ST131 Clone Research Articles

Molecular epidemiology of bloodstream-associated Escherichia coli ST131 H30-Rx subclone infection in a region with high quinolone resistance.

Bloodstream infections caused by Escherichia coli ST131 and ST131 H30-Rx subclones have emerged worldwide. This study was carried out to evaluate the prevalence of the ST131-Rx subclone and characterize the virulence properties of the Rx isolates among the bloodstream E. coli isolates. A total of 297 non-duplicated E. coli bloodstream isolates were studied. Antibiotic susceptibilities were tested using the disc diffusion method. PCR amplification and sequencing was used to identify ST131 and H30-Rx, the virulence gene, the β-lactamase and virotype. Quinolone resistance among bacteraemic E. coli strains was 51 %, and it was 98 % among E. coli ST131 isolates. The ST131 isolates accounted for 16 % (49) of all isolates and all ST131 isolates belonged to the extraintestinal pathogenic E. coli. The proportion of H30 subclone among the ST131 isolates was 98 %, and 75 % of H30 isolates belonged to the H30-Rx subclone. The prevalence of ST131 increased from 13 to 23 % in 4 years; however, there was a decrease in the ratio of H30-Rx infections. CTX-M-15 was detected in 85 % of ST131 and all of the H30-Rx isolates. The virulence genes associated with adhesion, cell protection, iron uptake and toxins (papA, iha, kpsMTII, iut and sat) were more common in ST131 than in non-ST131 isolates. Most of the ST131 and H30-Rx isolates were of the C virotype. All papA-positive isolates were in virotype C. The E. coli ST131 clone has increased rapidly among bloodstream isolates. However, a decrease in the proportion of the H30-Rx subclone in the quinolone-resistant population suggests the possibility of dissemination of other virulent and quinolone-resistant subclones in hospital settings.

Clonal dissemination of KPC-2 producing Klebsiella pneumoniae ST11 clone with high prevalence of oqxAB and rmtB in a tertiary hospital in China: results from a 3-year period.

BackgroundCarbapenemase-producing Klebsiella pneumoniae (CPKP) strains have emerged as a major problem for healthcare systems. The aim of this study was to determine the circulating clones and analyze the clinical and molecular characteristics of CPKP in our hospital.MethodsA total of 74 carbapenemase producers collected from our hospital from 2012 to 2014 were analyzed for the prevalence of extended-spectrum β-lactamase (ESBLs), plasmid-mediated quinolone resistance genes (PMQRs), exogenously acquired 16S rRNA methyltransferase (16S-RMTase), and plasmid-mediated AmpC enzyme (pAmpCs) by PCR and DNA sequencing. The sequence types (STs) of the carbapenemase producers were analyzed by multi-locus sequence typing (MLST). And Pulsed-field gel electrophoresis (PFGE) was performed to investigate the genetic relationship of KPC-2 producing strains. Clinical data were retrieved from the medical records.ResultsKPC-2 (n = 72) was the predominant enzyme followed by NDM-1 (n = 2); The genes blaCTX-M, blaSHV, blaTEM-1, blaDHA-1, rmtB, armA, oqxA, oqxB, and qnrB were present in 29 (39.2 %), 27 (36.5 %), 46 (62.2 %), 2 (2.7 %), 25 (33.8 %), 1 (1.4 %), 60 (81.1 %) and 56 (75.7 %), 6 (8.1 %) isolates, respectively. MLST analysis revealed 10 different STs. The most dominant ST was ST11 (78.4 %, 58/74), followed by ST15 (8.1 %, 6/74). PFGE patterns of the KPC-2 producing K. pneumoniae isolates exhibited clonal dissemination of ST11 and ST15 clones as well as a genetic diversity of the remaining strains.ConclusionThe intra- and inter-hospital cross-transmission of KPC-2-producing K. pneumoniae ST11 co-carrying oqxAB and rmtB in our hospital strongly suggested that rapid identification of colonized or infected patients and screening of carriers is quite necessary to prevent a scenario of endemicity.

Low intestinal colonization of Escherichia coli clone ST131 producing CTX-M-15 in Jordanian infants.

Over a period of 3 years' study (2012-2014), a total of 518 faecal samples were collected and cultured to isolate Escherichia coli. Of these, 338 (65.3%) E. coli isolates were recovered from infants, and 142/338 (42%) were multidrug-resistant (MDR) to ≥ 3 drug classes using the antimicrobial susceptibility disc diffusion method. A total of 125/142 (88%) of E. coli isolates were extended-spectrum β-lactamase (ESBL) producers. blaCTX-M-15 types were observed in 80/125 (64%) of the isolates, and 60/80 (75%) were positive for blaCTX-M-15. Out of 338 E. coli isolates, 9 (2.6%) were positive for ST131/O25b clone and each isolate was associated with several plasmids of different sizes (1-21.2 kb). The identities of these nine isolates were confirmed by sequencing for presence of pabB (347 bp) and trpA (427 bp) genes. This study demonstrates low prevalence rate of the highly virulent E. coli ST131 clone producing blaCTX-M-15 in the intestines of Jordanian infants.

A potential camel reservoir for extended-spectrum β-lactamase-producing Escherichia coli causing human infection in Saudi Arabia.

The prevalence of antimicrobial resistance is continuing to increase. Consequently, efficient approaches to identify sources of resistance are required. This study aimed to compare Escherichia coli isolates from the intestinal tract of camels with isolates from human urinary tract infections (UTIs) in Al Ahsa Province, Kingdom of Saudi Arabia (KSA), for antimicrobial resistance and identification of extended-spectrum β-lactamases (ESBLs). A microbiological study was conducted on 100 samples of cecal contents from camels and 100 urine samples from female UTI patients, to isolate and confirm E. coli using the VITEK 2 Automated System. Sensitivity patterns and identification of ESBLs were analyzed using the antimicrobial susceptibility test. Molecular techniques were used to detect E. coli drug-resistant clones. The presence rate of E. coli in camels was 26.0% (n = 26/100), and in human samples, the rate of E. coli was 33.0% (n = 33/100). ESBLs were reported for the first time in KSA, in 26.9% (n = 5/26) of camel samples and 36.4% (n = 8/33) of human samples. The multi-drug resistance (MDR) index was 0.13 and 0.17, for camels and humans, respectively. Escherichia coli drug-resistant O25b:H4-sequence type 131(ST131) clone was detected in two camel and two human isolates. This study demonstrates a high presence rate of ESBL-producing E. coli (ESBL-EC) in camels for the first time in KSA. Confirmation of MDR strains and E. coli ST131 clone in human and camel isolates suggests that camels could be a potential reservoir for resistant E. coli strains contributing to the increase in antimicrobial resistance in KSA.

Nosocomial dissemination of VIM-2-producing ST235 Pseudomonas aeruginosa in Lithuania.

Pseudomonas aeruginosa multidrug resistance, and particularly the production of carbapenemases linked to international high-risk clones, is of growing concern. While high levels of carbapenem resistance (>60 %) have been reported in Lithuania, so far, there is no information on the underlying mechanisms. Thus, the aim of this work was to determine the molecular epidemiology and prevalence of acquired carbapenemases among 73 carbapenem-resistant P. aeruginosa isolates recovered in a hospital from Kaunas, Lithuania in 2011-2012. The presence of acquired carbapenemases was evaluated through phenotypic (modified Hodge test, cloxacillin inhibition test, double-disc synergy test) and genetic methods [polymerase chain reaction (PCR) and sequencing]. Clonal relatedness was assessed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Acquired β-lactamases were detected in 19 (26 %) of the isolates, whereas resistance was exclusively chromosomal (OprD inactivation ± AmpC hyperproduction) in the remaining 54 (74 %) isolates. The acquired β-lactamases detected included 16 VIM-2, one PER-1 and two GES enzymes. PFGE revealed that 15 of the 16 VIM-2 isolates belonged to a single clone, identified as the international high-risk clone ST235 by MLST. bla VIM-2 was preceded by aacA7 in a class I integron, similar to epidemic ST235 isolates described in nearby countries. Additionally, sequencing of bla GES revealed the presence of the carbapenem-hydrolysing enzyme GES-5 in one of the isolates and a novel GES variant, designated GES-27, in the other. GES-27 differed from GES-5 by a single amino acid substitution, proline 167, that was replaced by glutamine. Increasing emergence and dissemination of concerning resistance mechanisms and international clones warrants global surveillance and control strategies.

The clinical impact of ST131 H30-Rx subclone in urinary tract infections due to multidrug-resistant Escherichia coli.

In this study, risk factors for ST131 H30 and H30-Rx subclones among urinary tract infections (UTIs) caused by multidrug-resistant (MDR) Escherichia coli were described. Urine samples were collected from consecutive outpatients registered to the outpatient clinics of Başkent University Hospital (Ankara, Turkey) with complaints of acute cystitis in 2011. A total of 107 MDR E. coli isolates were included in the study. Of the 107 isolates studied, 26 (24.3%) were typed as ST131 clone. Extended-spectrum β-lactamase (ESBL)-producers accounted for 59 (55.1%) of the 107 isolates. Among the 59 ESBL-positive isolates, 18 (31%) were found to belong to the ST131 clone. Of the 18 ESBL-positive ST131 isolates, 17 (94%) were defined as H30 subclone, among which 16 (94%) represented the H30-Rx subclone. Among the 48 ESBL-negative isolates, 8 (17%) ST131 isolates were detected, 7 (88%) of which belonged to H30 subclone; 5 (71%) of the H30 subclone isolates were classified under H30-Rx subclone. In multivariate analysis, hospitalisation within last year was the only host risk factor associated with MDR E. coli ST131 H30-Rx subclone UTI (OR=3.5, 95% CI 1.04-12.17; P=0.042). CTX-M-15 production was found to be highly associated with the presence of ST131 H30-Rx subclone (OR=4.8, 95% CI 1.54-15.32; P=0.007). In conclusion, urinary MDR E. coli ST131 H30-Rx subclone was found to be important in the dissemination of MDR UTIs in the community. Approximately 20% of the MDR isolates were H30-Rx subclone. Infection with this subclone was found to be healthcare-associated.

Comparison of the microbiological characteristics and virulence factors of ST131 and non-ST131 clones among extended-spectrum β-lactamase–producing Escherichia coli causing bacteremia

We evaluated the molecular epidemiology and microbiological characteristics of extended-spectrum β-lactamase–producing Escherichia coli (ESBL-EC) isolates that cause bacteremia in Korean hospitals, focusing especially on ST131. Our data suggest that ST131 isolates possessed more virulence traits and showed more multidrug resistance patterns than non-ST131 isolates. Among CTX-M-15 producers, the frequency of serum resistance was significantly higher in ST131 than in non-ST131. As in other parts of the world, the ESBL-EC ST131 clone has emerged and disseminated in both community and hospital settings in Korea, including in blood isolates in patients with bacteremia.

Characterization of the extra-intestinal pathogenic Escherichia coli ST131 clone among isolates recovered from urinary and bloodstream infections in the United Kingdom.

The multidrug-resistant ST131-O25b clone of Escherichia coli is well established as a significant cause of extra-intestinal infections worldwide. However, there have been only two small regional studies comparing ST131 isolates from the UK. Therefore, we characterized 143 ST131 E. coli (38 urinary, 105 bloodstream) collected between January 2011 and March 2012 from 38 centres located across the UK and Republic of Ireland. Phenotypic and genotypic characterization of clonal isolates revealed high rates of resistance to amoxicillin-clavulanate (56 %), cefotaxime (32 %), ciprofloxacin (79 %), temocillin (69 %, bloodstream isolates only), gentamicin (67 %) and trimethoprim-sulfamethoxazole (59 %). The most frequently detected extended-spectrum beta-lactamase was CTX-M-15 (87 %), predominantly encoded on IncF plasmids, although it was also associated with IncU plasmids in two isolates. The majority of UK ST131 clonal isolates possessed the O25b antigen (97 %) and the H30 fimH allele (92 %), but three serogroups (O19a, O136 and O153) novel to ST131 were identified among our strains. Contrary to previous reports, UK ST131-O16 isolates were typically susceptible to ciprofloxacin and lacked beta-lactamase genes (n = 12/12). In summary, ST131 strains of E. coli circulating in the UK possess characteristic clonal features, but are becoming more diverse than other international ST131 populations.

Bloodstream Infections due to Enterobacteriaceae Among Neonates in Poland – Molecular Analysis of the Isolates

Bloodstream infections (BSIs) are associated with a significantly increased risk of fatality. No report has been found about the molecular epidemiology of Enterobacteriaceae causing BSI in neonates in Poland. The aim of this work was to determine the antibiotic resistance profiles, virulence gene prevalence, the epidemiological and genetic relationships among the isolates from Enterobacteriaceae causing BSI in neonates with birth weight < 1501 g. Antimicrobial susceptibility testing was performed. PCR was performed to identify the presence of common beta-lactamase genes, virulence genes. PFGE and MLST were performed. The surveillance group contained 1,695 newborns. The incidence rate for BSIs was 5.9%, the fatality rate 15%. The most common species were Escherichia coli (n = 24) and Klebsiella pneumoniae (n = 16). CTX-M-15 was found in 6 E. coli, 8 K. pneumoniae, 1 Enterobacter cloacae strains. Among E. coli fimH (83.3%), ibeA (37.5%), neuC (20.8%) were the most frequent. PFGE demonstrated unique pulsotypes among E. coli. E. coli ST131 clone was found in 7 E. coli strains. PFGE of 16 K. pneumoniae strains showed 8 pulsotypes. Five isolates from one NICU belonged to one clone. MLST typing revealed 7 different ST with ST336 as the most prevalent. This study provides information about resistance, virulence and typing of Enterobacteriaceae strains causing BSI among neonates. E. coli and Klebsiella spp. isolated in this study have completely different epidemiology from each other.

Srr2, a multifaceted adhesin expressed by ST-17 hypervirulent Group B Streptococcus involved in binding to both fibrinogen and plasminogen.

The Group B Streptococcus (GBS) 'hypervirulent' ST-17 clone is strongly associated with invasive neonatal meningitis. Comparative genome analyses revealed that the serine-rich repeat (Srr) glycoprotein Srr2 is a cell wall-anchored protein specific for ST-17 strains, the non-ST-17 isolates expressing Srr1. Here, we unravel the binding capacity of GBS Srr proteins to relevant components of the host fibrinolysis pathway. We demonstrate that: (i) Srr2 binds plasminogen and plasmin whereas Srr1 does not; (ii) the ability of ST-17 strains to bind fibrinogen reflects a high level surface display of Srr2 combined with a higher affinity of Srr2 than Srr1 to bind this ligand; and (iii) Srr2 binding to host plasma proteins results in the formation of bacterial aggregates that are efficiently endocytosed by phagocytes. Importantly, we show that Srr2 increased bacterial survival to phagocytic killing and bacterial persistence in a murine model of meningitis. We conclude that Srr2 is a multifaceted adhesin used by the ST-17 clone to hijack ligands of the host coagulation system, thereby contributing to bacterial dissemination and invasiveness, and ultimately to meningitis.

An ST11 clone of Pasteurella multocida, widely spread among farmed rabbits in the Iberian Peninsula, demonstrates respiratory niche association

Pasteurella multocida is a veterinary pathogen causing diseases with considerable economic repercussions in a wide range of animal hosts. In rabbits, P. multocida infections cause a variety of clinical manifestations including rhinitis, pneumonia, septicemia, abscesses, mastitis, and pyometra. In this study, 100 P. multocida isolates from different commercial rabbit farms located throughout the Iberian Peninsula were molecularly characterized by capsular typing, detection of four virulence-associated genes (tbpA, toxA, hgbB, and pfhA), and multilocus sequence typing (MLST). Rabbit P. multocida isolates belonged to three different capsular types: A (47.0%), D (28.0%), and F (25.0%). One group of P. multocida isolates of capsular type D and positive for the hgbB gene was significantly associated with the clinical presentation of respiratory disease (OR 5.91; 95%CI, 1.63–21.38). These isolates belonged to same sequence type, ST11, in the P. multocida Multi-host MLST database. The ST11 clone also includes isolates from porcine and avian pneumonia. This clonal group of epidemiologically unrelated P. multocida isolates could be a virulent clone with some degree of specificity for respiratory disease. These findings could be relevant in the development of vaccines for pasteurellosis prevention, especially respiratory disease.

Molecular characterization of extended-spectrum β-lactamase-producing Escherichia coli and Klebsiella spp. isolates in Mongolia.

The aim of this study was to determine the molecular characteristics of β-lactamase genes in extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae isolates from Mongolia. Fifty-six ESBL-producing Enterobacteriaceae isolates were collected, of which 46 were Escherichia coli, seven were Klebsiella pneumoniae, and three were K. oxytoca. Minimum inhibitory concentrations for selected antibiotics were tested using the agar dilution method, and the β-lactamase genes were determined using polymerase chain reaction combined with sequencing. Pulsed-field gel electrophoresis (PFGE) was used for genotyping all isolates, and phylogenetic grouping was performed on ESBL-producing E. coli isolates. Conjugation tests combined with plasmid digestion assays were used to determine whether there was a horizontal spread in Mongolia. Among the 56 ESBL-producing isolates, 43 isolates (76.8%) were resistant to fluoroquinolones, but all isolates were susceptible to carbapenems and amikacin. The polymerase chain reaction sequencing results showed that the dominant CTX-M genotype was CTX-M-15 (19/46, 41.3%) in the ESBL-producing E. coli isolates. By contrast, CTX-M-14 and CTX-M-3 were the major genotypes found in Klebsiella spp. Phylogenetic analysis revealed that 21 ESBL-producing E. coli isolates belonged to group D (21/46, 45.6%), followed by group A (13/46, 28.3%), group B2 (11/46, 23.9%), and group B1 (1/46, 2.2%). Only four E. coli isolates (4/46, 8.7%) belonged to the ST131 clone. PFGE showed that the ESBL-producing Enterobacteriaceae were genetically unrelated. The conjugation assay showed that two plasmids harboring CTX-M-15 in E. coli isolates were genetic unrelated, whereas seven plasmids harboring CTX-M-14 (5/7 and 2/7) and four plasmids harboring CTX-M-55 (4/4) showed genetic relatedness, indicating the dissemination of resistance plasmids in this area.

Emergence and spread of O16-ST131 and O25b-ST131 clones among faecal CTX-M-producing Escherichia coli in healthy individuals in Hunan Province, China.

ObjectivesThe objectives of this study were to determine CTX-M-producing Escherichia coli ST131 strain prevalence in stool specimens from healthy subjects in central China and to molecularly characterize clonal groups.MethodsFrom November 2013 to January 2014, stool specimens from healthy individuals in Hunan Province were screened for ESBL-producing E. coli using chromogenic medium and CTX-M genotypes and phylogenetic groups were determined. ST131 clonal groups were detected by PCR and characterized for antibiotic resistance, fimH, gyrA and parC alleles, plasmid-mediated quinolone resistance determinants, virulence genotypes and PFGE patterns.ResultsAmong 563 subjects, 287 (51.0%) exhibited the presence of faecal ESBL-producing E. coli, all of which produced CTX-M enzymes. The most common CTX-M genotypes were CTX-M-14 (48.4%), CTX-M-15 (27.5%) and CTX-M-27 (15.0%). Of the 287 CTX-M-producing isolates, 32 (11.1%) belonged to the ST131 clone. O16-ST131 isolates were dominant (75%) and contained the fimH41 allele. The remaining eight (25%) ST131 isolates were of the O25b subgroup and contained fimH30 or fimH41. Ciprofloxacin resistance was found in 100% of the O25b-ST131 isolates, whereas only 8% of the O16-ST131 isolates were resistant. All of the O25b-ST131 isolates except one showed gyrA1AB and parC1aAB mutations; most of the O16-ST131 isolates had gyrA1A and parC1b mutations. The virulence genotypes of O16-ST131 resembled those of the O25b-ST131 isolates. The 32 ST131 isolates formed one large group at the 64% similarity level. They comprised 15 PFGE groups (defined at ≥85% similarity).ConclusionsO16-ST131 isolates have emerged as the predominant type of ST131 isolate in faecal CTX-M-producing E. coli in healthy individuals in China.

Characterization of Inhibitor-Resistant TEM β-Lactamases and Mechanisms of Fluoroquinolone Resistance in Escherichia coli Isolates.

The aim of present work was to characterize the inhibitor-resistant TEM (IRT) β-lactamases produced by Escherichia coli in Hospital Clínico San Carlos (Madrid, Spain). Mechanisms of fluoroquinolone resistance among IRT-producing strains were also studied. Isolates with susceptibility to cephalosporins and amoxicillin-clavulanate (AMC) resistance were collected in our hospital (November 2011-July 2012) from both outpatients and hospitalized patients. Among 70 AMC-resistant E. coli strains, 28 (40%) produced IRT enzymes. Most of them were uropathogens (82.1%) and recovered from outpatients (75%). Seven different IRT enzymes were identified with TEM-30 (IRT-2) being the most prevalent, followed by TEM-40 (IRT-11). A high rate of ciprofloxacin resistance was found among IRT-producing strains (50%). Most of the ciprofloxacin-resistant isolates showed ciprofloxacin minimum inhibitory concentration >32 mg/L and contained two mutations in both gyrA and parC genes. Four IRT enzyme producers harbored the qnr gene. ST131 clone was mainly responsible for both IRT enzyme production and ciprofloxacin resistance. In conclusion, data from this study show that the frequency of IRT producers was 40% and a high rate of ciprofloxacin resistance was found among IRT-producing isolates. Current and future actions should be taken into account to avoid or reduce the development of AMC and fluoroquinolone resistance in E. coli.

Molecular characterization of a multidrug resistance IncF plasmid from the globally disseminated Escherichia coli ST131 clone.

Escherichia coli sequence type 131 (E. coli ST131) is a recently emerged and globally disseminated multidrug resistant clone associated with urinary tract and bloodstream infections. Plasmids represent a major vehicle for the carriage of antibiotic resistance genes in E. coli ST131. In this study, we determined the complete sequence and performed a comprehensive annotation of pEC958, an IncF plasmid from the E. coli ST131 reference strain EC958. Plasmid pEC958 is 135.6 kb in size, harbours two replicons (RepFIA and RepFII) and contains 12 antibiotic resistance genes (including the bla CTX-M-15 gene). We also carried out hyper-saturated transposon mutagenesis and multiplexed transposon directed insertion-site sequencing (TraDIS) to investigate the biology of pEC958. TraDIS data showed that while only the RepFII replicon was required for pEC958 replication, the RepFIA replicon contains genes essential for its partitioning. Thus, our data provides direct evidence that the RepFIA and RepFII replicons in pEC958 cooperate to ensure their stable inheritance. The gene encoding the antitoxin component (ccdA) of the post-segregational killing system CcdAB was also protected from mutagenesis, demonstrating this system is active. Sequence comparison with a global collection of ST131 strains suggest that IncF represents the most common type of plasmid in this clone, and underscores the need to understand its evolution and contribution to the spread of antibiotic resistance genes in E. coli ST131.

Insight into neonatal septicaemic Escherichia coli from India with respect to phylogroups, serotypes, virulence, extended-spectrum-β-lactamases and association of ST131 clonal group.

The study characterizes a collection of 67 neonatal septicaemic Escherichia coli isolates on the basis of phylogroup, serotype, virulence, antibiotic resistance and also the association of CTX-M-producing E. coli and the ST131 clone in a developing country. Phylogroups B2 and D were predominant (33% and 19%, respectively). The most prevalent virulence factors (VFs) were traT (69%) and iucC (68%) and most VFs were concentrated in the B2 isolates. High levels of resistance (⩾70%) to cefotaxime, ciprofloxacin and trimethoprim/sulfamethoxazole was recorded but meropenem remained the most active antimicrobial. Six (9%) of the study isolates belonged to the ST131 clone, five of which were from the same hospital, and were either indistinguishable or closely related by pulsed-field gel electrophoresis. Although the prevalence of CTX-M-15-producing isolates was high (81%), the ST131 clone was relatively infrequent (11%) in extended spectrum β-lactamase (ESBL)-producers. The ST131 clone was characterized by the presence of bla CTX-M-15, qnrS, aac(6')-Ib-cr, IncF plasmids and virulence determinants such as iucC, papC, traT, usp, hlyA, iroN E.coli , cnf, and sat. We conclude that clonal spread of ST131 did not contribute directly to the high prevalence of CTX-M-15 in our settings.

Molecular epidemiology of carbapenemase-producing Klebsiella pneumoniae in a hospital in Madrid: Successful establishment of an OXA-48 ST11 clone

Here we report a retrospective clinical and molecular study conducted in a tertiary care facility in southern Madrid, Spain, from January 2009 to February 2014 to investigate the epidemiology of carbapenemase-producing Klebsiella pneumoniae (CPKp). Carbapenemase genes were identified in 97 non-duplicate K. pneumoniae isolates, including 59 harbouring blaOXA-48, 37 harbouring blaVIM-1 and 1 harbouring blaKPC-2. Pulsed-field gel electrophoresis (PFGE) analysis verified the presence of 20 different clonal types, whilst multilocus sequence typing (MLST) assigned the isolates to eight sequence types (STs). A gradual increase was noted in the number of CPKp isolated, ranging from 0.8% in 2009 to 4.3% in 2013. A large outbreak was also identified, initiated in 2013 owing to a blaOXA-48 and blaCTX-M-15 co-producing ST11 clone and involving a total of 44 patients. Whole-genome sequencing was used to characterise the resistome of a representative isolate from this outbreak. Bioinformatics analysis revealed the presence of 121 genes related to antibiotic and antiseptic resistance, mutations in the ompk35 and ompk36 genes, and the presence of the blaOXA-48 gene on a 62 811bp IncL/M-type plasmid as part of a Tn1999.2 composite transposon. These results portray the increasing trend in carbapenemase-producing isolates in this hospital and highlight the successful establishment of a blaOXA-48 and blaCTX-M-15 co-producing ST11 clone that has led to the displacement of previous circulating clones.

Emergence of Antibiotic Resistance-Associated Clones Among Escherichia coli Recovered From Newborns With Early-Onset Sepsis and Meningitis in the United States, 2008-2009.

Escherichia coli associated with early-onset sepsis (EOS) have historically been antibiotic-susceptible and K1-encapsulated. In the era of emerging antibiotic resistance, however, the clonal makeup of E coli associated with EOS has not been well characterized. Escherichia coli isolates were collected from 28 cases of EOS and early-onset meningitis (EOM) from April 2008 through December 2009, during a parent study conducted at National Institute of Child Health and Human Development Neonatal Research Network centers from February 2006 through December 2009. Clinical and microbiologic data were collected for the parent study. We applied polymerase chain reaction- and sequence-based molecular techniques to determine clonal, virulence-associated and antibiotic resistance-associated traits of the E coli isolates. Among 28 E coli strains, phylogroup B2 strains predominated (68%), of which more than half were K1-encapsulated (53%). Phylogroup D strains were prominent as well (18%), but none were K1-encapsulated. Across the strain collection, the rate of ampicillin resistance was high (78%). The sole strain resistant to either extended-spectrum cephalosporins or fluoroquinolones represented ST131 H30-Rx, the multidrug-resistant subclone that has emerged worldwide in the last decade. This strain encoded extended-spectrum β-lactamase CTX-M-15 and carried an IncF plasmid of type F2:A1:B-. In this collection of EOS/EOM-associated E coli isolates, we observed a high rate of ampicillin resistance, a low rate of fluoroquinolone resistance, and no aminoglycoside resistance, with resistance to third-generation cephalosporins appearing in only a single strain, from the worldwide emerging ST131 clone. Ongoing surveillance of antibiotic resistance among EOS isolates is warranted, to ensure that standard empiric regimens remain effective.

Prevalence and distribution of beta-lactamase coding genes in third-generation cephalosporin-resistant Enterobacteriaceae from bloodstream infections in Cambodia.

Resistance to third-generation cephalosporins in Gram-negative bacteria is emerging in Asia. We report the prevalence and distribution of extended-spectrum beta-lactamase (ESBL), AmpC beta-lactamase and carbapenemase-coding genes in cefotaxime-resistant Enterobacteriaceae isolates from bloodstream infections (BSI) in Cambodia. All Enterobacteriaceae isolated from BSI in adult patients at Sihanouk Hospital Centre of HOPE, Phnom Penh, Cambodia (2007–2010) were assessed. Antimicrobial susceptibility testing was carried out by disc diffusion and MicroScan according to Clinical and Laboratory Standards Institute (CLSI) guidelines. Screening for ESBL, plasmidic AmpC and carbapenemase-coding genes was performed by multiplex polymerase chain reaction (PCR) sequencing assays. Identification of the ST131 clone was performed in all CTX-M-positive Escherichia coli, using PCR targeting the papB gene. Out of 183 Enterobacteriaceae, 91 (49.7 %) isolates (84 BSI episodes) were cefotaxime-resistant: E. coli (n = 68), Klebsiella pneumoniae (n = 17) and Enterobacter spp. (n = 6). Most episodes were community-acquired (66/84; 78.3 %). ESBLs were present in 89/91 (97.8 %) cefotaxime-resistant isolates: 86 (96.6 %) were CTX-M, mainly CTX-M-15 (n = 41) and CTX-M-14 (n = 21). CTX-M of group 1 were frequently associated with TEM and/or OXA-1/30 coding genes and with phenotypic combined resistance to ciprofloxacin, sulphamethoxazole–trimethoprim and gentamicin (39/50, 78.0 %). Plasmidic AmpC (CMY-2 and DHA-1 types) were found alone (n = 2) or in combination with ESBL (n = 4). Eighteen E. coli isolates were identified as B2-ST131-O25B: 11 (61.1 %) carried CTX-M-14. No carbapenemase-coding genes were detected. ESBL among Enterobacteriaceae from BSI in Cambodia is common, mainly associated with CTX-M-15 and CTX-M-14. These findings warrant urgent action for the containment of antibiotic resistance in Cambodia.

Sequence type 131 fimH30 and fimH41 subclones amongst Escherichia coli isolates in Australia and New Zealand

The clonal composition of Escherichia coli causing extra-intestinal infections includes ST131 and other common uropathogenic clones. Drivers for the spread of these clones and risks for their acquisition have been difficult to define. In this study, molecular epidemiology was combined with clinical data from 182 patients enrolled in a case–control study of community-onset expanded-spectrum cephalosporin-resistant E. coli (ESC-R-EC) in Australia and New Zealand. Genetic analysis included antimicrobial resistance mechanisms, clonality by DiversiLab (rep-PCR) and multilocus sequence typing (MLST), and subtyping of ST131 by identification of polymorphisms in the fimH gene. The clonal composition of expanded-spectrum cephalosporin-susceptible E. coli and ESC-R-EC isolates differed, with six MLST clusters amongst susceptible isolates (median 7 isolates/cluster) and three clusters amongst resistant isolates, including 40 (45%) ST131 isolates. Population estimates indicate that ST131 comprises 8% of all E. coli within our population; the fluoroquinolone-susceptible H41 subclone comprised 4.5% and the H30 subclone comprised 3.5%. The H30 subclone comprised 39% of all ESC-R-EC and 41% of all fluoroquinolone-resistant E. coli within our population. Patients with ST131 were also more likely than those with non-ST131 isolates to present with an upper than lower urinary tract infection (RR=1.8, 95% CI 1.01–3.1). ST131 and the H30 subclone were predominant amongst ESC-R-EC but were infrequent amongst susceptible isolates where the H41 subclone was more prevalent. Within our population, the proportional contribution of ST131 to fluoroquinolone resistance is comparable with that of other regions. In contrast, the overall burden of ST131 is low by global standards.