Comparison of CTX-M encoding plasmids present during the early phase of the ESBL pandemic in western Sweden.

Plasmids encoding blaCTX-M genes have greatly shaped the evolution of E. coli producing extended-spectrum beta-lactamases (ESBL-E. coli) and adds to the global threat of multiresistant bacteria by promoting horizontal gene transfer (HGT). Here we screened the similarity of 47 blaCTX-M -encoding plasmids, from 45 epidemiologically unrelated and disperse ESBL-E. coli strains, isolated during the early phase (2009-2014) of the ESBL pandemic in western Sweden. Using optical DNA mapping (ODM), both similar and rare plasmids were identified. As many as 57% of the plasmids formed five ODM-plasmid groups of at least three similar plasmids per group. The most prevalent type (28%, IncIl, pMLST37) encoded blaCTX-M-15 (n = 10), blaCTX-M-3 (n = 2) or blaCTX-M-55 (n = 1). It was found in isolates of various sequence types (STs), including ST131. This could indicate ongoing local HGT as whole-genome sequencing only revealed similarities with a rarely reported, IncIl plasmid. The second most prevalent type (IncFII/FIA/FIB, F1:A2:B20) harboring blaCTX-M-27, was detected in ST131-C1-M27 isolates, and was similar to plasmids previously reported for this subclade. The results also highlight the need for local surveillance of plasmids and the importance of temporospatial epidemiological links so that detection of a prevalent plasmid is not overestimated as a potential plasmid transmission event in outbreak investigations.

Comparative Genomic Analysis of ST131 Subclade C2 of ESBL-Producing E. coli Isolates from Patients with Recurrent and Sporadic Urinary Tract Infections.

The global emergence of extended-spectrum beta-lactamase-producing Escherichia coli (ESBL-E. coli), mainly causing urinary tract infections (UTI), is a major threat to human health. ESBL-E. coli sequence type (ST) 131 is the dominating clone worldwide, especially its subclade C2. Patients developing recurrent UTI (RUTI) due to ST131 subclade C2 appear to have an increased risk of recurrent infections. We have thus compared the whole genome of ST131 subclade C2 isolates from 14 patients with RUTI to those from 14 patients with sporadic UTI (SUTI). We aimed to elucidate if isolates causing RUTI can be associated with specific genomic features. Paired isolates from patients with RUTI were identical, presenting 2-18 single nucleotide polymorphism (SNP) differences for all six patients investigated. Comparative genomic analyses, including virulence factors, antibiotic resistance, pangenome and SNP analyses did not find any pattern associated with isolates causing RUTI. Despite extensive whole genome analyses, an increased risk of recurrences seen in patients with UTI due to ST131 subclade C2 isolates could not be explained by bacterial genetic differences in the two groups of isolates. Hence, additional factors that could aid in identifying bacterial properties contributing to the increased risk of RUTI due to ESBL-E. coli ST131 subclade C2 remains to be explored.

Knockout of Targeted Plasmid-Borne ß-Lactamase Genes in an Extended-Spectrum-ß-Lactamase-Producing Escherichia coli Strain: Impact on Resistance and Proteomic Profile.

Resistance to ß-lactams is known to be multifactorial, although the underlying mechanisms are not well established. The aim of our study was to develop a system for assessing the phenotypic and proteomic responses of bacteria to antibiotic stress as a result of the loss of selected antimicrobial resistance genes. We applied homologous recombination to knock out plasmid-borne ß-lactamase genes (blaOXA-1, blaTEM-1, and blaCTX-M15) in Escherichia coli CCUG 73778, generating knockout clone variants lacking the respective deleted ß-lactamases. Quantitative proteomic analyses were performed on the knockout variants and the wild-type strain, using bottom-up liquid chromatography tandem mass spectrometry (LC-MS/MS), after exposure to different concentrations of cefadroxil. Loss of the blaCTX-M-15 gene had the greatest impact on the resulting protein expression dynamics, while losses of blaOXA-1 and blaTEM-1 affected fewer proteins' expression levels. Proteins involved in antibiotic resistance, cell membrane integrity, stress, and gene expression and unknown function proteins exhibited differential expression. The present study provides a framework for studying protein expression in response to antibiotic exposure and identifying the genomic, proteomic, and phenotypic impacts of resistance gene loss. IMPORTANCE The critical situation regarding antibiotic resistance requires a more in-depth effort for understanding underlying mechanisms involved in antibiotic resistance, beyond just detecting resistance genes. The methodology presented in this work provides a framework for knocking out selected resistance factors, to study the adjustments of the bacterium in response to a particular antibiotic stress, elucidating the genetic response and proteins that are mobilized. The protocol uses MS-based determination of the proteins that are expressed in response to an antibiotic, enabling the selection of strong candidates representing putative resistance factors or mechanisms and providing a basis for future studies to understand their implications in antibiotic resistance. This allows us to better understand how the cell responds to the presence of the antibiotic when a specific gene is lost and, consequently, identify alternative targets for possible future treatment development.

The impact of the ST131 clone on recurrent ESBL-producing E. coli urinary tract infection: a prospective comparative study.

The global emergence of extended-spectrum beta-lactamase-producing Escherichia coli (ESBL-E. coli), mainly causing urinary tract infections (UTI), is of great concern. Almost one third of patients with UTI, develop recurrent UTI (RUTI). We followed 297 patients for one year after their first episode of UTI due to ESBL-E. coli. Our aim was to evaluate the impact of the globally dominant sequence type (ST)131 clone and its clades, on the risk of subsequent recurrences with ESBL-E. coli. Isolates from patients developing RUTI (68/297) were compared with those from patients with sporadic UTI (SUTI, 229/297). No association was found between RUTI and the two most prevalent phylogroups B2 and D, blaCTX-M genes, or resistance profile. Half of the patients with RUTI were infected with ST131 isolates. Clade C2 were in dominance (50/119) among ST131 isolates. They were more common in patients with RUTI than SUTI (28% vs 13%) and multivariate analysis showed an increased odds-ratio (OR = 2.21, p = 0.033) for recurrences in patients infected with these isolates as compared to non-ST131 isolates. Detecting specific biomarkers, as ST131 clade C2, in ESBL-E. coli UTI isolates may aid in prediction of RUTI and improve diagnostics and care of patients with a risk of ESBL-E. coli recurrences.

Prevalence and patient related factors associated with Extended-Spectrum Beta-Lactamase producing Escherichia coli and Klebsiella pneumoniae carriage and infection among pediatric patients in Tanzania.

Extended-Spectrum Beta-Lactamase (ESBL) producing Enterobacteriaceae (EPE) is increasing worldwide, though less documented in low-income settings. Here we determined the prevalence of EPE infection and carriage, and patient factors associated with EPE-carriage among pediatric patients in three health care levels in Tanzania. Between January and April 2016, 350 febrile children (median age 21 months) seeking care at a university or a regional referral hospital, or a health centre in Moshi municipality, Tanzania, were included. Socio-demographic characteristics were collected using a questionnaire. Rectal swabs and blood cultures were collected from all children (n = 350) and urinary samples from 259 children at admission. ESBL-phenotype and antimicrobial susceptibility were determined for Klebsiella pneumoniae (K. pneumoniae) and Escherichia coli (E. coli) isolates. Only one EPE case (E. coli) in blood and four in urine (one E. coli and three K. pneumoniae) were found, whereas (n = 90, 26%) of the children were colonized in feces (ESBL-E. coli; n = 76, ESBL-K. pneumoniae, n = 14). High resistance rates were seen in fecal ESBL-E. coli (n = 76) against trimethoprim-sulfamethoxazole (n = 69, 91%), gentamicin (n = 51, 67%), ciprofloxacin (n = 39, 51%) and chloramphenicol (n = 27, 35%) whereas most isolates were sensitive to amikacin (n = 71, 93%). Similar rates were seen for fecal ESBL-K. pneumoniae. Resistance to first line antibiotics were also very high in fecal E. coli not producing ESBL. No sociodemographic factor was associated with EPE-carriage. Children colonized with EPE were younger than 12 months (n = 43, 48%) and often treated with antibiotics (n = 40, 44%) in the previous two months. After adjustment for age children admitted to the intensive care unit had higher odds of EPE fecal carriage compared with those in the general wards (OR = 3.9, 95%CI = 1.4-10.4). Despite comparatively high rates of fecal EPE-carriage and previous antibiotic treatment, clinical EPE cases were rare in the febrile children. The very high resistant rates for the EPE and the non-ESBL producing E. coli to commonly used antibiotics are worrying and demand implementation of antibiotic stewardship programs in all levels of health care in Tanzania.

Identity of blaCTX-M Carrying Plasmids in Sequential ESBL-E. coli Isolates from Patients with Recurrent Urinary Tract Infections.

Plasmid-mediated multidrug resistance in E. coli is becoming increasingly prevalent. Considering this global threat to human health, it is important to understand how plasmid-mediated resistance spreads. From a cohort of 123 patients with recurrent urinary tract infections (RUTI) due to extended spectrum beta-lactamase (ESBL)-producing Escherichia coli (ESBL E. coli), only five events with a change of ESBL E. coli strain between RUTI episodes were identified. Their blaCTX-M encoding plasmids were compared within each pair of isolates using optical DNA mapping (ODM) and PCR-based replicon typing. Despite similar blaCTX-M genes and replicon types, ODM detected only one case with identical plasmids in the sequential ESBL E. coli strains, indicating that plasmid transfer could have occurred. For comparison, plasmids from seven patients with the same ESBL E. coli strain reoccurring in both episodes were analyzed. These plasmids (encoding blaCTX-M-3, blaCTX-M-14, and blaCTX-M-15) were unaltered for up to six months between recurrent infections. Thus, transmission of blaCTX-M plasmids appears to be a rare event during the course of RUTI. Despite the limited number (n = 23) of plasmids investigated, similar blaCTX-M-15 plasmids in unrelated isolates from different patients were detected, suggesting that some successful plasmids could be associated with specific strains, or are more easily transmitted.

Genomic and Proteomic Characterization of the Extended-Spectrum ß-Lactamase (ESBL)-Producing Escherichia coli Strain CCUG 73778: A Virulent, Nosocomial Outbreak Strain.

Escherichia coli strain CCUG 78773 is a virulent extended-spectrum ß-lactamase (ESBL)-producing ST131-O25b type strain isolated during an outbreak at a regional university hospital. The complete and closed genome sequence, comprising one chromosome (5,076,638 bp) and six plasmids (1718-161,372 bp), is presented. Characterization of the genomic features detected the presence of 59 potential antibiotic resistance factors, including three prevalent ß-lactamases. Several virulence associated elements were determined, mainly related with adherence, invasion, biofilm formation and antiphagocytosis. Twenty-eight putative type II toxin-antitoxin systems were found. The plasmids were characterized, through in silico analyses, confirming the two ß-lactamase-encoding plasmids to be conjugative, while the remaining plasmids were mobilizable. BLAST analysis of the plasmid sequences showed high similarity with plasmids in E. coli from around the world. Expression of many of the described virulence and AMR factors was confirmed by proteomic analyses, using bottom-up, liquid chromatography-tandem mass spectrometry (LC-MS/MS). The detailed characterization of E. coli strain CCUG 78773 provides a reference for the relevance of genetic elements, as well as the characterization of antibiotic resistance and the spread of bacteria harboring ESBL genes in the hospital environment.

Recurrence of urinary tract infections with extended-spectrum ß-lactamase-producing Escherichia coli caused by homologous strains among which clone ST131-O25b is dominant.

OBJECTIVES: Bacterial features associated with recurrent urinary tract infections (RUTIs) due to extended-spectrum ß-lactamase-producingEscherichia coli (ESBL-E. coli) are not well understood. In this study, phylogenetic groups and ST131 subclones were investigated to assess strain homology of ESBL-E. coli isolates in patients with RUTIs in inpatient and outpatient settings in western Sweden. METHODS: Almost all isolates (319/356) from 123 patients with 2-7 episodes (median 2 episodes) of ESBL-E. coli UTI within 1 year were examined for seven E. coli phylogroups, the ST131-O25b clone and its subclone fimH30-Rx. Antimicrobial resistance and ESBL genes were determined for the index isolates. A subset of isolates was typed using pulsed-field gel electrophoresis (PFGE). RESULTS: The same phylogroup and ST131 subclones were seen for all recurrences in 119/123 patients, and PFGE confirmed strain homology in recurrences for 43/44 patients tested. Phylogroup B2 dominated (56%), followed by D (19%) and F (10%). ST131-O25b andfimH30-Rx isolates were detected in 44% and 30%, respectively. CTX-M group 1 (71%) predominated. Elderly patients were in the majority. There were no associations between patient demographics or time to recurrence and bacterial characteristics. The fimH30-Rx subclone was associated with a higher number of recurrences (P = 0.015) compared with the remaining B2 isolates. CONCLUSION: In ESBL-E. coli RUTI, most recurrences were caused by the initial infecting strain. The high frequency of the multidrug-resistant fimH30-Rx subclone and its association with multiple recurrences warrants further attention and early detection of this subclone in patients at risk of developing RUTI with ESBL-E. coli.

Interspecies plasmid transfer appears rare in sequential infections with extended-spectrum ß-lactamase (ESBL)-producing Enterobacteriaceae.

From a cohort of 1836 Swedish patients infected with ESBL-producing Enterobacteriaceae (EPE) during 2004-2014, 513 patients with recurrent EPE infection were identified. Only in 14 of the 513 patients was a change of species (ESBL-E. coli to ESBL-K. pneumoniae or vice versa) found between the index and subsequent infection. Eleven sequential urine isolates from 5 of the 14 patients were available for further analysis of possible transfer of ESBL-carrying plasmids. The plasmid content was studied using optical DNA mapping (ODM), PCR-based replicon typing, and ESBL gene sequencing. ODM allowed us to directly compare whole plasmids between isolates and found similar ESBL-carrying plasmids in 3 out of the 5 patients. The ODM results and the rarity in shift of species between ESBL-E. coli and ESBL-K. pneumoniae imply that in recurrent EPE infections interspecies plasmid transfer is uncommon.

The resistomes of six carbapenem-resistant pathogens - a critical genotype-phenotype analysis.

Carbapenem resistance is a rapidly growing threat to our ability to treat refractory bacterial infections. To understand how carbapenem resistance is mobilized and spread between pathogens, it is important to study the genetic context of the underlying resistance mechanisms. In this study, the resistomes of six clinical carbapenem-resistant isolates of five different species - Acinetobacter baumannii, Escherichia coli, two Klebsiella pneumoniae, Proteus mirabilis and Pseudomonas aeruginosa - were characterized using whole genome sequencing. All Enterobacteriaceae isolates and the A. baumannii isolate had acquired a large number of antimicrobial resistance genes (7-18 different genes per isolate), including the following encoding carbapenemases: blaKPC-2, blaOXA-48, blaOXA-72, blaNDM-1, blaNDM-7 and blaVIM-1. In addition, a novel version of blaSHV was discovered. Four new resistance plasmids were identified and their fully assembled sequences were verified using optical DNA mapping. Most of the resistance genes were co-localized on these and other plasmids, suggesting a risk for co-selection. In contrast, five out of six carbapenemase genes were present on plasmids with no or few other resistance genes. The expected level of resistance - based on acquired resistance determinants - was concordant with measured levels in most cases. There were, however, several important discrepancies for four of the six isolates concerning multiple classes of antibiotics. In conclusion, our results further elucidate the diversity of carbapenemases, their mechanisms of horizontal transfer and possible patterns of co-selection. The study also emphasizes the difficulty of using whole genome sequencing for antimicrobial susceptibility testing of pathogens with complex genotypes.

Methicillin-resistant Staphylococcus argenteus misidentified as methicillin-resistant Staphylococcus aureus emerging in western Sweden.

Two strains included in a whole-genome sequencing project for methicillin-resistant Staphylococcus aureus (MRSA) were identified as non-Staphylococcus aureus when the sequences were analysed using the bioinformatics software ALEX (www.1928diagnostics.com, Gothenburg, Sweden). Sequencing of the sodA gene of these strains identified them as Staphylococcus argenteus. The collection of MRSA in western Sweden was checked for additional strains of this species. A total of 18 strains of S. argenteus isolated between 2011 and December 2017 were identified.

Subsequent infection with extended-spectrum ß-lactamase-producing Enterobacteriaceae in patients with prior infection or fecal colonization.

In clinical practice, there is a growing need to assess the impact of prior colonization or infection with extended-spectrum ß-lactamase-producing Enterobacteriaceae (EPE) on new EPE infections. We have investigated the frequency of, and duration to, a subsequent EPE infection in patients with prior fecal carriage or infection with EPE. Culture data for 3272 EPE-positive patients in Western Sweden during 2004-2014 were evaluated. The median follow-up time was 3.7 years. The first recorded EPE-positive fecal screen, or clinical (urine, blood) culture, and subsequent EPE-positive clinical samples were analyzed, focusing on the first and last recurrence of EPE infection. ESBL Escherichia coli dominated (95%). Almost all (94%) patients initially positive in fecal screen (n = 1436) and 72 and 71% of those initially positive in urine (n = 1717) and blood (n = 119) had no further EPE clinical isolates. Subsequent EPE bacteremia was only detected in 0.7, 1.6, and 4.2% of the respective patient group. Recurrent EPE-positive urine cultures occurred in 27% (460/1717), most (75%) within 6 months, and rarely (13%) after 1 year. Repeated EPE-positive clinical samples were significantly (p < 0.01) more common in patients > 65 years and in men with ESBL Klebsiella pneumoniae. In our low-endemic setting, subsequent EPE infections in previously colonized patients were rare. On the other hand, in patients previously EPE-positive in urine or blood, subsequent EPE urinary tract infections were common, especially within 6 months and in patients > 65 years old.

Development of a rapid MALDI-TOF MS based epidemiological screening method using MRSA as a model organism.

In this study we present a method using whole cell MALDI-TOF MS and VITEK MS RUO/SARAMIS as a rapid epidemiological screening tool. MRSA was used as a model organism for setting up the screening strategy. A collection of well-characterised MRSA strains representing the 19 most common Pulsed-Field Gel Electrophoresis (PFGE)-types in the region of South-West Sweden for the past 20 years was analysed with MALDI-TOF MS. A total of 111 MRSA strains were used for creating 19 PFGE-specific Superspectra using VITEK MS RUO/SARAMIS. Prior to performing the final analysis, the 19 Superspectra were combined into ten groups displaying similar peak patterns, hereafter named "MALDI-types". Two-hundred fifty-five MRSA strains were analysed to test the constructed Superspectra/MALDI-type database. Matches to the Superspectra above a threshold of 65% (corresponding to the number of matched peaks in the Superspectrum) were considered as positive assignment of a strain to a MALDI-type. The median peak matching value for correct assignment of a strain to a MALDI-type was 78% (range 65.3-100%). In total, 172 strains (67.4%) were assigned to the correct MALDI-type and only 5.5% of the strains were incorrectly assigned to another MALDI-type than the expected based on the PFGE-type of the strain. We envision this methodology as a cost-efficient step to be used as a first screening strategy in the typing scheme of MRSA isolates, to exclude epidemiological relatedness of isolates or to identify the need for further typing.

Corrigendum: Rapid identification of intact bacterial resistance plasmids via optical mapping of single DNA molecules.

This corrects the article DOI: 10.1038/srep30410.

Transfer and Persistence of a Multi-Drug Resistance Plasmid in situ of the Infant Gut Microbiota in the Absence of Antibiotic Treatment.

The microbial ecosystem residing in the human gut is believed to play an important role in horizontal exchange of virulence and antibiotic resistance genes that threatens human health. While the diversity of gut-microorganisms and their genetic content has been studied extensively, high-resolution insight into the plasticity, and selective forces shaping individual genomes is scarce. In a longitudinal study, we followed the dynamics of co-existing Escherichia coli lineages in an infant not receiving antibiotics. Using whole genome sequencing, we observed large genomic deletions, bacteriophage infections, as well as the loss and acquisition of plasmids in these lineages during their colonization of the human gut. In particular, we captured the exchange of multidrug resistance genes, and identified a clinically relevant conjugative plasmid mediating the transfer. This resistant transconjugant lineage was maintained for months, demonstrating that antibiotic resistance genes can disseminate and persist in the gut microbiome; even in absence of antibiotic selection. Furthermore, through in vivo competition assays, we suggest that the resistant transconjugant can persist through a fitness advantage in the mouse gut in spite of a fitness cost in vitro. Our findings highlight the dynamic nature of the human gut microbiota and provide the first genomic description of antibiotic resistance gene transfer between bacteria in the unperturbed human gut. These results exemplify that conjugative plasmids, harboring resistance determinants, can transfer and persists in the gut in the absence of antibiotic treatment.

Genome Dynamics of Escherichia coli during Antibiotic Treatment: Transfer, Loss, and Persistence of Genetic Elements In situ of the Infant Gut.

Elucidating the adaptive strategies and plasticity of bacterial genomes in situ is crucial for understanding the epidemiology and evolution of pathogens threatening human health. While much is known about the evolution of Escherichia coli in controlled laboratory environments, less effort has been made to elucidate the genome dynamics of E. coli in its native settings. Here, we follow the genome dynamics of co-existing E. coli lineages in situ of the infant gut during the first year of life. One E. coli lineage causes a urinary tract infection (UTI) and experiences several alterations of its genomic content during subsequent antibiotic treatment. Interestingly, all isolates of this uropathogenic E. coli strain carried a highly stable plasmid implicated in virulence of diverse pathogenic strains from all over the world. While virulence elements are certainly beneficial during infection scenarios, their role in gut colonization and pathogen persistence is poorly understood. We performed in vivo competitive fitness experiments to assess the role of this highly disseminated virulence plasmid in gut colonization, but found no evidence for a direct benefit of plasmid carriage. Through plasmid stability assays, we demonstrate that this plasmid is maintained in a parasitic manner, by strong first-line inheritance mechanisms, acting on the single-cell level, rather than providing a direct survival advantage in the gut. Investigating the ecology of endemic accessory genetic elements, in their pathogenic hosts and native environment, is of vital importance if we want to understand the evolution and persistence of highly virulent and drug resistant bacterial isolates.

Variation in Staphylococcus aureus Colonization in Relation to Disease Severity in Adults with Atopic Dermatitis during a Five-month Follow-up.

The aim of this study was to monitor Staphylococcus aureus colonization and disease severity in adults with atopic dermatitis (AD) during 5 months. Twenty-one patients attended 3 visits each for severity SCORing of Atopic Dermatitis (SCORAD) assessment, quantitative cultures from the skin and conventional cultures from the anterior nares, tonsils and perineum. S. aureus isolates were typed for strain identity with pulsed-field gel electrophoresis (PFGE). Seventy-one percent of patients were colonized with S. aureus on lesional skin at least once. Density (colony-forming units (CFU)/cm2) was higher on lesional skin than on non-lesional skin (p < 0.05). Density on lesional skin and number of colonized body sites were positively correlated with SCORAD (p = 0.0003 and p = 0.007, respectively). Persistent carriers of the same strain on lesional skin had higher mean SCORAD index than intermittent/non-carriers (36.3 and 17.1, respectively, p = 0.002). The results show a temporal correlation between several aspects of S. aureus colonization and disease severity in AD raising the question of the importance of this in pathogenesis and treatment.

Typing and Characterization of Bacteria Using Bottom-up Tandem Mass Spectrometry Proteomics.

Methods for rapid and reliable microbial identification are essential in modern healthcare. The ability to detect and correctly identify pathogenic species and their resistance phenotype is necessary for accurate diagnosis and efficient treatment of infectious diseases. Bottom-up tandem mass spectrometry (MS) proteomics enables rapid characterization of large parts of the expressed genes of microorganisms. However, the generated data are highly fragmented, making downstream analyses complex. Here we present TCUP, a new computational method for typing and characterizing bacteria using proteomics data from bottom-up tandem MS. TCUP compares the generated protein sequence data to reference databases and automatically finds peptides suitable for characterization of taxonomic composition and identification of expressed antimicrobial resistance genes. TCUP was evaluated using several clinically relevant bacterial species (Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus pneumoniae, Moraxella catarrhalis, and Haemophilus influenzae), using both simulated data generated by in silico peptide digestion and experimental proteomics data generated by liquid chromatography-tandem mass spectrometry (MS/MS). The results showed that TCUP performs correct peptide classifications at rates between 90.3 and 98.5% at the species level. The method was also able to estimate the relative abundances of individual species in mixed cultures. Furthermore, TCUP could identify expressed ß-lactamases in an extended spectrum ß-lactamase-producing (ESBL) E. coli strain, even when the strain was cultivated in the absence of antibiotics. Finally, TCUP is computationally efficient, easy to integrate in existing bioinformatics workflows, and freely available under an open source license for both Windows and Linux environments.

Evaluation of MLVA for epidemiological typing and outbreak detection of ESBL-producing Escherichia coli in Sweden.

BACKGROUND: To identify the spread of nosocomial infections and halt outbreak development caused by Escherichia coli that carry multiple antibiotic resistance factors, such as extended-spectrum beta-lactamases (ESBLs) and carbapenemases, is becoming demanding challenges due to the rapid global increase and constant and increasing influx of these bacteria from the community to the hospital setting. Our aim was to assess a reliable and rapid typing protocol for ESBL-E. coli, with the primary focus to screen for possible clonal relatedness between isolates. All clinical ESBL-E. coli isolates, collected from hospitals (n = 63) and the community (n = 41), within a single geographical region over a 6 months period, were included, as well as clinical isolates from a polyclonal outbreak (ST131, n = 9, and ST1444, n = 3). The sporadic cases represented 36 STs, of which eight STs dominated i.e. ST131 (n = 33 isolates), ST648 (n = 10), ST38 (n = 9), ST12 and 69 (each n = 4), ST 167, 405 and 372 (each n = 3). The efficacy of multiple-locus variable number tandem repeat analysis (MLVA) was evaluated using three, seven or ten loci, in comparison with that of pulsed-field gel electrophoresis (PFGE) and multi locus sequence typing (MLST). RESULTS: MLVA detected 39, 55 and 60 distinct types, respectively, using three (GECM-3), seven (GECM-7) or ten (GECM-10) loci. For GECM-7 and -10, 26 STs included one type and eleven STs each included several types, the corresponding numbers for GECM-3 were 29 and 8. The highest numbers were seen for ST131 (7,7 and 8 types, respectively), ST38 (5,5,8) and ST648 (4,5,5). Good concordance was observed with PFGE and GECM-7 and -10, despite fewer types being identified with MLVA; 78 as compared to 55 and 60 types. The lower discriminatory power of MLVA was primarily seen within the O25b-ST131 lineage (n = 34) and its H30-Rx subclone (n = 21). Epidemiologically unrelated O25b-ST131 isolates were clustered with O25b-ST131 outbreak isolates by MLVA, whereas the ST1444 outbreak isolates were accurately distinguished from unrelated isolates. CONCLUSION: MLVA, even when using only three loci, represents an easy initial typing tool for epidemiological screening of ESBL-E. coli. For the ST131-O25b linage, complementary methods may be needed to obtain sufficient resolution.

Draft Genome Sequence of Extended-Spectrum-ß-Lactamase-Producing Escherichia coli Strain CCUG 62462, Isolated from a Urine Sample.

The draft genome sequence has been determined for an extended-spectrum-ß-lactamase (ESBL)-producing (blaCTX-M-15) Escherichia coli strain (CCUG 62462), composed of 119 contigs and a total size of 5.27 Mb. This E. coli is serotype O25b and sequence type 131, a pandemic clonal group, causing worldwide antimicrobial-resistant infections.