereas fimH 30 and fimH 22 alleles were the most frequently detected among O25b:H4 isolates. The majority (381/490; 78%) of ST131 isolates belonged to H 30 subclone, and 302 of 381 (79%) H 30 isolates belonged to the H 30-Rx subclone. Of the 27 O16:H5 isolates, 48% produced CTX-M-14; however, none produced CTX-M-15. In contrast, 46% of O25b:H4 isolates produced CTX-M-15 while only 2% produced CTX-M-14. More than a half of the O16:H5 isolates (56%) showed the ExPEC status which was significantly more prevalent within O25b:H4 isolates (81%) ( P < 0.01), especially among H 30-Rx (97%) isolates. In the present study, a modified virotype scheme was applied within which approximately half (52%) of the O16:H5 isolates showed the C1 specific virotype. Despite their low virulence-gene score (mean of virulence genes 6.4 versus 8.5 in O25b:H4 isolates), six out of the 10 O16:H5 isolates assayed showed high virulence in the mouse model of sepsis (killed 90–100% of mice challenged). Furthermore, four O16:H5 isolates of virotypes A and C1, carrying K2 variant of group II capsule, showed lethality at 24 h. Thus, certain O16:H5 fimH 41 isolates show a similar in vivo virulence to that reported with the highly virulent O25b:H4 H 30-Rx isolates (Mora et al., PLOS ONE 2014, e87025), supporting their potential virulence for humans. [ABSTRACT FROM AUTHOR]

Objectives Determining the prevalence of children in day-care centres (DCCs) carrying faecal extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae and molecularly characterizing those belonging to the Escherichia coli species. Methods Stools were collected from children's diapers (January–April 2012) in randomly chosen DCCs and plated onto ChromID™ ESBL. Colonies growing on this medium were identified by the Vitek 2® system and tested for antibiotic susceptibility and for ESBL production by the double-disc synergy test. ESBL genotypes were determined as well as phylogenetic groups, ERIC-2 (enterobacterial repetitive intergenic consensus) PCR profiles and sequence types (STs) for the E. coli isolates. Serotypes, virotypes, fimH alleles, ESBL-carrying plasmids and PFGE patterns were determined for the ST131 E. coli isolates. Results Among 419 children from 25 participating DCCs, 1 was colonized by CTX-M-15-producing Klebsiella pneumoniae and 27 (6.4%) by E. coli, which all produced CTX-M enzymes [CTX-M-15 (37%), CTX-M-1 (26%), CTX-M-14 (22%), CTX-M-27 (11%) and CTX-M-22 (4%)]. The 27 E. coli isolates, 55.5% belonging to group B2, displayed 20 ERIC-2 PCR profiles and 16 STs. The ST131 E. coli isolates were dominant (44%), displayed serotypes O25b:H4 and O16:H5, fimH alleles 30 and 41 and virotypes A and C. According to the PFGE patterns, one strain of E. coli ST131 producing a CTX-M-15 enzyme carried by an IncF F2:A1:B− plasmid had spread within one DCC. Conclusions This study shows a notable prevalence (6.4%) of DCC children with faecal CTX-M-producing E. coli isolates comprising a high proportion of E. coli ST131 isolates, suggesting that these children might be a reservoir of this clone. [ABSTRACT FROM PUBLISHER]

Escherichia coli sequence type (ST)131 is an emerging disseminated public health threat implicated in multidrug-resistant extraintestinal infections worldwide. Although the majority of ST131 isolates belong to O25b:H4 serotype, new variants with different serotypes, STs using the discriminative multilocus sequence typing scheme of Pasteur Institute, and virulence-gene profiles (virotypes) have been reported with unknown implications on the pattern of spread, persistence and virulence. The aim of the present study was to compare virulence in a mouse subcutaneous sepsis model of representative ST131 clinical isolates belonging to 2 serotypes (O25b:H4, O16:H5) and nine virotypes and subtypes (A, B, C, D1, D2, D3, D4, D5 and E). Fourteen out of the 23 ST131 isolates tested (61%) killed 90 to 100% of mice challenged, and 18 of 23 (78%) at least 50%. Interestingly, different virulence patterns in association with virotypes were observed, from highly rapid lethality (death in less than 24 h) to low final lethality (death at 7 days) but with presence of an acute inflammation. This is the first study to assess virulence of ST131 isolates belonging to serotype O16:H5, which exhibited virotype C. In spite of their low virulence-gene score, O16:H5 isolates did not show significant differences in final lethality compared with highly virulent O25b:H4 isolates of virotypes A, B and C, but killed mice less rapidly. Significant differences were found, however, between virotypes A, B, C (final lethality ≥80% of mice challenged) and virotypes D, E. Particularly unexpected was the low lethality of the newly assigned virotype E taking into account that it exhibited high virulence-gene score, and the same clonotype H30 as highly virulent O25b:H4 isolates of virotypes A, B and C. In vivo virulence diversity reported in this study would reflect the genetic variability within ST131 clonal group evidenced by molecular typing. [ABSTRACT FROM AUTHOR]

The objectives of this study were to investigate the presence of Shiga toxin-producing Escherichia coli (STEC) strains in wildlife that have spread in Europe, living near human settlements; to analyze their epidemiological role in maintenance and transmission to domestic livestock; and to assess the potential health risk of wildlife-carried strains. STEC strains were recovered from 53% of roe deer, 8.4% of wild boars, and 1.9% of foxes sampled in the northwest of Spain (Galicia). Of the 40 serotypes identified, 21 were classified as seropathotypes associated with human disease, accounting for 81.5% of the wildlife-carried STEC strains, including the enterohemorrhagic serotypes O157:H7-D-eae-β1, O26:[H11]-B1-eae-β1, O121:H19-B1-eae-ϵ1, and O145:[H28]-D-eae-γ1. None of the wildlife-carried strains belonged to the highly pathogenic serotype O104:H4-B1 from the recent Germany outbreak. Forty percent of wildlife-carried STEC strains shared serotypes, phylogroups, intimin types, and Stx profiles with isolates from human patients from the same geographic area. Furthermore, wildlife-carried strains belonging to serotypes O5:HNM-A, O26:[H11]-B1, O76:H19-B1, O145:[H28]-D, O146:H21-B1, and O157:H7-D showed pulsed-field gel electrophoresis (PFGE) profiles with >85% similarity to human-pathogenic STEC strains. We also found a high level of similarity among STEC strains of serotypes O5:HNM-A, O26:[H11]-B1, and O145:HNM-D of bovine (feces and beef) and wildlife origins. Interestingly, O146:H21-B1, the second most frequently detected serotype in this study, is commonly associated with human diarrhea and isolated from beef and vegetables sold in Galicia. Importantly, at least 3 STEC isolates from foxes (O5:HNM-A-eae-β1, O98:[H21]-B1-eae-ζ1, and O146:[H21]-B1) showed characteristics similar to those of human STEC strains. In conclusion, roe deer, wild boar, and fox in Galicia are confirmed to be carriers of STEC strains potentially pathogenic for humans and seem to play an important role in the maintenance of STEC. [ABSTRACT FROM AUTHOR]

Abstract: CTX-M enzymes, mainly CTX-M-14 and CTX-M-15, have emerged as the most prevalent extended-spectrum β-lactamase (ESBL) type produced by Escherichia coli in Spain, with successful dissemination of clonal group O25b:H4-B2-ST131 producing CTX-M-15 within the hospital and community settings. However, until now CTX-M-14-producing E. coli in Spain had been shown to belong to a wide variety of serotypes with no predominance of a certain clonal group. In the present study, 654 E. coli strains positive for ESBL production obtained between 2005 and 2008 from inpatients and outpatients of four hospitals in Galicia, northwest Spain, were analysed. The strains were characterised with regard to ESBL enzymes, serotype, virulence genes, phylogenetic group, multilocus sequence type, and pulsed-field gel electrophoresis of XbaI-digested DNA. As a result, the emergence of certain clonal groups of extraintestinal pathogenic E. coli producing CTX-M-14 has been detected in this geographic area, including O1:HNM-D-ST59, O15:H1-D-ST393/ST1394, O20:H34/HNM-D-ST354, O25b:H4-B2-ST131 and ONT:H21,42-B1-ST101. These five clonal groups showed a high virulence potential as they harboured more than eight virulence factors, which could explain their successful dissemination. [ABSTRACT FROM AUTHOR]

Objectives Having shown that the Xeral-Calde Hospital in Lugo (Spain) has been concerned by Escherichia coli clone O25:H4-ST131 producing CTX-M-15 (Nicolas-Chanoine et al. J Antimicrob Chemother 2008; 61: 273–81), the present study was carried out to evaluate the prevalence of this clone among the extended-spectrum β-lactamase (ESBL)-producing E. coli isolates and also to molecularly characterize the E. coli isolates producing ESBL other than CTX-M-15. Methods In the first part of this study, 105 ESBL-producing E. coli isolates (February 2006 to March 2007) were characterized with regard to ESBL enzymes, serotypes, virulence genes, phylogenetic groups, multilocus sequence typing (MLST) and PFGE. In the second part of this study, 249 ESBL-producing E. coli isolates (April 2007 to May 2008) were investigated only for the detection of clone O25b:H4-ST131 producing CTX-M-15 using a triplex PCR developed in this study and based on the detection of the new operon afa FM955459 and the targets rfbO25b and 3′ end of the blaCTX-M-15 gene. Results Of the 105 ESBL-producing E. coli isolates, 60 (57.1%) were positive for CTX-M-14, 23 (21.9%) for CTX-M-15, 10 (9.5%) for SHV-12 and 7 (6.7%) for CTX-M-32. Serotypes, virulence genes, phylogenetic groups and molecular typing by PFGE demonstrated high homogeneity within those producing CTX-M-15 and high diversity within E. coli producing CTX-M-14 and other ESBLs. By PFGE, CTX-M-15-producing E. coli isolates O25b:H4 belonging to the phylogenetic group B2 and MLST profile ST131 were grouped in the same cluster. The epidemic strain of clone O25b:H4-ST131 represented 23.1%, 22.5% and 20.0% of all ESBL-producing E. coli isolated in 2006, 2007 and 2008, respectively. Conclusions CTX-M-type ESBLs, primarily CTX-M-14 and CTX-M-15, have emerged as the predominant types of ESBL produced by E. coli isolates in Lugo. In view of the reported findings, long-term care facilities for elderly people may represent a significant reservoir for E. coli clone O25b:H4-ST131 producing CTX-M-15. The triplex PCR developed in this work will be useful for rapid and simple detection of this clone. [ABSTRACT FROM AUTHOR]

Food-producing animals can harbor Escherichia coli strains with potential to cause diseases in humans. In this study, the presence of enteropathogenic E. coli (EPEC) was investigated in fecal samples from 130 healthy sheep (92 lambs and 38 adults) raised for meat in southern Brazil. EPEC was detected in 19.2% of the sheep examined, but only lambs were found to be positive. A total of 25 isolates was characterized and designated atypical EPEC (aEPEC) as tested negative for bfpA gene and BFP production. The presence of virulence markers linked to human disease as ehxA , paa , and lpfA O113 was observed in 60%, 24%, and 88% of the isolates, respectively. Of the 11 serotypes identified, eight were described among human pathogenic strains, while three (O1:H8, O11:H21 and O125:H19) were not previously detected in aEPEC. Associations between intimin subtypes and phylogroups were observed, including eae -θ2/A, eae -β1/B1, eae -α2/B2 and eae -γ1/D. Although PFGE typing of 16 aEPEC isolates resulted in 14 unique pulsetypes suggesting a genetic diversity, specific clones were found to be distributed in some flocks. In conclusion, potentially pathogenic aEPEC strains are present in sheep raised for meat, particularly in lambs, which can better contribute to dissemination of these bacteria than adult animals. [ABSTRACT FROM AUTHOR]