Sequence Type ST131 Research Articles

Extended-spectrum β-lactamase-producing Enterobacteriaceae from animal origin and wastewater in Tunisia: first detection of O25b-B23-CTX-M-27-ST131 Escherichia coli and CTX-M-15/OXA-204-producing Citrobacter freundii from wastewater.

This study aimed to isolate and characterise extended-spectrum β-lactamase-producing Enterobacteriaceae (ESBL-E) isolates from animals and wastewater in Tunisia. ESBL-E from wastewater (n=123 samples), faeces of healthy animals (poultry, sheep, goats and calves) (n=140) and raw milk from healthy cows (n=42) and goats (n=20) were investigated. Antimicrobial susceptibility was determined according to CLSI recommendations. The blaTEM, blaSHV, blaCTX-M and blaOXA-48 genes were analysed by PCR and sequencing. Phylogenetic groups were determined by PCR for Escherichia coli isolates. The clonality of E. coli and Klebsiella pneumoniae isolates was determined by XbaI-PFGE and MLST. A total of 81 E. coli, 20 K. pneumoniae, 4 Enterobacter cloacae, 1 Citrobacter freundii and 1 Citrobacter braakii were isolated. The blaCTX-M-1 and blaCTX-M-15 genes were predominant in E. coli and K. pneumoniae isolates. E. cloacae and C. braakii isolates harboured the blaSHV-12 gene. The C. freundii isolated from wastewater carried blaCTX-M-15, blaTEM-1 and blaOXA-204. E. coli isolates belonged to phylogroups A (37), B1 (25), B2 (7) and D (12). Seventy-eight E. coli isolates were typeable by PFGE and were classified into 34 pulsotypes. The K. pneumoniae isolates belonged to 11 pulsotypes. The E. coli isolates belonged to sequence types ST131, ST224, ST162, ST845, ST5204, ST69, ST141 and ST10. The K. pneumoniae isolates belonged to ST405, ST147, ST564, ST307, ST152, ST45, ST661 and ST1564. This is the first report of O25b-B23-CTX-M-27-ST131 E. coli isolates and of C. freundii carrying blaCTX-M-15, blaTEM-1 and blaOXA-204 in Tunisia.

Dissemination of CTX-M-Producing Escherichia coli in Freshwater Fishes From a French Watershed (Burgundy).

The burden of extended-spectrum β-lactamases producing Escherichia coli (ESBL-Ec), has increased over several decades. Freshwater ecosystems are suspected to play an important ecological and evolutionary role in driving the dissemination of antimicrobial resistance. The aim of our study was to decipher the occurrence of ESBL-Ec in a small watershed (Ouche river, Burgundy, France), targeting environmental matrices and fishes. Among cefotaxime resistant E. coli (ctxR Ec) isolates, we detected and characterized 36 ESBL-Ec from water, biofilm and fish guts. ctxR Ec and ESBL-Ec were found in samples from sites near the first small town, located downstream from the watershed which was studied. Treatment of urban wastewater by waste water treatment plants (WWTP), might therefore be a major potential source of ctxR Ec and thus of ESBL-Ec. Prevalence of total E. coli and ctxR Ec in fish guts ranged between 0 to 92% and 0 to 85%; respectively, depending on the sampling site and the fish species. The diet of fish (predator or omnivore) seems to strongly influence the prevalence of total E. coli and ESBL-Ec. Extended spectrum beta-lactamases produced by the isolates from this study belonged to the CTX-M family (CTX-M group 1 and 9). Moreover, some environmental ESBL-Ec proved to share genotypic features (MLST types) with isolates which originated from 8 WWTP effluents discharged in the Ouche river and with the sequence type ST131, which is widely described in clinical isolates. Ninety-seven % (97%) of ESBL-Ec from the study harbored additional antibiotic resistances and can thus be considered as multi drug resistant (MDR) bacteria. Finally, 53% of the ESBL-Ec strains harbored class 1 integron-integrase (intl1). These results are discussed with the perspective of defining indicators of antibiotic resistance contamination in freshwater ecosystems.

A Nosocomial Respiratory Infection Outbreak of Carbapenem-Non-Susceptible <i>Escherichia coli</i> ST131 with Multiple Transmissible <i>blaKPC-2</i> Carrying Plasmids

Background: Escherichia coli sequence type (ST) ST131 is one of the most widespread and successful opportunistic pathogenic E. coli clones known as multiple-resistance. It can cause community- and hospital-acquired urinary tract infection, abdominal and pelvic infection, and bacteraemia with high morbidity and mortality. Currently, carbapenems have been considered the treatment of choice for E. coli ST131 infections. Although carbapenem resistance is still rare, the resistance rate to carbapenems is growing slowly mainly due to the acquisition of β-lactamase and carbapenemase. In this study, we report a nosocomial outbreak of predominantly respiratory infections caused by blaKPC-2 carrying E. coli ST131 in a Chinese hospital and revealed the transmission of carbapenem-non-susceptible E. coli, as well as the acquisition and spread of blaKPC-2 by E. coli ST131. Methods: A total of 45 carbapenem-non-susceptible E.coli strains were collected in one Beijing hospital from July 2012 to December 2013. The characteristic of these strains including antimicrobial susceptibility, carbapenem resistance mechanism, plasmid profiles and genetic background were determined by two-fold broth microdilution method, PCR, PFGE-S1 southern blot hybridization and whole genome sequencing respectively. Phylogenetic relationships between these strains were inferred. The genetic context of blaKPC-2 was analyzed to trace the spread of blaKPC-2. Findings: 45 carbapenem-non-susceptible E. coli strains were isolated from patients mainly with respiratory infections in this nosocomial outbreak. These strains carried blaKPC-2 and belonged to ST131. These strains are closely related and divided into multiple lineages in the phylogenetic tree. It was initially prevalent only in the ward E02, then spread to other wards and multiple lineages co-evolved, resulting in a nosocomial respiratory infection outbreak of carbapenem-non-susceptible E. coli ST131 in these geriatric wards. The blaKPC-2 positive plasmid profiles suggested that the carbapenem resistance was due to the acquisition of multiple transmissible plasmids carrying blaKPC-2 by E. coli ST131. These plasmids with different backbone sequence exhibit wealthy multidrug resistance (MDR) regions and cointegrate structure at the mediate of IS26. Diverse multidrug resistance elements were transferred and rearranged between these plasmids with different backbone sequence at the mediate of IS26, resulting in the rapid spread of antibiotic-resistant genes and blaKPC-2 between these strains. Interpretation: E. coli ST131 has become a multidrug epidemic clone that can cause hospital-acquired respiratory infections that have been less aware. Our research indicates that clinical attention should be paid to the importance of E. coli ST131 in respiratory infections. The emergence of blaKPC in E. coli ST131 may herald the beginning for transmission of blaKPC with high prevalence in communities and hospitals. Close monitoring of blaKPC-2 positive ST131 in hospitals and communities are necessary for control and proper treatment of ST131 infections. Funding Statement: This work was supported by the National Natural Science Foundation of China (81861138053，31761133004 and 81861138004) and the Ministry of Science and Technology (2018ZX10733-402). Declaration of Interests: All authors declare that there are no conflicts of interest. Ethics Approval Statement: Not needed.

Isolation of the first New Delhi metallo-ß-lactamase-1 (NDM-1)-producing and colistin-resistant Klebsiella pneumoniae sequence type ST15 from a digestive carrier in Albania, May 2018.

Carbapenemases represent a public health threat, as they can spread through horizontal gene transfer and cause outbreaks. New Delhi metallo-ß-lactamase-1 (NDM-1) is a metallo-ß-lactamase that has spread rapidly in the last decade, causing worldwide alarm. This study aimed to describe the first isolate of NDM-1-producing and extensively drug resistant Klebsiella pneumoniae in Albania, its clinical context and genetic characterization. Strain was isolated from both oral and rectal intensive care unit admission screening swabs of a 70-year-old male patient with no history of international travel in the previous 6 months. Sequencing was performed by Illumina NextSeq500 platform, with a paired-end run of 2 by 150bp, after Nextera XT paired-end library preparation. Sequencing reads were assembled using SPAdes Genome (version 3.6.1) with accurate de novo settings. The assembled contigs were uploaded into the online tools: BIGSdb-Kp, ResFinder and PlasmidFinder. Isolate was resistant to all tested antibiotics but tigecycline and trimethoprim-sulfamethoxazole. Sequencing revealed the presence of acquired resistance genes conferring resistance to β-lactams (blaNDM-1, blaCMY-6, blaCTX-M-15and blaSHV-28), aminoglycosides (rmtC, aac(6')-Ib3), fluoroquinolones (oqxA, oqxB, aac(6')-Ib-cr), fosfomycin (fosA) and sulfonamides (sul1). The blaNDM-1 gene was located on an IncA/C2 plasmid. Plasmid mediated mcr-1 to mcr-8 genes were absent in both isolates. Resistance to colistin was due to an amino acid substitution (Thr157Pro) in PmrB protein. NDM-1-producing Enterobacteriaceae are spreading in the Balkans. Identification of NDM-1-producing and extensively drug resistant K. pneumoniae ST15 in Albania is a cause for serious concern. There should be a continuous national and Balkan multinational surveillance of blaNDM-1-carrying isolates.

Methicillin-resistant Staphylococcus aureus: livestock-associated, antimicrobial, and heavy metal resistance.

Staphylococcus aureus is an opportunistic pathogen of human and other mammals that is of increasing clinical and veterinary importance due to its ability to rapidly develop antimicrobial resistance. The injudicious use of antibiotics has given rise to the emergence of antibiotic resistant S. aureus strains, most importantly methicillin-resistant Staphylococcus aureus (MRSA). The emergence of livestock-associated MRSA (LA-MRSA) has highlighted the importance of directed research toward its prevention, as well as the need for the discovery and development of more efficient treatment than is currently available. Furthermore, the treatment of MRSA is complicated by the co-selection of heavy metal and antibiotic resistance genes by microorganisms. Livestock and livestock production systems are large reservoirs of heavy metals due to their use in feed as well as environmental contaminant, which has allowed for the selection of LA-MRSA isolates with heavy metal resistance. The World Health Organization reported that Africa has the largest gaps in data on the prevalence of antimicrobial resistance, with no reports on rates for LA-MRSA harboring heavy metal resistance in South Africa. This review aimed to report the emergence of LA-MRSA in South Africa, specifically the most frequent sequence type ST398, globally. Furthermore, we aimed to highlight the importance of LA-MRSA in clinical and food security, as well as this research gap in South Africa. This review sheds light on the prevalence of heavy metals in livestock farms and abattoirs, and focuses on the phenomenon of the co-selection of heavy metal and antibiotic resistance genes in MRSA, emphasizing the importance of a focused direction for research in humans, animals as well as environment using one-health approach.

Escherichia coli Sequence Type 410 Is Causing New International High-Risk Clones.

Escherichia coli sequence type 410 (ST410) has been reported worldwide as an extraintestinal pathogen associated with resistance to fluoroquinolones, third-generation cephalosporins, and carbapenems. In the present study, we investigated national epidemiology of ST410 E.coli isolates from Danish patients. Furthermore, E.coli ST410 was investigated in a global context to provide further insight into the acquisition of the carbapenemase genes blaOXA-181 and blaNDM-5 of this successful lineage. From 127 whole-genome-sequenced isolates, we reconstructed an evolutionary framework of E.coli ST410 which portrays the antimicrobial-resistant clades B2/H24R, B3/H24Rx, and B4/H24RxC. The B2/H24R and B3/H24Rx clades emerged around 1987, concurrently with the C1/H30R and C2/H30Rx clades in E.coli ST131. B3/H24Rx appears to have evolved by the acquisition of the extended-spectrum β-lactamase (ESBL)-encoding gene blaCTX-M-15 and an IncFII plasmid, encoding IncFIA and IncFIB. Around 2003, the carbapenem-resistant clade B4/H24RxC emerged when ST410 acquired an IncX3 plasmid carrying a blaOXA-181 carbapenemase gene. Around 2014, the clade B4/H24RxC acquired a second carbapenemase gene, blaNDM-5, on a conserved IncFII plasmid. From an epidemiological investigation of 49 E.coli ST410 isolates from Danish patients, we identified five possible regional outbreaks, of which one outbreak involved nine patients with blaOXA-181- and blaNDM-5-carrying B4/H24RxC isolates. The accumulated multidrug resistance in E.coli ST410 over the past two decades, together with its proven potential of transmission between patients, poses a high risk in clinical settings, and thus, E.coli ST410 should be considered a lineage with emerging "high-risk" clones, which should be monitored closely in the future.IMPORTANCE Extraintestinal pathogenic Escherichia coli (ExPEC) is the main cause of urinary tract infections and septicemia. Significant attention has been given to the ExPEC sequence type ST131, which has been categorized as a "high-risk" clone. High-risk clones are globally distributed clones associated with various antimicrobial resistance determinants, ease of transmission, persistence in hosts, and effective transmission between hosts. The high-risk clones have enhanced pathogenicity and cause severe and/or recurrent infections. We show that clones of the E.coli ST410 lineage persist and/or cause recurrent infections in humans, including bloodstream infections. We found evidence of ST410 being a highly resistant globally distributed lineage, capable of patient-to-patient transmission causing hospital outbreaks. Our analysis suggests that the ST410 lineage should be classified with the potential to cause new high-risk clones. Thus, with the clonal expansion over the past decades and increased antimicrobial resistance to last-resort treatment options, ST410 needs to be monitored prospectively.

Fecal carriage of multidrug-resistant Escherichia coli by community children in southern Taiwan

BackgroundThe emergence of multidrug-resistant (MDR) Escherichia coli (E. coli), particularly E. coli sequence type ST131, is becoming a global concern. Commensal bacteria, an important reservoir of antibiotic resistance genes, facilitate the spread of such genes to pathogenic bacterial strains. The objective of the study is to investigate the fecal carriage of MDR E. coli and ST131 E. coli in community children in Southern Taiwan.MethodsIn this prospective study, stool samples from children aged 0–18 years were obtained within 3 days of hospitalization from October 2013 to September 2014. Children with a history of underlying diseases, antibiotic treatment, or hospitalization in the 3 months before specimen collection were excluded. E. coli colonies were selected and tested for antimicrobial susceptibility, and O25b-ST131, multilocus sequence typing, and blaCTX-M gene groups were detected.ResultsAmong 157 E. coli isolates, the rates of nonsusceptibility to ampicillin, amoxycillin + clavulanate, trimethoprim–sulfamethoxazole, and cefazolin were 70, 65.6, 47.1, and 32.5%, respectively. Twenty-nine (18.5%) isolates were nonsusceptible to ciprofloxacin. MDR E. coli accounted for 58 (37%) of all isolates. Thirteen (8.3%) isolates produced extended-spectrum β-lactamase (ESBL). Furthermore, 26 (16.6%) and 13 (8.3%) isolates were O25b and ST131 positive, respectively. Five (38.5%) of the 13 ESBL-producing E. coli belonged to blaCTX-M group 9, among which were CTXM-14 and 4 (80%) were O25b–ST131 positive. Compared with the non-ESBL and ciprofloxacin-susceptible groups, the ESBL and ciprofloxacin-nonsusceptible groups showed significantly higher rates of O25b–ST131 positivity.ConclusionsThe prevalence of the fecal carriage of nonsusceptible E. coli in children was high; among these E. coli, 37% were MDR, 18.5% were nonsusceptible to ciprofloxacin, and 8.3% produced ESBL. O25b–ST131 was the most common ESBL-producing E. coli clonal group present in the feces of children, and the ESBL and ciprofloxacin-nonsusceptible groups showed significantly higher rates of O25b–ST131 positivity.

Multispecies and Clonal Dissemination of OXA-48 Carbapenemase in Enterobacteriaceae From Companion Animals in Germany, 2009—2016

The increasing spread of carbapenemase-producing Enterobacteriaceae (CPE) poses a serious threat to public health. Recent studies suggested animals as a putative source of such bacteria. We investigated 19,025 Escherichia coli, 1607 Klebsiella spp. and 570 Enterobacter spp. isolated from livestock, companion animal, horse, and pet samples between 2009 and 2016 in our routine diagnostic laboratory for reduced susceptibility to carbapenems (CP) by using meropenem-containing media. Actively screened CP non-susceptible strains as well as 367 archived ESBL/AmpC-β-lactamase-producing Enterobacteriaceae were then tested for the presence of CP genes by PCRs. Among 21,569 isolates, OXA-48 could be identified as the sole carbapenemase type in 137 (0.64%) strains. The blaOXA-48 gene was located on an ∼60-kb IncL plasmid and sequence analysis revealed high similarity to reference plasmid pOXA-48a, which has been involved in the global spread of the blaOXA-48 gene in humans for many years. Klebsiella pneumoniae was the predominant OXA-48 producer (n = 86; 6.6% of all K. pneumoniae isolates), followed by E. cloacae (n = 28; 5.0%), Klebsiella oxytoca (n = 1; 0.3%), and E. coli (n = 22, 0.1%). OXA-48 was not found in livestock, but in dogs (120/3182; 3.8%), cats (13/792; 1.6%), guinea pig (1/43; 2.3%), rat (1/23; 4.3%), mouse (1/180; 0.6%), and one rabbit (1/144; 0.7%). Genotyping identified few major clones among the different enterobacteria species, including sequence types ST11 and ST15 for K. pneumoniae, ST1196 for E. coli, and ST506 and ST78 for E. cloacae, most of which were previously involved in the dissemination of multidrug-resistant strains in humans. The majority of OXA-48 isolates (n = 112) originated from a university veterinary clinic (UVC), while animals from further 16 veterinary institutions were positive. Clonal analyses suggested nosocomial events related to different species and STs in two veterinary clinics and horizontal transfer of the pOXA-48-like plasmid between bacterial species and animals. A systematic monitoring is urgently needed to assess the dissemination of CPE not only in livestock but also in companion animals and veterinary clinics.

Effective management of an outbreak with multiresistent Klebsiella pneumoniae in aneurorehabilitation unit

In addition to acute care hospitals, rehabilitation centres are increasingly confronted with multi-resistant pathogens. Long durations of stay and intensive treatments impose special hygienic challenges. We investigated an extended spectrum beta-lactamase-Klebsiella pneumoniae (ESBL-K.pneumoniae) outbreak in aneurorehabilitation centre. We defined confirmed cases as patients who stayed in the centre during the outbreak period and from whom ESBL-K.pneumoniae was isolated with the outbreak sequence type. Probable cases had an epidemiological link to at least one confirmed case but no isolate for typing. Next generation sequencing (NGS) was performed on 53isolates from patients. Environmental sampling was performed. Systematic microbiological screening was implemented and ESBL-K.pneumoniae-positive patients were cohorted in adesignated ward. We identified 30confirmed and 6probable cases. NGS revealed three genetic clusters: Cluster1 - the outbreak cluster - with isolates of 30cases (sequence type ST15), Cluster2 with 7patients (ST405) and Cluster3 with 8patients (ST414). In two patients, the outbreak strain developed further antibiotic resistance, one with colistin resistance and the other carbapenem resistance. The outbreak ceased after strict isolation measures. Epidemiology and NGS results paired with the effectiveness of cohorting suggest that transmission occurred mainly from person to person in this outbreak. There was an apparent association of the probability to acquire ESBL-K.pneumoniae and treatment intensity, whereas infection rate was related to morbidity. The identification of the outbreak clone and additional clusters plus the development of additional antibiotic resistance shows the relevance of NGS and highlights the need for timely and efficient outbreak management.

Comparative analysis of Klebsiella pneumoniae strains isolated in 2012–2016 that differ by antibiotic resistance genes and virulence genes profiles

The antibacterial resistance and virulence genotypes and phenotypes of 148 non-duplicate Klebsiella pneumoniae strains collected from 112 patients in Moscow hospitals in 2012–2016 including isolates from the respiratory system (57%), urine (30%), wounds (5%), cerebrospinal fluid (4%), blood (3%), and rectal swab (1%) were determined. The majority (98%) were multidrug resistant (MDR) strains carrying blaSHV (91%), blaCTX-M (74%), blaTEM (51%), blaOXA (38%), and blaNDM (1%) beta-lactamase genes, class 1 integrons (38%), and the porin protein gene ompK36 (96%). The beta-lactamase genes blaTEM-1, blaSHV-1, blaSHV-11, blaSHV-110, blaSHV-190, blaCTX-M-15, blaCTX-M-3, blaCTX-M-55, blaOXA-48, blaOXA-244, and blaNDM-1 were detected; class 1 integron gene cassette arrays (aadA1), (dfrA7), (dfrA1-orfC), (aadB-aadA1), (dfrA17-aadA5), and (dfrA12-orfF-aadA2) were identified. Twenty-two (15%) of clinical K. pneumoniae strains had hypermucoviscous (HV) phenotype defined as string test positive. The rmpA gene associated with HV phenotype was detected in 24% of strains. The intrapersonal mutation of rmpA gene (deletion of one nucleotide at the polyG tract) was a reason for negative hypermucoviscosity phenotype and low virulence of rmpA-positive K. pneumoniae strain KPB584. Eighteen virulent for mice strains with LD50 ≤ 104 CFU were attributed to sequence types ST23, ST86, ST218, ST65, ST2174, and ST2280 and to capsular types K1, K2, and K57. This study is the first report about hypervirulent K. pneumoniae strain KPB2580-14 of ST23K1 harboring extended-spectrum beta-lactamase CTX-M-15 and carbapenemase OXA-48 genes located on pCTX-M-15-like and pOXA-48-like plasmids correspondingly.

Dissemination of carbapenem-resistant Klebsiella pneumoniae harboring KPC-carrying plasmid pKPC_P16, a pKPC_LK30 variant, in northern Taiwan

The prevalence of carbapenem-resistant Klebsiella pneumoniae (CRKP) was up to 30% between 2014 and 2016 in the study hospital. Of these 77 CRKP isolates, 22 isolates with sequence type ST11 carried the new pKPC_P16 plasmid, a pKPC_LK30 variant, and were widely disseminated between 2014 and 2015 in northern Taiwan.

Molecular epidemiology of carbapenemase-producing Enterobacteriaceae infection/colonisation in a hospital in Madrid

IntroductionA description is presented on the molecular epidemiology of carbapenemase-producing enterobacteriaceae infection in a tertiary hospital. Material and methodsA study was made on all the carbapenemase-producing enterobacteriaceae isolations obtained between February 2015 and March 2016 in the Hospital Universitario 12 de Octubre (Madrid). Phenotypic and molecular methods were used. ResultsA total of 7 bacterial species were identified, with the majority being Klebsiella pneumoniae (K. pneumoniae) (78.9%) and Enterobacter cloacae (E. cloacae) (16.4%). The resistance of K. pneumoniae and E. cloacae for carbapenems was 88.7 and 88.6% for ertapenem, 21.4 and 54.3% for imipenem, and 20.8 and 34.3% for meropenem, respectively. The most frequent carbapenemase type was OXA-48 (91.1%) and VIM (71.4%) in E. cloacae. A total of 9 K. pneumoniae clonal types were identified, including a majority pertaining to the sequence type ST11. In E. cloacae, 16 clonal types were identified. ConclusionsThe current increase in carbapenemase-producing enterobacteriaceae is mainly due to the spread of OXA-48-producing K. pneumoniae.

Draft genome sequence of a tigecycline-resistant KPC-2-producing Klebsiella pneumoniae ST15 clinical isolate from China.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) infections have emerged worldwide as a serious challenge to public health. Tigecycline is regarded as one of the last-resort antimicrobials for CRKP infections. Here we report the draft genome sequence of a clinical tigecycline-resistant KPC-2-producing K. pneumoniae isolate (TRKP1). Whole-genome sequencing of strain TRKP1 was performed using an Illumina Hiseq™ X Ten system. The generated reads were assembled using CLC Genomics Workbench. Functional annotation was performed by the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) server. Bioinformatics analysis was further performed. Klebsiella pneumoniae TRKP1 belongs to sequence type ST15, corresponding to clonal complex CC23. The total genome length of the isolate was 5609823bp, harbouring 19 genes relevant to antibiotic resistance. Antimicrobial susceptibility testing revealed that the isolate is resistant to all antibiotics except colistin. The genome sequence of K. pneumoniae TRKP1 will facilitate the understanding of drug resistance mechanisms and genomic features of clinically isolated tigecycline-resistant CRKP.

Adhesion and invasion to epithelial cells and motility of extended-spectrum β-lactamase-producing Escherichia coli reveal ST131 superiority: a comparative in vitro study of extraintestinal pathogenic E. coli lineages.

Extended-spectrum β-lactamase (ESBL)-producing extraintestinal pathogenic Escherichia coli (ExPEC) sequence type ST131 is pandemic, and it is the major contributor to antibiotic resistance in E. coli. Despite its epidemiological superiority, the physiological reasons that decipher its success remain elusive. We aimed to compare the adhesion, invasion and motility potential of ST131 versus other E. coli lineages. In this in vitro comparative study, 14 ESBL-producing ExPEC community-onset bacteremia isolates were chosen from a reported clinical collection (Karfunkel D, Carmeli Y, Chmelnitsky I, Kotlovsky T, Navon-Venezia S. Eur J Clin Microbiol Infect Dis 2013;32:513-521). Isolates were divided into two groups, ST131 (n=7) and 'non-ST131', sporadic sequence types (STs) (n=7). Virulence and adhesion genes were screened by PCR in all isolates. Virotyping and serotyping were performed for ST131 isolates. Adhesion and invasion to Caco-2 epithelial cells, and motility on semi-solid agar were quantified and compared between the two groups. Fluorescence microscopy using anti-LPS E. coli antibodies was used for visualization and confirmation of adhesion and invasion. ST131 isolates belonged to the O25b:H4-B2 subclone. Two ST131 virotypes were found, A (two blaCTX-M-15 H30-Rx) and C (two blaCTX-M-15 H30-Rx and three blaCTX-M-14 H30 isolates). The average number of adhesion and virulence genes carried by ExPEC ST131 isolates and non-ST131 isolates was 5.3 and 3.7, respectively (P<0.05). Group analysis showed that ST131 surpassed non-ST131 lineages in all three physiological properties: adherence (17.1 vs 13.1 %, P<0.001), invasion (0.4 vs 0.17 %, P<0.01), and swarming motility on all media tested (P<0.05). This study demonstrates ST131 superiority that may explain its improved gut-colonization and dissemination capabilities within the host. These insights are an important step in our understanding of ST131 epidemiological success.

Extensively Drug-Resistant Klebsiella pneumoniae Causing Nosocomial Bloodstream Infections in China: Molecular Investigation of Antibiotic Resistance Determinants, Informing Therapy, and Clinical Outcomes.

The rise in diversity of antimicrobial resistance phenotypes seen in Klebsiella pneumoniae is becoming a serious antibiotic management problem. We sought to investigate the molecular characteristics and clinical implications of extensively drug-resistant (XDR) K. pneumoniae isolated from different nosocomial bloodstream infections (BSIs) patients from July 2013 to November 2015. Even in combination treatment, meropenem did not protect against mortality of BSIs patients (P = 0.015). In contrast, tigecycline in combination with other antimicrobial agents significantly protected against mortality (P = 0.016). Antimicrobial susceptibility tests, molecular detection of antibiotic resistance determinants, conjugation experiments, multilocus sequence typing (MLST), S1-PFGE, Southern blot, SDS-PAGE, immunoblot analysis, and pulsed-field gel electrophoresis (PFGE) were used to characterize these isolates. These XDR K. pneumoniae strains were resistant to conventional antimicrobials except tigecycline and polymyxin B and co-harbored diverse resistance determinants. rmtB, blaKPC−2 as well as blaCTX−M−9 were located on a transferable plasmid of ~54.2 kb and the most predominant replicon type was IncF. 23 of the 35 isolates belonging the predominant clone were found to incorporate the globally-disseminated sequence type ST11, but others including a unique, previously undiscovered lineage ST2281 (allelic profile: 4-1-1-22-7-4-35) were also found and characterized. The porins OmpK35 and OmpK36 were deficient in two carbapenemase-negative carbapenem-resistant strains, suggesting decreased drug uptake as a mechanism for carbapenem resistance. This study highlights the importance of tracking hospital acquired infections, monitoring modes of antibiotic resistance to improve health outcomes of BSIs patients and to highlight the problems of XDR K. pneumoniae dissemination in healthcare settings.

Genome sequencing and comparative analysis of an NDM-1-producing Klebsiella pneumoniae ST15 isolated from a refugee patient

The escalating problem of antibiotic resistance, specifically cabarpenemase and extended-spectrum β-lacatamase (ESBL) producing K. pneumoniae strains, is directly correlated with increased patient morbidity and mortality and prolonged hospitalization and costs. In this study, a comprehensive genomic analysis encompassing the resistomics, virulence repertoire and mobile genetic elements of an NDM-1 positive ESBL-producing K. pneumoniae EA-MEH ST15 isolated from a urine sample collected from a Syrian refugee was conducted. Illumina paired-end libraries were prepared and sequenced resulting in 892,300 high-quality reads. The initial assembly produced 329 contigs with a combined 5,954,825 bp and a 56.5% G+C content. Resistome analysis revealed the presence of several β-lactamases including NDM-1, SHV-28, CTX-M-15 and OXA-1 in addition to 18 other genes encoding for resistance, among which are aph(3′)-Ia, aac(6′)Ib-cr, armA, strB, strA and aadA2 genes. Additionally, five plasmids IncFIB(Mar), IncHI1B, IncFIB(pKPHS1), IncFIB(K) and IncFII(K) and four integrated phages were detected. In silico MLST analysis revealed that the isolate was of sequence type ST15. To our knowledge this is the first in-depth genomic analysis of a NDM-1 positive K. pneumoniae ST15 in Lebanon associated with the recent population migration. The potential dissemination of such MDR strains is an important public health concern.

Epidemiología molecular de las infecciones/colonizaciones por enterobacterias productoras de carbapenemasas en un hospital de Madrid

IntroductionA description is presented on the molecular epidemiology of carbapenemase-producing enterobacteriaceae infection in a tertiary hospital. Material and methodsA study was made on all the carbapenemase-producing enterobacteriaceae isolations obtained between February 2015 and March 2016 in the Hospital Universitario 12 de Octubre (Madrid). Phenotypic and molecular methods were used. ResultsA total of 7 bacterial species were identified, with the majority being Klebsiella pneumoniae (K. pneumoniae) (78.9%) and Enterobacter cloacae (E. cloacae) (16.4%). The resistance of K. pneumoniae and E. cloacae for carbapenems was 88.7 and 88.6% for ertapenem, 21.4 and 54.3% for imipenem, and 20.8 and 34.3% for meropenem, respectively. The most frequent carbapenemase type was OXA-48 (91.1%) and VIM (71.4%) in E. cloacae. A total of 9K. pneumoniae clonal types were identified, including a majority pertaining to the sequence type ST11. In E. cloacae, 16 clonal types were identified. ConclusionsThe current increase in carbapenemase-producing enterobacteriaceae is mainly due to the spread of OXA-48-producing K. pneumoniae.

National survey of colistin resistance among carbapenemase-producing Enterobacteriaceae and outbreak caused by colistin-resistant OXA-48-producing Klebsiella pneumoniae, France, 2014.

From January 2014 to December 2014, 972 consecutive non-replicate carbapenemase-producing Enterobacteriaceae isolates from colonised or infected patients were collected at the Associated French National Reference Centre as part of the French national survey on antimicrobial resistance. It included 577 Klebsiella spp. (59%), 236 Escherichia coli (24%), 108 Enterobacter spp. (11%), 50 Citrobacter spp. (5%), and a single Salmonella spp. isolate (0.1%). Of 561 K. pneumoniae isolates, 35 were found to be resistant to colistin (6.2%). PFGE analysis revealed a clonal outbreak involving 15 K. pneumoniae isolates belonging to sequence type ST11, recovered in a single hospital in the Picardie region in northern France. Those clonally related isolates showed variable levels of resistance to colistin, ranging from 4 to 64 mg/L. They harboured the blaOXA-48 carbapenemase gene and the blaCTX-M-15 extended-spectrum beta-lactamase gene. Among the 91 Enterobacter cloacae isolates, seven were resistant to colistin and produced different types of carbapenemases. Surprisingly, none of the E. coli and Citrobacter spp. isolates showed resistance to colistin. This national survey including carbapenemase-producing isolates recovered in 2014 reported a high rate of colistin resistance in K. pneumoniae and E. cloacae (6.2% and 7.7%, respectively) in France.

High prevalence of extended-spectrum ß-lactamase producing enterobacteriaceae among clinical isolates in Burkina Faso.

BackgroundNothing is known about the epidemiology and resistance mechanisms of extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBL-PE) in Burkina Faso. The objective of this study was to determine ESBL-PE prevalence and to characterize ESBL genes in Burkina Faso.MethodsDuring 2 months (June-July 2014), 1602 clinical samples were sent for bacteriologic investigations to the microbiology laboratories of the tree main hospitals of Burkina Faso. Isolates were identified by mass spectrometry using a matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) BioTyper. Antibiotic susceptibility was tested using the disk diffusion method on Müller-Hinton agar. The different ESBL genes in potential ESBL-producing isolates were detected by PCR and double stranded DNA sequencing. Escherichia coli phylogenetic groups were determined using a PCR-based method.ResultsESBL-PE frequency was 58 % (179 strains among the 308 Enterobacteriaceae isolates identified in the collected samples; 45 % in outpatients and 70 % in hospitalized patients). The CTX-M-1 group was dominant (94 %, CTX-M-15 enzyme), followed by the CTX-M-9 group (4 %). ESBL producers were more often found in E. coli (67.5 %) and Klebsiella pneumoniae (26 %) isolates. E. coli isolates (n = 202; 60 % of all Enterobacteriaceae samples) were distributed in eight phylogenetic groups (A = 49, B1 = 15, B2 = 43, C = 22, Clade I = 7, D = 37, F = 13 and 16 unknown); 22 strains belonged to the sequence type ST131. No association between a specific strain and ESBL production was detected.ConclusionsThis report shows the alarming spread of ESBL genes in Burkina Faso. Public health efforts should focus on education (population and healthcare professionals), surveillance and promotion of correct and restricted antibiotic use to limit their dissemination.

Countrywide dissemination of a DHA-1-type plasmid-mediated AmpC β-lactamase-producing Klebsiella pneumoniae ST11 international high-risk clone in Hungary, 2009–2013

The first plasmid-mediated AmpC β-lactamase-producing Klebsiella pneumoniae (pAmpC KP) isolate was detected in December 2009 in Hungary. Hungarian microbiological laboratories were asked to send all KP strains showing cefoxitin resistance and decreased susceptibility or resistance to any third-generation cephalosporins to the Reference Laboratories at the National Center for Epidemiology. Investigation was conducted in order to outline spatio-temporal distribution and genetic characterization of pAmpC-KP isolates in Hungary. Between December 2009 and December 2013, 312 consecutive KP clinical isolates were confirmed as producing pAmpCs. All isolates showed resistance to third-generation cephalosporins, aminoglycosides and fluoroquinolones, and 77 % were non-susceptible to at least one carbapenem. Analysis of β-lactamase genes showed blaDHA-1 in all and additionally blaCTX-M-15 in 90 % of isolates. PFGE typing revealed 12 pulsotypes; of these, KP053 (262/312) and KP070 (38/312) belonged to sequence type ST11 and comprised 96 % of the isolates. The blaDHA-1 and blaCTX-M-15 co-producing KP053/ST11 clone affected 234 patients and spread to 55 healthcare centres across Hungary during the study period. Three KP053 isolates were also resistant to colistin. In two of these, the mgrB gene was truncated by IS10R, while in the third isolate, insertional inactivation of mgrB by ISKPn14 was identified. Hungary is the first European country showing endemic spread of blaDHA-1 facilitated by the international high-risk clone ST11. The rapid countrywide spread of this multidrug-resistant clone seriously endangers Hungarian healthcare facilities and warrants strengthening of infection control practices and prudent use of carbapenems and colistin.