Plasmid-mediated AmpC Β-lactamase Research Articles

Antimicrobial Resistance Profiles of Adherent Invasive Escherichia coli Show Increased Resistance to β-Lactams.

The adherent invasive Escherichia coli (AIEC) pathotype has been associated with the aetiology of Crohn’s disease (CD). Scarce reports have shown the antimicrobial resistance (AMR) profiles of AIEC. Despite antibiotics not being recommended to treat CD, antimicrobial therapy could be useful in stratified patients, such as AIEC carriers. We examined the antimicrobial resistance profiles of AIEC strains to identify which therapies could be effective or confer a risk for such patients. Phenotypic resistance to 30 antimicrobials was tested according to CLSI standards. AIEC (n = 22) and non-pathogenic E. coli (non-AIEC) strains (n = 37) isolated from the gut mucosa of 31 CD patients and 18 controls were studied. De novo genome sequencing was carried out for 39 of the 59 strains, and AMR genes were searched using the DeepARG database in these genomes and 33 additional AIEC publicly available genomes. The strains isolated from CD and controls showed similar phenotypic AMR profiles. The genomic analysis did not reveal an increased prevalence of AMR genes. However, AIEC strains were more frequently resistant to β-lactams than non-AIEC strains (11 AIEC (50%) and 5 non-AIEC (22%) strains were resistant to at least one β-lactam; p < 0.042). Two AIEC strains were resistant to expanded-spectrum cephalosporins. One strain carried a plasmid-mediated AmpC β-lactamase (CMY-69), and the other presented mutations in the promotor of the intrinsic chromosomal AmpC related to the hyperproduction of this enzyme. The rest of the strains were resistant to β-lactams not including expanded-spectrum cephalosporins. The majority carried TEM-related β-lactamases. Genomic analysis including external AIEC revealed that the gene sul1 encoding for sulphonamide resistance was more frequent in AIEC strains than non-AIEC strains (34.6% vs. 9.5%, p = 0.030). AMR in AIEC is a matter of concern regarding the putative implication of the pathotype in CD. The high proportion of AIEC resistant to β-lactams warrants caution about the risk there may be in the use of these antimicrobials in AIEC-colonized CD patients.

Evaluation of the Association Between Gastric Acid Suppression and Risk of Intestinal Colonization With Multidrug-Resistant Microorganisms

Acid suppressants inhibit gastric acid secretion and disrupt the intestinal microbiome. Whether acid suppression increases the risk of colonization with multidrug-resistant microorganisms (MDROs) is unclear. To systematically examine the association of use of acid suppressants with the risk of colonization with MDROs and to perform a meta-analysis of current evidence. PubMed, Embase, the Web of Science Core Collection, and the Cochrane Central Register of Controlled Trials were searched from database inception through July 8, 2019. Study selection was performed independently by 2 authors (R.P.J.W. and C.M.J.E.V.-G.) on the basis of predefined selection criteria; conflicts were resolved by consensus or by an adjudicator (K.v.D.). Human observational studies (case control, cohort, and cross-sectional) and clinical trial designs were selected if they quantified the risk of MDRO colonization in users of acid suppressants in comparison with nonusers. The Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) and Meta-analysis of Observational Studies in Epidemiology (MOOSE) recommendations were followed. Data were extracted independently by the same 2 authors, and adjudication was conducted when necessary. Risk of bias was assessed according to a modified Newcastle-Ottawa Scale. Pooled odds ratios (ORs) were estimated using random-effects models; heterogeneity was evaluated using the I2 method. The primary outcome measure was intestinal colonization with MDROs of the Enterobacterales order (producing extended-spectrum β-lactamases, carbapenemases, or plasmid-mediated AmpC β-lactamases), vancomycin-resistant enterococci, methicillin-resistant or vancomycin-resistant Staphylococcus aureus, or multidrug-resistant Pseudomonas or Acinetobacter species. A total of 26 observational studies including 29 382 patients (11 439 [38.9%] acid suppressant users) met the selection criteria. Primary meta-analysis of 12 studies including 22 305 patients that provided adjusted ORs showed that acid suppression increased the odds of intestinal carriage of MDROs of the Enterobacterales order and of vancomycin-resistant enterococci by roughly 75% (OR = 1.74; 95% CI, 1.40-2.16; I2 = 68%). The odds were concordant with the secondary pooled analysis of all 26 studies (OR = 1.70; 95% CI, 1.44-1.99; I2 = 54%). Heterogeneity was partially explained by variations in study setting and the type of acid suppression. Acid suppression is associated with increased odds of MDRO colonization. Notwithstanding the limitations of observational studies, the association is plausible and is strengthened by controlling for confounders. In view of the global increase in antimicrobial resistance, stewardship to reduce unnecessary use of acid suppressants may help to prevent MDRO colonization.

Extended-spectrum beta-lactamases and plasmid diversity in urinary isolates of Escherichia coli in Croatia: a nation-wide, multicentric, retrospective study.

In recent years, a dramatic increase in the prevalence of Escherichia coli strains producing extended-spectrum β-lactamases (ESBLs) has been observed-both in the community and in healthcare settings. This multicentric study aimed to characterize ESBLs produced by E. coli isolates causing hospital-onset and community urinary tract infections, as well as to compare their antimicrobial sensitivity patterns, β-lactamase content and plasmid types. Phenotypic tests for the detection of ESBLs and plasmid-mediated AmpC β-lactamases were initially pursued, followed by molecular detection of resistance genes, plasmid characterization, genotyping with pulsed-field gel electrophoresis and whole genome sequencing (WGS). The isolates exhibited high level of resistance to expanded-spectrum cephalosporins (ESC) and carried CTX-M (cefotaximase-Munich) or TEM (Temoniera) β-lactamases. All six representative isolates subjected to WGS belonged to the widespread clone ST131. In conclusion, our study demonstrated dissemination of group 1 CTX-M positive E. coli in different geographic regions of Croatia, but also different components of the health care systems (hospitals, nursing homes and the community) and confirmed the switch from SHV-2 (suphydril variant) and SHV-5 ESBLs to the nation-wide predominance of group 1 CTX-M β-lactamases. Different plasmids were shown to be associated with the dissemination of blaCTX-M genes in different geographic regions of Croatia.

Occurrence of ESBL-Producing Escherichia coli ST131, Including the H30-Rx and C1-M27 Subclones, Among Urban Seagulls from the United Kingdom.

Antimicrobial resistance is a public health concern. Understanding any role that urban seagulls may have as a reservoir of resistant bacteria could be important for reducing transmission. This study investigated fecal Escherichia coli isolates from seagulls (herring gulls and lesser black-backed gulls) to determine the prevalence of extended-spectrum cephalosporin-resistant (ESC-R) and fluoroquinolone-resistant E. coli among gull species from two cities (Taunton and Birmingham) in the United Kingdom (UK). We characterized the genetic background and carriage of plasmid-mediated resistance genes in extended-spectrum β-lactamase (ESBL)-producing E. coli obtained from these birds. Sixty ESC-R E. coli isolates were obtained from 39 seagulls (39/78, 50%), of which 28 (28/60, 46.7%) were positive for plasmid-mediated CTX-M and/or AmpC β-lactamase resistance genes. Among these, blaCTX-M-15, blaCTX-M-14, and blaCMY-2 predominated. Three isolates belonging to the B2-ST131 clone were detected, of which two harbored blaCTX-M-15 (typed to C2/H30Rx) and one harbored blaCTX-M-27 and was typed to C1/H30-R (recently described as the C1-M27 sublineage). The plasmid-mediated quinolone resistance (PMQR) gene carriage prevalence (11.7%) consisted of aac(6')-Ib-cr and qnrB genes. No carbapenem or colistin resistance genes were detected. Urban seagulls in the UK are colonized and can spread major antimicrobial-resistant E. coli isolates harboring ESBL and PMQR determinants, including clinically important strains such as the pandemic clone B2-ST131 and the C1-M27 subclade. This is the first report of ST131-C1-M27 subclade in wildlife in the UK and in seagulls worldwide.

Effect of challenge dose of plasmid-mediated extended-spectrum β-lactamase and AmpC β-lactamase producing Escherichia coli on time-until-colonization and level of excretion in young broilers

Plasmid-mediated extended-spectrum β-lactamase and AmpC β-lactamase (ESBL/pAmpC) producing bacteria are present at all levels of the broiler production pyramid. Young birds can be found positive for ESBL/pAmpC-producing Escherichia coli shortly after arrival at farm. The aim of this study was to determine the effect of different challenge doses of ESBL/pAmpC-producing E. coli on time-until-colonization and the level of excretion in young broilers. One-day-old broilers (specific-pathogen free (SPF) and conventional Ross 308) were housed in isolators and challenged with 0.5 ml ESBL/pAmpC-producing E. coli strains of varying doses (101–105 CFU/ml). Presence and concentration (CFU/gram feces) of ESBL/pAmpC-producing E. coli and total E. coli were determined longitudinally from cloacal swabs, and in cecal content 72 h after challenge. Higher challenge doses resulted in shorter time-until-colonization. However, even the lowest dose (101 CFU/ml) resulted in colonization of the broilers which excreted >106 CFU/gram feces 72 h after inoculation. Conventional broilers were colonized later than SPF broilers, although within 72 h after challenge all broilers were excreting ESBL/pAmpC-producing E. coli. A probabilistic model was used to estimate the probability of colonization by initial inoculation or transmission. The higher the dose the higher the probability of excreting ESBL/pAmpC-producing E. coli as a result of inoculation. In conclusion, low initial doses of ESBL/pAmpC-producing E. coli can result in rapid colonization of a flock. Interventions should thus be aimed to eliminate ESBL/pAmpC-producing bacteria in the environment of the hatchlings and measures focusing at reducing colonization and transmission of ESBL/pAmpC-producing E. coli should be applied shortly after hatching.

Detection of multidrug resistance and extended-spectrum/plasmid-mediated AmpC beta-lactamase genes in Enterobacteriaceae isolates from diseased cats in Italy.

The aim of this study was to determine the antimicrobial susceptibility of Enterobacteriaceae isolated from cats affected by diseases commonly encountered in practice, and to characterise the third-generation cephalosporin (3GC)-resistance molecular mechanisms involved. Clinical samples (n = 100) included 58 rectal swabs from cats with diarrhoea, 31 nasal swabs from cats with clinical signs of upper respiratory tract disease, four ear swabs from cats with otitis, three conjunctival swabs from cats with conjunctivitis, two oral swabs from cats with stomatitis, one swab from a skin abscess and one urine sample from a cat with cystitis. A total of 125 Enterobacteriaceae were isolated from 90 cats. Escherichia coli was the most frequently isolated species (n = 65), followed by Enterobacter species (n = 20), Proteus species (n = 13), Citrobacter species (n = 12) and others (n = 15). Bacterial susceptibility testing was performed with respect to eight antimicrobial classes. Beta (β)-lactamase genes were identified by PCR and nucleotide sequencing. Overall, the higher frequency of resistance was to amoxicillin-clavulanate (61.3%), trimethoprim/sulfamethoxazole (33.6%) and cefotaxime (32.8%). Thirty-six percent of the isolates (n = 45) were resistant to 3GCs. Of these isolates, 34 were tested by PCR and nucleotide sequencing and 23 were confirmed as encoding β-lactamase genes. Fourteen 3GC-resistant isolates harboured extended-spectrum β-lactamases (ESBLs) belonging to groups CTX-M-1 (n = 12, two of which were CTX-M-79), CTX-M-2 (n = 1) and CTX-M-9 (n = 1), as well as SHV-12 (n = 1) and TEM-92 (n = 1). Nine isolates had CMY-2 plasmid-mediated AmpC β-lactamases (pAmpC). Thirty-one percent (n = 39) of the isolates were multidrug resistant (MDR) and were isolated from 34% (n = 31/90) of the cats. A high frequency of MDR and ESBL/pAmpC β-lactamase-producing Enterobacteriaceae were detected among bacteria isolated from a feline population in southern Italy with a variety of common clinical conditions, which poses limitations on therapeutic options for companion animals. We describe the first detection of CTX-M-79 and TEM-92 ESBL genes in isolates from cats.

Induction of plasmid-mediated AmpC β-lactamase DHA-1 by piperacillin/tazobactam and other β-lactams in Enterobacteriaceae.

Chromosomal AmpC β-lactamase induction by several types of β-lactams has been reported, but not enough data are available on DHA-1 β-lactamase, a plasmid-mediated AmpC β-lactamase. Therefore, we evaluated the DHA-1 β-lactamase induction by various antibiotics including piperacillin/tazobactam (PIP/TZB) in this study. Six strains (Enterobacter cloacae 2 strains, Citrobacter freundii 1 strain, Serratia marcescens 2 strain, and Morganella morganii 1 strain) possessing chromosomal inducible AmpC β-lactamase were used as controls. Four strains (Escherichia coli 2 strains, Klebsiella pneumoniae 1 strain, and C. koseri 1 strain) possessing DHA-1 β-lactamase were used. The β-lactamase activities were determined by a spectrophotometer using nitrocefin. β-lactamase induction by PIP, PIP/TZB was not observed in any strains and β-lactamase induction by third- and fourth-generation cephems was not observed in most strains. The induction ratios of the chromosomal AmpC β-lactamase in the reference group by PIP/TZB were <1.51, and those of the DHA-1 β-lactamase were <1.36, except for K. pneumoniae Rkp2004 (2.22). The β-lactamase induction by first- and second-generation cephems, flomoxef, and carbapenem differed in each strain. Cefmetazole (CMZ) strongly induced β-lactamase. This study demonstrated that the induction of DHA-1 β-lactamase was similar to that of chromosomal AmpC using various Enterobacteriaceae, although the induction of β-lactamase in both groups by PIP/TZB was low. We also reported that the induction of PIP/TZB, a β-lactamase inhibitor combination antibiotic, against various AmpC-producing Enterobacteriaceae, including DHA-1 producers, was low.

Phenotypic and genotypic characterization of plasmid-mediated AmpC β-lactamases in Klebsiella pneumoniae isolates from Mansoura University Children Hospital

Phenotypic and genotypic characterization of plasmid-mediated AmpC β-lactamases in Klebsiella pneumoniae isolates from Mansoura University Children Hospital

Multidrug-resistant Escherichia coli isolated from canine faeces in a public park in Quito, Ecuador.

This study focused on estimating the prevalence of extended-spectrum β-lactamases (ESBLs), plasmid-mediated AmpC β-lactamases, carbapenemases and MCR-1-producing Escherichia coli in canine faeces from a public park in Quito, Ecuador. Phenotypic and genotypic characterisation of E. coli isolated from 50 canine faecal samples recovered from a city park in Quito was performed. In addition, a multiple choice survey was conducted among 50 dog owners. Of the 50 faecal samples, 20 (40.0%) presented E. coli resistant to ceftriaxone. Moreover, 23 E. coli isolates were recovered for further analysis. All of the isolates showed as multidrug-resistant (MDR) phenotype (resistant to three or more antibiotic families). Resistance to carbapenems, tigecycline and amikacin was not observed. No major clonal relatedness was observed among the resistant isolates. The ESBL genes blaCTX-M-15, blaCTX-M-55 and blaCTX-M-65 were the most common. Two isolates harboured the blaCMY-2 gene and one isolate harboured both mcr-1 and blaCTX-M-65. Statistical analysis showed that older people were more conscious of collecting and disposing of dog faeces than subjects aged <35 years (P < 0.05). The finding of MDR E. coli in dog faeces in a city park in Ecuador illustrates the importance of analysing canine faeces in public settings (e.g. parks, playgrounds) as part of surveillance programmes for MDR E. coli. In addition, this research might be a sentinel sampling method to gain a better understanding of community sources of antibiotic-resistant Enterobacteriaceae at human-animal-environment interfaces.

Characteristics of third-generation cephalosporin-resistant Salmonella from retail chicken meat produced by integrated broiler operations

Integrated broiler operations, which control and operate vertically through all phases of the chicken industry, have applied biosecurity and sanitation practices, housing technologies, feeding regimens, and antibiotic applications in different ways to improve food safety. The objective of this study was to compare the prevalence and antimicrobial resistance profiles of Salmonella isolates recovered from 6 different integrated broiler operations and to analyze the characteristics of extended-spectrum β-lactamase (ESBL)- and plasmid-mediated AmpC β-lactamase (pAmpC)-producing Salmonella isolates. Among 336 chicken meat samples, 57 were observed to be positive for Salmonella. However, the prevalence varied from 6.8% to 45.8% in chicken meat, indicating variations in Salmonella occurrence among the operations. Salmonella Albany was the dominant serovar, followed by Salmonella Virchow. In the antimicrobial resistance test, nalidixic acid-resistant isolates were the most prevalent (73.7%), followed by isolates resistant to ampicillin (49.1%) and tetracycline (42.1%). Among 14 third-generation cephalosporin-resistant isolates, 9 (64.3%) ESBL/pAmpC-producing isolates were only obtained from 2 operations: blaCTX-M-15 (n = 7) and blaCTX-M-79 (n = 1) for ESBL genes and blaCMY-2 (n = 1) for pAmpC. All ESBL/pAmpC-positive isolates exhibited high minimum inhibitory concentrations (≥128 μg/mL) of most cephalosporins and showed multidrug resistance. The transfer of ESBL/pAmpC genes was confirmed in transconjugants, which had the same genes and similar resistance patterns as those of the donor. Our findings suggest that Salmonella with resistance to third-generation cephalosporins can now be found in association with integrated broiler operations, providing data to support the development of monitoring and prevention programs for the development and spread of antimicrobial resistance in integrated broiler operations.

Detection of plasmid-mediated quinolone resistance genes in β-lactamase-producing Escherichia coliisolates from layer hens

This study was conducted to investigate the presence of plasmid-mediated quinolone resistance (PMQR) genes in β-lactamase-producing Escherichia coli isolates from layer hens and to characterize their molecular background. Among 142 E. coli isolates, 86 (60.6%) showed multidrug resistance and 15 (10.6%) were found to be β-lactamase-producing E. coli. Extended-spectrum β-lactamase (ESBL) and plasmid-mediated AmpC (pAmpC) β-lactamase genes, blaCTX-M-14 and blaCMY-2, were identified in three and six E. coli isolates, respectively. The non-ESBL or pAmpC gene, blaTEM-1, was found in eight of the isolates. Two isolates had both genes, blaCTX-M-14 and blaTEM-1. Among the 15 β-lactamase-producing E. coli, six PMQR genes, qnrS1 (n = 3) and qnrB4 (n = 3), were identified. Among the six PMQR-positive E. coli isolates, four exhibited double amino acid exchanges at both gyrA and parC with ciprofloxacin and enrofloxacin minimum inhibitory concentrations of ≥32 and ≥16μg/mL, respectively. Additionally, five transconjugants (33.3%) showed a transferability of β-lactamase and PMQR genes. Pulsed-field gel electrophoresis (PFGE) analysis was conducted to investigate the 15 β-lactamase-producing E. coli isolates. In PFGE, E. coli included three PFGE patterns showing the same farms and in accordance with both β-lactamase and PMQR genes and the antimicrobial resistance pattern. Layer hens may act as a reservoir of antibiotic-resistant bacteria, and the PMQR gene in β-lactamase-producing E. coli isolates from layer hens has the potential to enter the food chain. Therefore, our findings suggest that comprehensive surveillance of antimicrobial use in laying operation systems is necessary.

Revealing mcr-1-positive ESBL-producing Escherichia coli strains among Enterobacteriaceae from food-producing animals (bovine, swine and poultry) and meat (bovine and swine), Portugal, 2010–2015

We screened 1840 Enterobacteriaceae isolates from food-producing animals, meat, meat products and animal feed, for the detection of plasmid-mediated colistin resistance, during 2010–2015. The mcr-1 gene was detected in 8.0% (97/1206) Escherichia coli and in 0.47% (3/634) Salmonella enterica isolates, with a high number of mcr-1 positive E. coli isolates (45.7%) being extended-spectrum β-lactamase or plasmid-mediated AmpC β-lactamase co-producers. No mcr-2 gene was detected. Our findings highlight the spread of mcr-1 genes within a wide-ranging sample of food-producing animals and meat, in Portugal.

Diversity of Plasmids and Genes Encoding Resistance to Extended Spectrum Cephalosporins in Commensal Escherichia coli From Dutch Livestock in 2007-2017.

Extended-spectrum β-lactamase (ESBL) and plasmid-mediated AmpC β-lactamase (pAmpC) genes confer resistance to extended spectrum cephalosporin’s. The spread of these genes is mostly facilitated by plasmid-mediated horizontal transfer. National surveillance activities to detect ESBL/pAmpC-producers in commensal bacteria from livestock are in place in the Netherlands since several years. This study aimed at reporting gene and plasmid diversity of commensal ESBL/pAmpC-producing Escherichia coli isolated from healthy animals during surveillance activities between 2007 and 2017. A collection of 2304 extended-spectrum cephalosporin-resistant (ESC-R) E. coli isolated from feces of broilers, dairy cattle, slaughter pigs, turkeys, ducks, and veal calves was investigated and ESBL/pAmpC genes were determined. Gene location of a selection of 473 E. coli isolates was determined and typing of plasmids linked to the ESBL/pAmpC genes was performed. Twenty-two different ESBL/pAmpC genes were identified with blaCTX-M-1 being the most prevalent gene in livestock (43.7%), followed by blaCMY -2 and blaSHV -12, independent of the animal source. Prevalence of typically human associated blaCTX-M-15 was highest in cattle. Less than 10% E. coli isolates owed their ESC-R phenotype to promoter mutations of the chromosomal ampC gene. Majority (92%) of ESBL/pAmpC genes analyzed were plasmid located, with IncI1α being the most represented plasmid family in isolates from all animals, followed by IncF (veal calves, dairy cattle and slaughter pigs), IncK (broilers and laying hens), IncX1 in broilers, and emerging IncX3 in broilers and dairy cattle. Prevalence and molecular diversity of ESC-R E. coli isolated from livestock over an 11-year period revealed a composite scenario of gene-plasmid combinations.

Characteristics of cephalosporin-resistant Salmonella isolates from poultry in Korea, 2010-2017.

Cephalosporins are the key drugs for the treatment of salmonellosis. Resistance to cephalosporins in Salmonella spp. has become a serious public health concern worldwide. Although the sales of cephalosporins have increased by five times from 2008 to 2016 in Korea, limited information is available on cephalosporin resistance in Korea. Therefore, we aimed to investigate the trends in prevalence and characteristics of cephalosporin-resistant Salmonella isolates from poultry in Korea in the period between 2010 and 2017. A total of 141 Salmonella isolates were collected from various poultry industry sources, and 38 (27.0%) among them showed resistance to cephalosporins. In particular, resistance to the following cephalosporins increased significantly over the seven-year period cephalothin (from 5.0% to 29.2%), cephalexin, cefuroxime, cefotaxime, and ceftazidime (from 0% to 25.0%), and cefepime (from 0% to 12.5%). In addition, 12 isolates carried a β-lactamase gene. A non-extended-spectrum β-lactamase (ESBL) and plasmid-mediated AmpC β-lactamase (pAmpC) gene, blaTEM-1, was found in three isolates in the periods of 2010 to 2011 and 2012 to 2013, respectively. The blaCTX-M-79 (n = 4) and blaCTX-M-15 (n = 1) for ESBL genes and blaCMY-2 (n = 1) for pAmpC genes were only present in the 2016-2017 period. All ESBL/pAmpC-positive isolates had high minimum inhibitory concentrations for most cephalosporins and showed multi-drug resistance. In a conjugation experiment, the transfer of blaCTX-M-79 and blaCMY-2 genes was confirmed in transconjugants, which showed similar pattern of antibiotic resistance. This demonstrates that ESBL/pAmpC-producing Salmonella isolates might be transmitted to humans through contaminated poultry products. These findings suggest the need for the development of monitoring program and the guidelines for prudent use of antimicrobial agents in the poultry industry in Korea.

First report of New Delhi metallo-β-lactamase-6 (NDM-6) among Klebsiella pneumoniae ST147 strains isolated from dialysis patients in Iran

There has been an alarming health-related concern about the growth of New Delhi metallo-β-lactamase. The aims of this study include the phenotypic detection of β-lactamases and molecular characterization of NDM in Klebsiella pneumoniae isolates in Tehran, Iran. A total of 120 K. pneumoniae isolates were collected from hospitalized haemodialysis patients, Tehran, Iran from March 2014 to February 2017. Antibiotic susceptibility tests were conducted using Kirby-Bauer disc diffusion and Broth Microdilution methods according to Clinical and Laboratory Standards Institute guidelines. Metallo-β-lactamase was detected using the Combined Disc Diffusion Test (CDDT), and production of carbapenemase was screened using the Modified Hodge Test. NDM-producing K. pneumoniae strains were screened for the presence of mcr-1 gene, β-lactamase genes, and 16S rRNA methylase genes by Polymerase Chain Reaction and sequencing. Molecular typing of the strains was determined using Repetitive Sequence Based-PCR and Multilocus Sequence Typing. The blaNDM-6 gene was detected in 3 (2.5%) out of 120 isolates from dialysis patients. Also, the three isolates were positive for blaCTX-M-15,blaTEM extended-spectrum β-lactamase genes, armA type plasmid-mediated 16S rRNA methylase and CMY-type plasmid-mediated AmpC β-lactamase.The isolates were identified as MLST sequence type 147 (ST147). This is the first report of blaNDM-6 in K. pneumoniae strains, isolated in Iran.

Antimicrobial resistance, virulence & plasmid profiles among clinical isolates of Shigella serogroups.

Background & objectives:Bacillary dysentery caused by Shigella spp. remains an important cause of the crisis in low-income countries. It has been observed that Shigella species have become increasingly resistant to most widely used antimicrobials. In this study, the antimicrobial resistance, virulence and plasmid profile of clinical isolates of Shigella species were determined.Methods:Sixty clinical Shigella isolates were subjected to whole-genome sequencing using Ion Torrent platform and the genome sequences were analyzed for the presence of acquired resistance genes, virulence genes and plasmids using web-based software tools.Results:Genome analysis revealed more resistance genes in Shigella flexneri than in other serogroups. Among β-lactamases, blaOXA-1 was predominantly seen followed by the blaTEM-1B and blaEC genes. For quinolone resistance, the qnrS gene was widely seen. Novel mutations in gyrB, parC and parE genes were observed. Cephalosporins resistance gene, blaCTX-M-15 was identified and plasmid-mediated AmpC β-lactamases genes were found among the isolates. Further, a co-trimoxazole resistance gene was identified in most of the isolates studied. Virulence genes such as ipaD, ipaH, virF, senB, iha, capU, lpfA, sigA, pic, sepA, celb and gad were identified. Plasmid analysis revealed that the IncFII was the most commonly seen plasmid type in the isolates.Interpretation & conclusions:The presence of quinolone and cephalosporin resistance genes in Shigella serogroups has serious implications for the further spread of this resistance to other enteric pathogens or commensal organisms. This suggests the need for continuous surveillance to understand the epidemiology of the resistance.

Detection of Plasmid-Mediated β-Lactamase Genes and Emergence of a Novel AmpC (CMH-1) in Enterobacter cloacae at a Medical Center in Southern Taiwan.

The plasmid-mediated extended-spectrum β-lactamases (ESBLs) and AmpC β-lactamases in Enterobacter spp. have increasingly been reported. In this study, we investigated the prevalence of the plasmid-mediated β-lactamases in Enterobacter cloacae from bloodstream isolates at a medical center in southern Taiwan. ESBL and ampC genes were detected by PCRs and DNA sequencing. Conjugation experiments were conducted to confirm the transferability of the genetic resistance trait. Among 41 non-repetitive blood isolates of cefuroxime-resistant E. cloacae, eight isolates exhibited ESBL phenotype confirmed by double-disk synergistic tests. Nearly all the strains were susceptible to carbapenems. The prevalence rate of the plasmid-mediated blaampC genes was 73% (30/41), including one blaDHA-1, one blaMIR-6, two novel blaCMH-1 genes and other blaACT-like genes. Coexistence of plasmid-mediated blaACT and ESBL genes (10 with blaSHV-12 and one with blaCTX-M-3) was observed. Successful transmissions of the blaACT and blaCMH-1 were demonstrated in some transconjugants. The inducible or derepressed CMH-1 had expanded activity of isolates versus ceftazidime. Enterobacterial repetitive intergenic consensus (ERIC)-PCR analysis and pulsotype showed distinct patterns suggesting non-clonal relationship. In conclusion, plasmid-mediated blaACT-like ampC genes in E. cloacae isolates have been highly prevalent in southern Taiwan and may continue genetic evolution, contributing to the complexities in antibiotic-resistant mechanisms.

1200. Molecular Epidemiology of Cephalosporinases and Extended Spectrum β-Lactamases (ESBLs) in Proteus mirabilis Isolates From Croatia: Following the Spread of Resistance Determinants Between Long-Term Care Facilities and the Community

BackgroundPrevious studies on P. mirabilis strains isolated from Croatian healthcare institutions revealed the predominance of TEM-52 extended spectrum β-lactamase (ESBL), as well as the emergence of plasmid AmpC β-lactamases. Our aim was to molecularly characterize cefalosporinases in P. mirabilis isolates from long-term care facilities (LTCFs) and to compare their resistance profile and dynamics with community isolates.MethodsFrom a total of 3,321 P. mirabilis isolates collected from two LCTFs and from outpatients between 2015 and 2017, 1.23% of them were resistant to third generation of cephalosporins. Antimicrobial sensitivity was tested by broth microdilution method. ESBLs and plasmid-mediated AmpC β-lactamases were detected with phenotypic inhibitor-based tests and polymerase chain reaction (PCR). Antibiotic resistance dissemination and genetic context of bla genes were interrogated by conjugal mating and PCR mapping, respectively. Plasmids were characterized by conjugation and transformation experiments, as well as PCR-based replicon typing.ResultsHigh-level of resistance to amoxicillin, co-amoxiclav, first, second and third generation of cephalosporins was found in all isolates. Three isolates tested positive in inhibitor-based test with clavulanic acid, and 38 both in Hodge test and combined disk test with phenylboronic acid, indicating the production of ESBLs and plasmid-mediated AmpC β-lactamases, respectively. Two ESBL-positive organisms yielded amplicons with primers for CTX-M β-lactamase of group 1 and one for TEM. All AmpC-positive organisms were identified by PCR as CMY (with an additional TEM). Insert sequence ISEcp-1 was found upstream of blaCMYi blaCTX-M genes. CTX-M positive strains harbored IncK plasmid, whereas AmpC-positive strains were negative for known plasmid types. This is also a first description of P. mirabilis harboring CTX-M-15 β-lactamase in Croatia.ConclusionOur study showed the persistence of CMY β-lactamases in one LTCF, but also the dissemination of characteristic resistance determinants to another LTCF and the community. Similar to some other studies, there was a clear trend of cephalosporinase dynamic switch from TEM variants to CMY and CTX-M, with impending consequences for treatment decisions.Disclosures All authors: No reported disclosures.

Multiple antibiotic resistances and virulence markers of uropathogenic Escherichia coli from Mexico

ABSTRACTVirulence and antibiotic resistance properties related to different Escherichia coli phylogenetic groups have not been studied in detail in Mexico. We aimed to identify patterns of virulence genes and multidrug resistance in phylogenetic groups of uropathogenic strains (UPEC). Strains of E. coli were isolated from outpatients with urinary tract infections (UTIs), who went to unit of the public health sector in the State of Mexico. E. coli virulence markers and phylogenetic groups were identified by PCR. Susceptibility to 12 antimicrobials was determined by Kirby-Bauer. E. coli was identified in 60.4% (n = 194) of the patients with UTIs. Phylogroups B2 51% (n = 99), A 13.4% (n = 26) and B1 10.3% (n = 20) were the most frequent. Resistance to three or up to eleven antibiotics was detected in most phylogroups (n = 188). The genes fimH (n = 146), feoB (n = 179), iutA (n = 178), sitA (n = 121), fyuA (n = 99), and traT (n = 142) were mainly detected in strains of phylogroups B2, A, B1, C, and D. Seventy-two patterns of virulence markers were distributed across eight E. coli phylogenetic groups. A high frequency of virulence markers and the multiple antibiotic resistance phenotypes was observed in the phylogroups. The genes of extended-spectrum β-lactamases (ESBLs) found with higher frequency among UPEC strains were blaTEM, blaSHV y blaCTX-M group 1, CIT (plasmid-mediated AmpC β-lactamase), and blaOXA-like. In conclusion, our findings show the importance of surveillance, permanent monitoring, and particularly controlled prescription of antibiotics by physicians in the social security health system to reduce the spread of highly virulent UPEC strains that are resistant to multiple antimicrobial agents.

Evaluation of the Inter- and Intrahospital Spread of Multidrug Resistant Gram-Negative Bacteria in Lithuanian Hospitals.

Spread of multidrug-resistant pathogenic bacteria became one of the greatest threats in healthcare worldwide. It is generally accepted that both inter- and intrahospital transmissions of these bacteria contribute significantly to this problem. The purpose of the current study was the evaluation of the inter- and intrahospital spread of multidrug resistant Gram-negative pathogenic bacteria in Lithuania. Clinical isolates of Acinetobacter sp., Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa were subjected for the screening for extended spectrum β-lactamase, carbapenemase, as well as plasmid-mediated AmpC β-lactamase genes. BOX-PCR genotyping was used for the genotyping of these isolates. Our results show that all four pathogens are involved in the intra- and/or interhospital dissemination between the Lithuanian healthcare institutions. The level of transmissions differed between pathogens, and the worst situation was detected for Acinetobacter sp. followed by E. coli. In almost all cases, transmissible strains had at least one gene conferring β-lactam resistance, thereby contributing to the dissemination of the resistance determinants in and between Lithuanian hospitals. Our study clearly demonstrated that immediate actions, more effective strategy, and surveillance are needed to confine and prevent further spread of multidrug resistant Gram-negative pathogenic bacteria in Lithuanian healthcare institutions.