Ciprofloxacin-resistant,CTX-M-15-producing Escherichia coli ST131 clone in extraintestinal infections in Italy

Quinolone and ß-lactam resistance mechanisms and clonal relationships were characterized among Escherichia coli isolates resistant to ciprofloxacin and extended-spectrum cephalosporins associated with human extra-intestinal infections in Rome. The E. coli. ST131 clone was found to be prevalent. This clone invariably carried a specific pattern of substitutions in the topoisomerase genes and all isolates but one produced CTX-M-15. One ST131 isolate produced SHV-12. The new ST131 variant described here is of particular concern because it combines fluoroquinolone resistance and chromosomally encoded CTX-M-15.     

NDM-1, OXA-48 and OXA-181 carbapenemase-producing Enterobacteriaceae in Sultanate of Oman

Twenty-two carbapenem-resistant enterobacterial isolates were recovered from patients hospitalized between October 2010 and March 2011 at the Royal Hospital of Muscat, Sultanate of Oman. Eleven NDM-1, five OXA-48 and one NDM-1 plus OXA-181 producers of diverse ST types were recovered from clinical samples. All carbapenemase genes were located on self-conjugative plasmids and were nearly always associated with other resistance determinants, including extended-spectrum β-lactamases and the ArmA methylase encoding resistance to aminoglycosides. This work highlights the dissemination of NDM-1 and OXA-48-type producers in the Middle East.     

Co-existence of NDM-1 and OXA-48 genes in Carbapenem Resistant Klebsiella pneumoniae clinical isolates in Kafrelsheikh,Egypt

BackgroundThe noteworthy spread of carbapenem-resistant K. pneumoniae (CR-KP) isolates represents a significant safety threat.ObjectiveDetermination of the carbapenemase genes incidence among CR-KP clinical isolates in Kafrelsheikh, Egypt.MethodsA total of 230 K. pneumoniae isolates were recovered from four hospitals in Kafrelsheikh, Egypt. Susceptibility testing was conducted using Kirby-Bauer method and automated-Vitek2 system. CR-KP isolates were tested using modified Hodge test (MHT) and combined disk synergy test. PCR and DNA sequencing were conducted for CR-KP isolates to recognize the included carbapenemase-genes.ResultsOut of 230 K. pneumoniae isolates, 50 isolates presented resistance to carbapenem (meropenem). All 50 CR-KP isolates were multidrug-resistant (MDR). Genes like blaNDM-1 and blaOXA-48 were the only detected genes among CR-KP with an incidence of 70.0% and 52.0%, respectively. Up to 74.0% of the tested isolates carried at least one of the two recorded genes, among them 48.0% co-harbored both blaNDM-1 and blaOXA-48 genes. The accession-numbers of sequenced blaNDM-1 and blaOXA-48 genes were {"type":"entrez-nucleotide","attrs":{"text":"MG594615","term_id":"1344457373"}}MG594615 and {"type":"entrez-nucleotide","attrs":{"text":"MG594616","term_id":"1344457375"}}MG594616, respectively.ConclusionThis study reported a high incidence of MDR profile with the emergence of blaNDM-1 and blaOXA-48 genes co-existence in CR-KP isolates in Kafrelsheikh, Egypt. Hence, more restrictions should be applied against the spread of such serious pathogens.     

Intrapatient emergence of OXA-247: a novel carbapenemase found in a patient previously infected with OXA-163-producing Klebsiella pneumoniae

Two genetically related Klebsiella pneumoniae strains carrying OXA-type carbapenemases were isolated from a single patient 1 month apart. Kpn163 harboured OXA-163 and Kpn247 a new variant named OXA-247 that showed susceptibility to carbapenems and expanded-spectrum cephalosporins similar to OXA-48. Our epidemiological, biochemical and molecular results suggest the intrapatient emergence of bla_OXA-247 from bla_OXA-163.     

Emergence of Klebsiella pneumoniae co-producing NDM-type and OXA-181 carbapenemases

The emergence of carbapenemase-producing Enterobacteriaceae is a rapidly evolving threat worldwide. Here, we report the molecular characterization of two Klebsiella pneumoniae isolates carrying both bla_OXA-181 and bla_NDM-1 or bla_NDM-5 isolated from epidemiologically unrelated patients in Singapore. The bla_OXA-181 genes were found existing in different genetic environments.     

Molecular epidemiology of OXA-48-producing Klebsiella pneumoniae in France

We characterized 53 OXA-48-producing Klebsiella pneumoniae (OXA-48-Kp) isolated between 2011 and 2013 in 21 French hospitals. All the isolates were genotyped using MLST and PFGE and the population structure of the species was determined by a nucleotide-based analysis of the entire K. pneumoniae MLST database. Most of the OXA-48-Kp isolates also produced CTX-M-15 and remained susceptible to imipenem and meropenem. The isolates were distributed into 20 STs, of which five were dominant (ST15, ST101, ST147, ST395 and ST405). All the OXA-48-Kp clustered in the major clade of K. pneumoniae KpI.     

Concurrent emergence of multidrug resistance and heat resistance by CTX-M-15-encoding conjugative plasmids in Klebsiella pneumoniae

A plasmid-encoded ClpK protein was recently identified as a predictor of a heat-resistant phenotype in the opportunistic pathogen Klebsiella pneumoniae. This study was undertaken to evaluate the presence of the clpK gene in extended-spectrum β-lactamase (ESBL)-producing K. pneumoniae and to assess the probable co-transfer of multi-resistance with the heat resistance phenotype. A Danish collection of 80 ESBL-producing K. pneumoniae bloodstream infection isolates was screened for clpK by colony hybridization. Nineteen isolates (24%) were positive for clpK; some of them representing major clones identified in Denmark. Among these, nine isolates belonged to a single K. pneumoniae CTX-M-15 clone with sequence type (ST)16 exhibiting a heat-resistant phenotype. This clone has a multi-hospital occurrence and has also been detected outside Denmark. Horizontal co-transfer of multiple antibiotic resistances, including the CTX-M-15 resistance determinant, and the heat resistance phenotype was observed. Thus, the clpK gene is harbored by different ESBL-producing K. pneumoniae isolates including a clone of ST16 internationally spread. The co-localization of clpK on transferable ESBL-encoding plasmids allowing co-dissemination of multiple drug resistance with bacterial heat resistance is a highly interesting phenomenon that may further complicate the prevention of spreading of certain successful clones of multi-resistant K. pneumoniae.     

Diversity identification of KPC- producing Klebsiella pneumoniae using multilocus variable number tandem repeat analysis

PurposeThe current study aimed to determine blaKPC, blaGES, blaVIM, blaNDM, blaOXA-23, and blaOXA-48 genes in clinical strains of Klebsiella pneumoniae isolated in Tehran, Iran to assess genetic diversity using MLVA as a typing method.MethodsA total of 181 K. pneumoniae isolates were obtained from various clinical samples. CLSI 2018 (clinical and laboratory standards institute) guidelines were used to determine antibiotic susceptibility and the Modified Hodge Test (MHT).To detect blaKPC, blaGES, blaVIM, blaNDM, blaOXA-23, and blaOXA-48, the polymerase chain reaction (PCR) method was used. The MLVA method was used to type K. pneumoniae isolates by using PCR for 8 Variable Number Tandem Repeats (VNTRs).ResultsImipenem was the most effective antibiotic against K. pneumoniae, with 36.5% susceptibility. 100 (55.24%) of the isolates tested positive for KPC, and 30 (30%) tested positive for six genes. Thirty MLVA genotypes were distinguished, and an examination of diversity indexes (DIs) for eight loci revealed that seven different alleles were the most polymorphic, with the highest DI of 0.780.ConclusionsThe present study showed that MLVA could be helpful for typing clinical strains of K. pneumoniae. Our K. pneumoniae isolates are thought to be derived from a small number of clones that have undergone minor genetic changes over time. The results also showed that this method had great potential to differentiate those strains with high phenotypic similarity. The current study has revealed some intriguing facts about K. pneumoniae genetic relatedness in Tehran, Iran.     

Presence and characterization of blaNDM-1-positive carbapenemase-producing Klebsiella pneumoniae from outpatients in Thailand

BackgroundPresently, community-associated carbapenemase-producing Enterobacterales (CPE) remains largely unknown and require public attention. This study aimed to investigate the presence of CPE from outpatients in Thailand.MethodsNon-duplicate stool (n = 886) and urine (n = 289) samples were collected from outpatients with diarrhea and urinary tract infection, respectively. Demographic data and characteristics of patients were collected. Isolation of CPE was performed by plating enrichment culture on agar supplemented with meropenem. Carbapenemase genes were screened by PCR and sequencing. CPE isolates were phenotypically and genotypically characterized.ResultsFifteen samples (1.3%, 14 stool and 1 urine) yielded bla_NDM-1-positive carbapenemase-producing Klebsiella pneumoniae (CPKP). Additional resistance to colistin and tigecycline was observed in 53.3% and 46.7% of isolates, respectively. Age >60 years was identified as a risk factor for patients with CPKP (P < 0.001, adjusted odds ratio = 11.500, 95% confidence interval = 3.223–41.034). Pulsed field gel electrophoresis revealed genetic diversity of CPKP isolates; however, clonal spread has been observed. ST70 (n = 4) was common, followed by ST147 (n = 3). bla_NDM-1 from all isolates were transferable and mainly resided on IncA/C plasmid (80%). All bla_NDM-1 plasmids remained stable in bacterial host for at least 10 days in antibiotic-free environments, regardless of replicon types.ConclusionThis study demonstrates that the prevalence of CPE among outpatients in Thailand remains low and the spread of bla_NDM-1-positive CPKP may be driven by IncA/C plasmid. Our results emphasize the need for a large-scale surveillance study to limit further spread of CPE in community.     

The methodology of surveillance for antimicrobial resistance and healthcare-associated infections in Europe (SUSPIRE): a systematic review of publicly available information

ObjectivesSurveillance is a key component of any control strategy for healthcare-associated infections (HAIs) and antimicrobial resistance (AMR), and public availability of methodologic aspects is crucial for the interpretation of the data. We sought to systematically review publicly available information for HAIs and/or AMR surveillance systems organized by public institutions or scientific societies in European countries.MethodsA systematic review of scientific and grey literature following the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines was performed. Information on HAIs and/or AMR surveillance systems published until 31 October 2016 were included.ResultsA total of 112 surveillance systems were detected; 56 from 20 countries were finally included. Most exclusions were due to lack of publicly available information. Regarding AMR, the most frequent indicator was the proportion of resistant isolates (27 of 34 providing information, 79.42%); only 18 (52.9%) included incidence rates; the data were only laboratory based in 33 (78.5%) of the 42 providing this information. Regarding HAIs in intensive care units, all 22 of the systems providing data included central line–associated bloodstream infections, and 19 (86.3%) included ventilator-associated pneumonia and catheter-associated urinary tract infections; incidence density was the most frequent indicator. Regarding surgical site infections, the most frequent procedures included were hip prosthesis, colon surgery and caesarean section (21/22, 95.5%).ConclusionsPublicly available information about the methods and indicators of the surveillance system is frequently lacking. Despite the efforts of European Centre for Disease Control and Prevention (ECDC) and other organizations, wide heterogeneity in procedures and indicators still exists.     

Characterisation of Virulence Genes Associated with Pathogenicity in Klebsiella pneumoniae

Purpose: This study was undertaken to characterise the virulence factors in clinical strains of Klebsiella pneumoniae and analyse their association with various infections caused and also to determine the association between virulence factors and antimicrobial resistance profile. Materials and Methods: A total number of 370 clinically significant, non-duplicate isolates of K. pneumoniae isolated from both hospitalised patients and patients attending clinics were included in this study. Polymerase chain reaction (PCR) was carried out for the detection of various virulence genes such as mucoviscosity-associated gene A (magA), gene associated with allantoin metabolism (allS), Klebsiella ferric iron uptake(Kfu), capsule-associated gene A (K2A), regulator of mucoid phenotype A (rmpA), enterobactin (entB), yersiniabactin (YbtS), aerobactin, Fimbrial adhesin (FimH) and uridine-diphosphate galacturonate 4-epimerase (uge). Antimicrobial susceptibility testing and PCR-based detection of beta-lactamase-encoding genes such as extended-spectrum beta-lactamases, AmpCs and carbapenemases were performed. Univariate analysis was done to find the association between virulence genes and mortality. Results: The siderophore, entB, was present in most (90.5%) of the isolates. Of the 370 isolates, 345 carried multiple virulence genes; 15 harboured single virulence genes and 10 did not harbour any of the studied virulence genes. The most common combination of occurrence was entB and FimH. A mortality rate of 12.75% (38/298) was observed among hospitalised patients. None of the virulence genes had any significant association with mortality. Conclusion: Pathogenic K. pneumoniae can harbour single to multiple virulence genes. Invasive infection with even a single virulence gene-harbouring K. pneumoniae can lead to poor outcomes. Both multidrug-resistant (MDR) and non-MDR K. pneumoniae can harbour a variety of virulence genes. None of the virulence genes have a significant association with mortality.     

The first major extended-spectrum beta-lactamase outbreak in Scandinavia was caused by clonal spread of a multiresistant Klebsiella pneumoniae producing CTX-M-15

Between May and December 2005, 64 multidrug-resistant isolates of Klebsiella pneumoniae were detected from patients admitted to Uppsala University Hospital. This represented a dramatic increase in ESBL-producing K. pneumoniae compared to previous years. To investigate the epidemiology and to characterize the resistance mechanisms of the isolates, a study was initiated. Antibiotic susceptibility was determined by means of the Etest and the disc diffusion method. Extended-spectrum beta-lactamase (ESBL) production was identified by clavulanic acid synergy test and confirmed with PCR amplification followed by DNA sequencing. DNA profiles of the isolates were examined with pulsed-field gel electrophoresis (PFGE). All isolates were resistant or exhibited reduced susceptibility to cefadroxil, cefuroxime, cefotaxime, ceftazidime, aztreonam, piperacillin/tazobactam, ciprofloxacin, tobramycin, and trimethoprim-sulfamethoxazole. They produced ESBL of the CTX-M-15 type, and the involvement of a single K. pneumoniae clone was shown. This is the first major clonal outbreak of multiresistant ESBL-producing K. pneumoniae in Scandinavia. The outbreak demonstrates the epidemic potential of enterobacteria containing ESBLs of the CTX-M type, even in a country with a relatively low selective pressure and a low prevalence of multiresistant bacteria.     

Multiclonal epidemic of Klebsiella pneumoniae isolates producing DHA-1 in a Spanish hospital

Between June 2007 and January 2008, 26 Klebsiella pneumoniae isolates carrying bla_DHA-1 on an IncL/M plasmid were obtained from clinical samples at Granollers Hospital, Barcelona, Spain. Three of the isolates also carried a bla_CTX-M-15 gene. The 26 isolates showed 11 pulsed-field gel electrophoresis (PFGE) patterns. Multilocus sequence typing showed that PFGE patterns A, B and C belonged to sequence type (ST)17, D to ST13, E to ST427, F and G to ST416, H to ST37, I to ST440, J to ST326, and K to ST428. Results demonstrated the effectiveness of the infection control programme in place at the centre. This study reports the first characterization of STs for bla_DHA-1-producing K. pneumoniae isolates.     

Inguinal skin colonization with multidrug-resistant bacteria among residents of elderly care facilities: Frequency,persistence, molecular analysis and clinical impact

Frequency, persistence and molecular characteristics of multidrug resistant bacteria colonizing inhabitants of long term care facilities are topics of current concern. We performed a point-prevalence survey of 402 residents in 7 elderly care facilities in Berlin, Germany. Inguinal swabs were analyzed for the presence of methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococci (VRE), and multidrug-resistant gram-negative bacteria. Three and six months following the initial investigation, all colonized residents were sampled again and the occurrence of intercurrent infections, hospital admissions and use of antimicrobials were registered. Genetic relatedness of the bacteria was investigated using multi-locus sequence typing (MLST), spa-typing and SmaI/XbaI-macrorestriction analysis. 33 (8.2%) residents were skin-colonized with multidrug-resistant bacteria. MRSA were found in 19 (4.7%) and ESBL-producing Enterobacteriaceae in 16 residents (3.98%). Independent risk factors for colonization with multidrug-resistant bacteria were a high level of care and the presence of chronic wounds. A large proportion of the observed bacteria persisted up to six months and showed a high degree of inter-individual diversity. Outcome analysis revealed that infections tend to occur slightly more often in residents colonized by multiresistant pathogens. We assume that a perceptible population of residents in nursing homes is at risk for individual colonization with multidrug-resistant bacteria as well as healthcare associated infections.     

Clonal dissemination of multilocus sequence type ST15 KPC‐2‐producing Klebsiella pneumoniae in Bulgaria

A total of 36 consecutive clinical and two fecal‐screening carbapenem‐resistant Klebsiella pneumoniae isolates from two Bulgarian university hospitals (Varna and Pleven) were investigated. Susceptibility testing, conjugation experiments, and plasmid replicon typing were carried out. Beta‐lactamases were characterized by isoelectric focusing, PCR, and sequencing. Clonal relatedness was investigated by RAPD and multilocus sequence typing (MLST). Most of the isolates demonstrated multidrug resistance profile. Amikacin and tigecycline retained good activity with susceptibility rates of 95 and 87%, respectively. The resistance rate to colistin was 63%. Six RAPD‐ and MLST‐types were identified: the dominating MLST‐type was ST15 (27 isolates), followed by ST76 (six isolates), and ST1350 (two isolates). ST101, ST258, and ST151 were detected once. All except one of the K. pneumoniae produced KPC‐2, mostly in combination with CTX‐M‐15, while for one isolate (ST101) the enzymes OXA‐48 and CTX‐M‐14 were found. All KPC‐2‐producing transconjugants revealed the presence of IncFII plasmid. The OXA‐48‐ and CTX‐M‐14‐producing isolate showed the presence of L/M replicon type. The dissemination of KPC‐2‐producing K.pneumoniae in Bulgaria is mainly due to the sustained spread of successful ST15 clone and to a lesser extent of ST76 clone. This is the first report of OXA‐48 producing ST101 K. pneumoniae in Bulgaria.     

Structure and gene cluster of the capsular polysaccharide of multidrug resistant carbapenemase OXA-48-producing Klebsiella pneumoniae strain KPB536 of the genetic line ST147

The Gram-negative opportunistic pathogen Klebsiella pneumoniae is a significant cause of community-acquired and healthcare-associated infections for which multidrug resistance is a concern worldwide. A major virulence determinant of K. pneumoniae is a polysaccharide capsule (CPS) which forms a barrier around the bacterial cell wall, providing protection from environmental pressures and immune responses of eukaryotic organisms. More than 70 chemical capsule structures of serologically typeable K. pneumoniae strains are known. However, there are little data on the CPS structure and cps gene cluster organization of clinical multidrug resistant K. pneumoniae strains.Our investigation of multidrug resistant carbapenemase OXA-48-producing K. pneumoniae strain KPB536 identified a capsular type that was structurally similar to K. pneumoniae K10 but different from any K. pneumoniae CPS reported so far. The content and organization of the cps gene cluster in K. pneumoniae KPB536 also was determined. The catalytic functions of glycosyltransferases coded by the cps_KPB536 gene cluster were assigned by comparison with those responsible for the synthesis of glycoside linkages in the CPSs of K. pneumoniae types K10 and K61.     

Further increase of vancomycin-resistant Enterococcus faecium, amikacin- and fluoroquinolone-resistant Klebsiella pneumoniae, and imipenem-resistant Acinetobacter spp. in Korea: 2003 KONSAR surveillance

Monitoring temporal trends of antimicrobial resistance can provide useful information for the empirical selection of antimicrobial agents to treat infected patients and for the control of nosocomial infections. In this study, we analyzed antimicrobial resistance of clinically relevant bacteria in 2003 at Korean hospitals and at a commercial laboratory. The following organism-antimicrobial agent resistance combinations were very prevalent: oxacillin-resistant Staphylococcus aureus (68%), expanded-spectrum cephalosporin-resistant Klebsiella pneumoniae (25%), and fluoroquinolone-resistant Escherichia coli (33%), Acinetobacter spp. (58%), and Pseudomonas aeruginosa (40%). Moreover, gradual increases in vancomycin-resistant Enterococcus faecium (20%), cefoxitin-resistant E. coli (10%) and K. pneumoniae (23%), and imipenem-resistant P. aeruginosa (20%) and Acinetobacter spp. (13%) were also observed. The resistance rates of Acinetobacter spp. to most antimicrobial agents at hospitals and at the commercial laboratory were similar. Among the Acinetobacter spp. isolated at a tertiary-care hospital, 46.2% were multidrug-resistant to 9-12 of 13 antimicrobial agents, and 18.3% were panresistant. The exclusion of duplicate isolates at a tertiary-care hospital significantly lowered the proportion of oxacillin-resistant S. aureus, vancomycin-resistant E. faecium, and fluoroquinolone-resistant E. coli.     

Antimicrobial susceptibility to parenteral and oral agents in a largely polyclonal collection of CTX-M-14 and CTX-M-15-producing Escherichia coli and Klebsiella pneumoniae

Activity of oral and parenteral antimicrobials against consecutively isolated extended-spectrum β-lactamase (ESBL)-producing Escherichia coli (n = 149) and Klebsiella pneumoniae (n = 20) was determined, and susceptibility test methods were compared for parenteral β-lactams. Polymerase chain reaction (PCR) targeting bla(CTX-M), bla(SHV) and bla(TEM), and DNA sequencing and epidemiological typing with pulsed-field gel electrophoresis were performed. PCR targeting pabB was screened for E. coli O25b-ST131. Minimum inhibitory concentrations (MICs) were determined using Etest and broth microdilution. Disc diffusion was performed according to European Committee on Antimicrobial Susceptibility Testing (EUCAST). Dominating genotypes were bla(CTX-M-15) (75%) and bla(CTX-M-14) (23%). Four E. coli clusters (7-18 isolates) were found. Forty-two per cent of E. coli belonged to O25b-ST131. Ciprofloxacin resistance was 72%, trimethoprim resistance was 70%. Among E. coli, resistance to mecillinam (13%), nitrofurantoin (7%) and fosfomycin (3%) was low, although resistance was high in K. pneumoniae (25%, 60%, 85%). Susceptibility to ertapenem was 99%, piperacillin-tazobactam 91%, tigecycline 96% and temocillin 76%. Susceptibility rates obtained with broth microdilution and Etest were in agreement for cefotaxime (2 vs 1%) and ceftazidime (9 vs 11%), but not for piperacillin-tazobactam (59 vs 91%). With disc diffusion major errors occurred with piperacillin-tazobactam (18/169). Several therapeutic alternatives exist for ESBL-producing E. coli, but few exist for K. pneumoniae. Disc diffusion and Etest can accurately predict susceptibility to cefotaxime and ceftazidime, but not to piperacillin-tazobactam with the present breakpoints.