Bla KPC Research Articles

Metagenomic analysis of urban wastewater resistome and mobilome: A support for antimicrobial resistance surveillance in an endemic country.

In developing countries, where high levels of antimicrobial resistance are observed in hospitals, the surveillance of this phenomenon in wastewater treatment plants (WWTPs) and the environment is very limited, especially using cutting-edge culture-independent methods. In this study, the composition of bacterial communities, the resistome and mobilome (the pool of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs), respectively) at a WWTP were determined using shotgun metagenomics and culture-based approaches. Wastewater samples were collected at four sampling points of a WWTP in Antioquia, Colombia. A total of 24 metagenomes were analyzed. Specifically, there were marked differences in bacterial community composition, resistome, and mobilome, according to the WWTP sampling points. Bacterial families of clinical importance such as Moraxellaceae, Aeromonadaceae, and Enterobacteriaceae were mainly detected in the WWTP influent and effluent samples. Genes encoding resistance to macrolide-lincosamide-streptogramin, β-lactams, and those conferring multidrug resistance (e.g., acrB, adeG, and mexD) were the most abundant. Moreover, some clinically important ARGs such as blaKPC-2 and blaCTX-M, and others not reported locally, such as blaTEM-196, blaGES-23, blaOXA-10, mcr-3, and mcr-5 were frequently detected. Co-occurrence network analyses indicated a significant association of ARGs such as blaOXA-58 and blaKPC genes with Aeromonadaceae and Enterobacteriaceae. Among the markers of MGEs, intI1 and ISCR8 were the most frequently detected. Altogether, this work reveals the importance of shotgun metagenomics and culture-based approaches in antimicrobial resistance studies. The findings also support that WWTPs are hotspots for antimicrobial resistance, whose analysis constitutes a powerful tool to predict the impact of antimicrobial resistance in a population.

Resistance evolution of hypervirulent carbapenem-resistant Klebsiella pneumoniae ST11 during treatment with tigecycline and polymyxin

ABSTRACT Hypervirulent carbapenem-resistant Klebsiella pneumoniae (hv-CRKP) has recently aroused increasing attention, especially ST11, the predominant CRKP clone in China. Here, we report a case of hv-CRKP-associated infection and reveal the in-host evolution of its mechanism of resistance to tigecycline and polymyxin under clinical therapy. A total of 11 K. pneumoniae carbapenemase (KPC)-producing CRKP strains were consecutively isolated from a male patient who suffered from continuous and multisite infections. String and antimicrobial susceptibility tests identified seven hypermucoviscous strains and three tigecycline-resistant and four colistin-resistant strains. Galleria mellonella larvae infection model confirmed the hypervirulence. Pulsed-field gel electrophoresis (PFGE) separated three PFGE clusters among all strains, and further Southern blotting detected that bla KPC-2 was located on the same-sized plasmid. Whole-genome sequencing showed that all strains belonged to the hv-CRKP ST11-KL64 clone. Diverse hypervirulence factors and resistance genes were identified. Further sequencing with the Nanopore platform was performed on the CRKP-Urine1 strain, which contained one virulence plasmid (pVi-CRKP-Urine1) and two resistance plasmids (pKPC-CRKP-Urine1 and pqnrS1-CRKP-Urine1). The gene mutations responsible for tigecycline or colistin resistance were then amplified with PCR followed by sequencing, which indicated that mutations of ramR and lon were the potential loci for tigecycline resistance and that the pmrB, phoQ and mgrB genes for colistin resistance. A novel frameshift mutation of lon was identified in the high-level tigecycline-resistant strain (MIC, 128 mg/L). The results indicate that the hypervirulent ST11-KL64 clone is a potential threat to antiinfection treatment and is capable of rapid and diverse evolution of resistance during tigecycline and polymyxin treatment.

Mutations in porin LamB contribute to ceftazidime-avibactam resistance in KPC-producing Klebsiella pneumoniae

Ceftazidime-avibactam (CAZ-AVI) shows promising activity against carbapenem-resistant Klebsiella pneumoniae (CRKP), however, CAZ-AVI resistance have emerged recently. Mutations in KPCs, porins OmpK35 and/or OmpK36, and PBPs are known to contribute to the resistance to CAZ-AVI in CRKP. To identify novel CAZ-AVI resistance mechanism, we generated 10 CAZ-AVI-resistant strains from 14 CAZ-AVI susceptible KPC-producing K. pneumoniae (KPC-Kp) strains through in vitro multipassage resistance selection using low concentrations of CAZ-AVI. Comparative genomic analysis for the original and derived mutants identified CAZ-AVI resistance-associated mutations in KPCs, PBP3 (encoded by ftsI), and LamB, an outer membrane maltoporin. CAZ-AVI susceptible KPC-Kp strains became resistant when complemented with mutated bla KPC genes. Complementation experiments also showed that a plasmid borne copy of wild-type lamB or ftsI gene reduced the MIC value of CAZ-AVI in the induced resistant strains. In addition, bla KPC expression level increased in four of the six CAZ-AVI-resistant strains without KPC mutations, indicating a probable association between increased bla KPC expression and increased resistance in these strains. In conclusion, we here identified a novel mechanism of CAZ-AVI resistance associated with mutations in porin LamB in KPC-Kp.

First report of the aac(6')-Ib-cr gene in Providencia stuartii isolates in Brazil.

INTRODUCTION: The aac(6’)-Ib-cr and bla KPC genes are spreading among Enterobacteriaceae species, including Providencia stuartii, in some countries of world. METHODS: These genes were investigated in 28 P. stuartii isolates from a public hospital in Recife, Pernambuco, Brazil, by PCR and sequencing. RESULTS: The aac(6’)-Ib-cr gene was detected in 16 resistant isolates, and the bla KPC gene was seen in 14.CONCLUSIONS: The presence of these genes in P. stuartii multi- and extensively drug-resistant isolates indicates that the resistance arsenal of this species is increasing, thus limiting the therapeutic options.

Carbapenem-Resistant Klebsiella pneumonia Isolated from Patients Admitted in Tertiary Care Hospital in Saudi Arabia

Background: Carbapenem-resistant klebsiella pneumoniae is an emerging threat worldwide causing high rates of morbidity and mortality
 Aim: To evaluate the prevalence of carbapenem-resistant K. pneumonia (CRKP), associated risk factors, type of infections caused by CRKP and their antimicrobial susceptibility. To evaluate Carbapenemase Detection Set (D70C) as screening test for CRKP
 Place and Duration of the Study: A cross sectional study and prospective cohort study was performed from June 2019 to February 2020 in intensive care unit and medical units of Al Quwayiyah General hospital.
 Methodology: 541 samples were collected from different patient sources. Klebsiella pneumoniae strain was only selected identified to the species level and AST was done using the Vitek-2. Minimum inhibitory concentration (MIC) of meropenem and imipenem was carried out. A Carbapenemase Detection Set (D70C) was used as screening test for CRKP while Modified Hodge test and multiplex PCR as confirmatory tests.
 Results: A total of 132 isolates were diagnosed as Enterobacteriaceae out of 541 patient samples.78 clinical isolates were klebsiella pneumoniae which were collected. Out of the 78 clinical isolates CRKP were 36 (46.2%) and CSKP were 42 (53.8%).) CRKP cases aged from (18-84 years) with the median patient age 59 year. Seventeen of 36 patients (47.2%) were males. the majority of the nosocomial CRKP infections were pneumonia 12 (33.3%) followed by urinary tract infection 9 (25%). The most common associated disease was diabetes (30%) followed by renal disease (27.8%). For invasive procedures, Urinary catheter was 27(75%) and 29(69%) followed by Mechanical ventilation 25(69.4%) and 22(52.4%) in CRKP and CSKP patients respectively. Reports of PCR for the 41 isolates which sent to regional laboratory for confirmation revealed that 36 isolates had carbapenemase genes; twenty eight (77.8%) K. pneumonia isolates positive for bla OXA-48 and 5 (13.9%) isolates were positive for blaNDM. in 2 (5.6%) bla KPC were detected, one isolate contained blaIMP. 5 isolates contain both blaOXA-48 and blaNDM. The sensitivity of MHT was analysed to be 91.7%. (95%Cl ratio 77.53% - 98.25%) and the specificity was 100% (95%Cl ratio 54.07% to 100%). The positive predictive value was 100% and the Negative predictive value was 66.7% ( 95%Cl ratio 40.36% to 85.53%). The sensitivity of Carbapenemase Detection Set (D70C) was 94.4% (81.34% to 99.32%) and the specificity was 80% (95%Cl ratio 28.36% to 99.49%). The positive predictive value was 97.1% (95%Cl ratio 85.46% to 99.49%).and the Negative predictive value was 66.7% (95%Cl ratio 32.67% to 89.18%).
 Conclusion: CRKP prevalence was 46.2% among K. pneumoniae isolates in Al Quwayiya General Hospital. Using invasive procedures such as urinary catheters or mechanical ventilator and misuse of antibiotics were risk factors associated with CRKP indicating that infection control guidelines and effective preventive measures should be strictly applied. It is very important to monitor and report changes in antimicrobial-resistant isolates but Carbapenemase Detection Set (D70C) has low specificity makes it less reliable and need PCR confirmation.

Reflections on the Predictability of Evolution: Toward a Conceptual Framework.

SummaryEvolution is generally considered to be unpredictable because genetic variations are known to occur randomly. However, remarkable patterns of repeated convergent evolution are observed, for instance, loss of pigments by organisms living in caves. Analogous phenotypes appear in similar environments, sometimes in response to similar constraints. Alongside randomness, a certain evolutionary determinism also exists, for instance, the selection of particular phenotypes subjected to particular environmental constraints in the “evolutionary funnel.” We pursue the idea that eco-evolutionary specialization is in some way determinist. The conceptual framework of phenotypic changes entailing specialization presented in this essay explains how evolution can be predicted. We also discuss how the predictability of evolution could be tested using the case of metabolic specialization through gene losses. We also put forward that microorganisms could be key models to test and possibly make headway evolutionary predictions and knowledge about evolution.

Colistin resistance in multidrug-resistant Klebsiella pneumoniae strains at a perinatal maternal institute in Lima, Peru, 2015-2018.

The objective of this study was to evaluate the presence of colistin- and carbapenemic-resistant genes in multidrug-resistant Klebsiella pneumoniae strains isolated at the Instituto Materno Perinatal de Lima (2015-2018). Susceptibility levels were analyzed by disk diffusion and microdilution. The presence of colistin- and carbapenemic-resistant genes was determined by polymerase chain reaction (PCR) and was related to clonality. A total of 36 K. pneumoniae strains were analyzed, 5 (13.8%) were resistant to colistin and belonged to different clonal groups. Only 2 strains were found with carbapenemases (bla KPC and bla NDM), and no colistin plasmid genes were detected. High resistance levels to the other tested antimicrobials were observed, except for amikacin (13.9%). The results highlight the presence of colistin-resistant strains (33.3% in 2018), a worrying situation as they are part of the latest treatment alternatives for infections caused by multiresistant pathogens.

Co-Occurrence of the bla KPC-2 and Mcr-3.3 Gene in Aeromonas caviae SCAc2001 Isolated from Patients with Diarrheal Disease.

PurposeTo characterize the genetic feature of a multi-drug-resistant Aeromonas caviae strain isolated from the diarrhea sample of a 45-year-old male patient with acute diarrhea.Materials and MethodsWhole-genome of the A. caviae strain SCAc2001 was sequenced via the Illumina system, followed by a series of bioinformatic analyses to describe the genetic feature.ResultsThe genome sequence of A. caviae SCAc2001 was assembled into 340 scaffolds (305 of them were > 1000 bp in length and 4,487,370 bp in total) with an average G+C content of 61.09%. Phylogenetic analysis showed that the A. caviae SCAc2001 strain was highly similar to the A. caviae strain R25-2 and T25-39. Resistome analysis identified that A. caviae SCAc2001 carried 13 antimicrobial resistance genes, including β-lactams (blaKPC, blaCTX-M-14, blaTEM-1, blaOXA-10, blaOXA-427, blaVEB-3 and blaMOX-6), aminoglycosides (aadA1), fluoroquinolones (aac(6ʹ)-Ib-cr), phenicol resistance (catB3), sulfonamide (sul1), trimethoprim (dfrA5) and colistin resistance (mcr-3.3).And also, A. caviae ScAc2001 carried 54 putative virulence genes including the type IV pilus, fimbria, flagellarthe, and hemolysin A encoding genes, and 12 pathogen–host interactions (PHI) genes. There were also four genomic islands and eight prophages in the genome of A. caviae ScAc2001. In addition, A. caviae SCAc2001 also carried three secondary metabolism products coding clusters including nonribosomal peptide synthetases (nrps), hserlactone and bacteriocin.ConclusionA. caviae ScAc2001 carries many resistance genes, a variety of virulence factors, PHI genes and four genomic islands and eight prophages, which poses a severe threat to infectious diseases control strategies, diagnosis methods and clinical treatment.

Whole genome sequence of blaNDM and blaKPC co-producing Klebsiella pneumoniae isolate KSH203 with capsular serotype K25 belonging to ST11 from China.

The aim of this study was to characterise a high biofilm-forming capacity, hypermucoviscous, blaKPC and blaNDM co-producing Klebsiella pneumoniae strain (KSH203). Antimicrobial susceptibility, biofilm formation and hypermucoviscous phenotype were determined by the disk diffusion method, crystal violet staining and positive string test, respectively. Whole-genome sequencing was performed using a PacBio RS II Sequencer. High-quality reads were de novo assembled using Celera Assembler v.8.0. Genome annotation was performed using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP), and the genome characteristics were analysed by bioinformatics methods. Klebsiella pneumoniae strain KSH203 was resistant to all antibiotics tested but was only intermediate-resistant to polymyxin B. This strain showed high biofilm-forming ability and a hypermucoviscous phenotype with serotype K25 belonging to the ST11 clone. KSH203 consists of a 5 464 059-bp single chromosome and four plasmids including pKSH203-NDM (53 144 bp), pKSH203-KPC (159 467 bp), pKSH203-CTX-M-3 (156 910 bp) and pKSH203-qnrS (253 705 bp). A total of 44 antimicrobial resistance genes and a large number virulence-associated genes were identified in the genome of strain KSH203. In this study, we illustrate the whole genome sequence of high biofilm-forming capacity, hypermucoviscous K. pneumoniae isolate KSH203 with capsular serotype K25 belonging to ST11 isolated from a patient in China, which carried a large number of antimicrobial resistance genes and virulence-associated genes. Future studies are needed to be aware of dissemination of this type of strain among environmental, animal and human isolates.

Multidrug-Resistance Genes in Pseudomonas aeruginosa from Wound Infections in a Tertiary Health Institution in Osogbo, Nigeria.

Multidrug Resistant Pseudomonas aeruginosa (MDRPA) is a ubiquitous opportunistic organism that poses threat to the management of infections globally. The objectives of the current research were to assess the antibiotic resistance profiles as well as Multiple Antibiotic Resistance (MAR) Index of clinical isolates of P. aeruginosa associated with wound infections. Presence of Extended Spectrum Beta Lactamase genes (bla CTX-M, bla SHV and bla TEM) and Carbapenemase genes (bla KPC and blaNDM) were also determined among the isolates. Swab samples were collected from 255 patients with wound infections. Bacterial identification was done by standard diagnostic tests. The identity of isolates was confirmed by the detection of the exoA gene using the PCR technique. Antibiotic susceptibility testing and resistance profile were determined using the disc diffusion method. Resistance genes were amplified by the PCR method. A total of 235 (92.2%) bacterial isolates were recovered from the wounds of the 255 patients, of these, 124 (52.8%) were Gram-negative bacilli while the remaining 111 (47.2%) were Gram-positive cocci. A total of 69 Pseudomonas aeruginosa strains were recovered from the wound specimens. Imipenem was the most effective antibiotic against these isolates (92.8% isolates were susceptible) while all isolates were resistant to Meropenem, Cefepime, Ticarcillin, Amoxicillin-clavulanic acid, Cefotaxime, Ampicillin and Cefpodoxime. All 69 Pseudomonas aeruginosa isolates were multidrug resistant (MDR). Of the isolates selected for PCR, all were positive for TEM, CTX-M and SHV genes while one-third were blaKPC and blaNDM producers. This study demonstrated high prevalence of carbapenem-resistant strains of P. aeruginosa, suggesting that there is an urgent need in Nigeria for the enactment and enforcement of policies and necessary laws restricting the availability and indiscriminate use of antibiotics.

Identification of plasmid IncQ1 and NTEKPC-IId harboring bla KPC-2 in isolates from Klebsiella pneumoniae infections in patients from Recife-PE, Brazil.

INTRODUCTION: This study investigated the genetic environment of bla KPC-2 in Klebsiella pnemoniae multi-drug resistant clinical isolates. METHODS: Four carbapenemase gene isolates resistant to carbapenems, collected from infected patients from two hospitals in Brazil, were investigated using polymerase chain reaction and plasmid DNA sequencing. RESULTS: The bla KPC-2 gene was located between ISKpn6 and a resolvase tnpR in the non-Tn4401 element (NTEKPC-IId). It was detected on a plasmid belonging to the IncQ1 group. CONCLUSIONS To our knowledge, this is the first report of the presence of the bla KPC-2 gene in the NTEKPC-IId element carried by plasmid IncQ1 from infections in Brazil.

Cointegration as a mechanism for the evolution of a KPC-producing multidrug resistance plasmid in Proteus mirabilis

ABSTRACT The incidence and transmission of Klebsiella pneumoniae carbapenemase (KPC) producing plasmids have been well documented. However, the evolutionary dynamics of KPC plasmids and their fitness costs are not well characterized. Here, two carbapenemase-producing plasmids from Proteus mirabilis, pT18 and pT211 (both carrying bla KPC-2), were characterized through whole genome sequencing. pT211 is a 24.2 kbp N-type plasmid that contains bla KPC-2 and a single copy of the IS6-family insertion sequence IS26. pT18 is a 59 kbp cointegrate plasmid comprised of sequences derived from three different plasmids: a close relative of pT211 (containing bla KPC-2), an FII-33 plasmid (bla TEM-1B, bla CTX-M-65, rmtB and fosA3) and a rolling-circle plasmid. The segments of pT18 derived from each of the different plasmids are separated by copies of IS26, and sequence analysis indicated that pT18 was likely generated by both conservative and replicative IS26-mediated cointegrate formation. pT18 and pT211 were transferred into Escherichia coli DH5α separately to assess the impact of plasmids on host fitness. Only DH5α harbouring pT18 grew slower than the wild type in antibiotic-free media. However, in sub-inhibitory concentrations of fosfomycin and amikacin, cells containing pT18 grew faster than the wild type, and the minimum concentrations of fosfomycin and amikacin required to observe an advantage for plasmid-carrying cells were 1/3 and 1/20 the DH5α MIC, respectively. This study highlights the importance of the role of cointegrate plasmids in the dissemination of antibiotic resistance genes between pathogenic bacterial species, and highlights the importance of sub-inhibitory concentrations of antibiotics to the persistence of such plasmids.

Prevalence of Klebsiella pneumoniae carbapenemase - and New Delhi metallo-beta-lactamase-positive K. pneumoniae in Sergipe, Brazil, and combination therapy as a potential treatment option.

INTRODUCTION:Carbapenem-resistant Klebsiella pneumoniae infection lacks treatment options and is associated with prolonged hospital stays and high mortality rates. The production of carbapenemases is one of the most important factors responsible for this multi-resistance phenomenon.METHODS:In the present study, we analyzed the presence of genes encoding carbapenemases in K. pneumoniae isolates circulating in one of the public hospitals in the city of Aracaju, Sergipe, Brazil. We also determined the best combination of drugs that display in vitro antimicrobial synergy. First, 147 carbapenem-resistant K. pneumoniae isolates were validated for the presence of blaKPC, bla GES, bla NDM, bla SPM, bla IMP, bla VIM, and bla OXA-48 genes using multiplex polymerase chain reaction. Thereafter, using two isolates (97 and 102), the role of double and triple combinational drug therapy as a treatment option was analyzed.RESULTS:Seventy-four (50.3%) isolates were positive for bla NDM, eight (5.4%) for bla KPC, and one (1.2%) for both bla NDM and bla KPC. In the synergy tests, double combinations were better than triple combinations. Polymyxin B and amikacin for isolate 97 and polymyxin B coupled with meropenem for isolate 102 showed the best response.CONCLUSIONS:Clinicians in normal practice use multiple drugs to treat infections caused by multi-resistant microorganism; however, in most cases, the benefit of the combinations is unknown. In vitro synergistic tests, such as those described herein, are important as they might help select an appropriate multi-drug antibiotic therapy and a correct dosage, ultimately reducing toxicities and the development of antibiotic resistance.

Prevalence, risk factors and molecular epidemiology of carbapenem-resistant Klebsiella pneumoniae in patients from Zhejiang, China, 2008–2018

ABSTRACT Carbapenem-resistant Klebsiella pneumoniae (CRKP) is emerging as a worldwide public health concern; however, the long-term molecular epidemiological surveillance of clinical CRKP in China is limited. We conducted a retrospective observational study (2008–2018) to assess the prevalence, susceptibility, risk factors and molecular epidemiology of clinical CRKP isolates. We found the prevalence of CRKP increased from 2.5%, 2008 to 15.8%, 2018. CRKP were significantly more frequent among hospitalized patients from ICU, and it was significantly more likely to be isolated from the capital city (Hangzhou) and the patients aged ≥60 years. Additionally, seasons and specimen types were associated with CRKP infections. The main CRKP sequence type (ST) was ST11, and bla KPC-2 was the most prevalent gene variant. Together these data reveal an increasing incidence and resistance trends among CRKP, especially the ST11-bla KPC-2-CRKP, in Zhejiang, during 2008–2018. Our findings are important for hospitals to limit its dissemination and optimize antibiotic administration.

Emergence of ST11-K47 and ST11-K64 hypervirulent carbapenem-resistant Klebsiella pneumoniae in bacterial liver abscesses from China: a molecular, biological, and epidemiological study

ABSTRACT Background: Multidrug-resistant bacteria, especially those with high virulence, are an emerging problem in clinical settings. Methods: We conducted a multicentre epidemiological and comparative genomic analysis on the evolution, virulence and antimicrobial resistance of carbapenem-resistant Enterobacteriaceae in patients with bacterial liver abscesses from 2012 to 2016. Results: A total of 477 bacterial isolates were collected. Enterobacteriaceae were the main pathogen (89.3%) with K. pneumoniae (52.4%) predominating followed by Escherichia coli (26.8%). All CRKps (3.2%) were of sequence type (ST) 11 and serotypes K47 or K64, and simultaneously possessed acquired bla KPC-2/bla KPC-5 and bla CTX-M-65 together with the multidrug transporter EmrE. Seven Hv-CRKps (five ST11-K47, two ST11-K64) were confirmed by bacteriological test, neutrophil killing assay and Galleria mellonella infection model. Genomic analysis indicated that the emergence of one ST11-K64 Hv-CRKp strain was related to the acquisition of rmpA/rmpA2 genes and siderophore gene clusters, while ST11-K47 Hv-CRKp lacked these traditional virulence genes. Further complete genome analysis of one ST11-K47 Hv-CRKp strain, R16, showed that it acquired a rare plasmid (pR16-Hv-CRKp1) carrying bla KPC-2, bla SHV-12, bla TEM-1, bla CTX-M-65, rmtB and a predicted virulence gene R16_5486 simultaneously. Conclusion: The emergence of the ST11-K47/K64 Hv-CRKps, which are simultaneously multidrug-resistant and hypervirulent, requires urgent control measures to be implemented.

High plasmid variability, and the presence of IncFIB, IncQ, IncA/C, IncHI1B, and IncL/M in clinical isolates of Klebsiella pneumoniae with bla KPC and bla NDM from patients at a public hospital in Brazil.

INTRODUCTION Antibiotic resistance in carbapenemase-producing Klebsiella pneumoniae is acquired and disseminated mainly by plasmids. Therefore, we aimed to investigate the occurrence of carbapenemase genes, analyze the genetic diversity by ERIC-PCR, and examine the most common plasmid incompatibility groups (Incs) in clinical isolates of K. pneumoniae from colonization and infection in patients from a hospital in Brazil. METHODS Twenty-seven isolates of carbapenem-resistant K. pneumoniae were selected and screened for the presence of carbapenemase genes and Incs by PCR, followed by amplicon sequencing. RESULTS The bla KPC and bla NDM genes were detected in 24 (88.8 %) and 16 (59.2 %) of the isolates, respectively. Thirteen isolates (48.1 %) were positive for both genes. The IncFIB (92.6 %) and IncQ (88.8 %) were the most frequent plasmids, followed by IncA/C, IncHI1B, and IncL/M, indicating that plasmid variability existed in these isolates. To our knowledge, this is the first report of IncHI1B in Brazil. We found eight isolates with clonal relationship distributed in different sectors of the hospital. CONCLUSIONS The accumulation of resistance determinants, the variability of plasmid Incs, and the clonal dissemination detected in K. pneumoniae isolates demonstrate their potential for infection, colonization, and the dissemination of different resistance genes and plasmids.

Frequency and molecular epidemiology of class A ESBLs producing Enteroinvasive Escherichia coli (EIEC) isolates among patients with diarrhea.

This study aimed to investigate the frequency and molecular epidemiology of class A ESBLs producing Enteroinvasive Escherichia coli (EIEC) isolates among patients with diarrhea. Antibiotic resistance is widespread among diarrheagenic Escherichia coli (DEC) in developing countries. Information regarding Extended-Spectrum β-Lactamases (ESBLs) in diarrheagenic pathogens should be considered in clinical management when an optimal treatment is required. A total of 581 stool samples were collected from patients with diarrhea in Ahvaz, Iran. PCR was used for the presence of the ipaH gene to confirm EIEC strains. The antibiotic resistance pattern of all EIEC isolates was determined by the disk diffusion method. EIEC isolates were screened for class A β-lactamase genes. Genotyping of harboring β-lactamase genes was performed by Multi-Locus VNTR Analysis (MLVA). Among 13 EIEC isolates, 9 isolates (69.2%) were found ESBL positive by double-disk synergy test (DDST) and PCR. Furthermore, bla CTX-M-15 and bla CTX-M-1 genes were detected in 77.8% (n=7) and 44.5% (n=4) of the bla CTX-M-1 group. On the other hand, the bla TEM-1 gene was detected in 66.6% (n=6). None of the isolates had bla SHV-1, bla KPC, or bla GES genes. Six MLVA genotypes were identified. The current study revealed that the presence of ESBLs genes mediates the resistance of EIEC isolates to the majority of antibiotics in this region. The presence of ESBLs genes in different MLVA types showed that one specific clone was not responsible for spreading the EIEC isolates.

Colistin-resistance gene mcr in clinical carbapenem-resistant Enterobacteriaceae strains in China, 2014–2019

ABSTRACT To investigate whether introduction of colistin into the clinical settings selected colistin-resistant CRE, we performed molecular epidemiological study of 1868 CRE strains collected from different geographical locales in China during the period 2014–2019. 1755 (96.18%) isolates carried the carbapenemase genes bla KPC and bla NDM; 14 Escherichia coli isolates (0.75%) carrying mcr-1 and bla NDM (MCR-CREC) were also identified. Importantly, the number and relative prevalence of MCR-CREC isolates increased from 5 (0.41%) to 9 (1.38%) after introduction of polymyxin into clinical practice. Consistently, results of genetic analysis indicated that MCR-CREC strains collected before December 2017 were genetically diverse, yet those collected after that date exhibited more closely related genetic profiles, indicating that specific MCR-CREC strains were rapidly selected as a result of increased usage of colistin in clinical settings. The resistance level of MCR-CREC isolates to colistin increased after the introduction of polymyxin into clinical use with the MIC to colistin from <2 mg/L in 80% strains to 2 mg/L in 100% strains. Further dissemination of MCR-CREC strains, which exhibit resistance to the last-line drugs of carbapenems and colistin, is expected to pose a severe threat to human health.

Clonal dissemination of KPC-2-producing Klebsiella pneumoniae ST11 and ST48 clone among multiple departments in a tertiary teaching hospital in Jiangsu Province, China.

The world-wide prevalence of carbapenem-resistant Klebsiella pneumoniae (CRKP) poses a threat to the public health. The objective of this study was to determine the epidemiological and molecular patterns of KPC-producing Klebsiella pneumoniae (K. pneumoniae) clinical isolates. In this study, a total of 82 non-duplicated CRKP isolates were analyzed for the prevalence of resistant determinants including carbapenemase, extended spectrum β-lactamase (ESBLs), and AmpC as well as integrons and cassette regions by polymerase chain reaction (PCR) and DNA sequencing. The genetic relatedness was investigated by pulsed field gel electrophoresis (PFGE) and multi-locus sequencing typing (MLST). Overall, bla KPC-2 (n=75) was the predominant carbapenemase gene, followed by high prevalence of bla SHV (92.7%) and bla CTX-M (90.2%). PFGE and MLST analysis revealed that 65 out of 68 KPC-2-producing CRKP belonged to the ST11 clone and were distributed mainly in the department of neurology ICU. Moreover, first report on clonal dissemination of KPC-2-producing CRKP ST48 clone and NDM-5-producing CRKP ST337 clone was also identified. Class I integron were detected in 17 (20.7%) of 82 isolates with aadA2 being the most common cassette. And a novel cassette array of integron, aac(6')-II-bla CARB/PSE-1 was identified. All in all, KPC-2-producing CRKP ST11 and ST48 clone were widely disseminated in multiple departments of our hospital, which triggers the need for active surveillance and implementation of infection control measures.

The in vitro activity of polymyxin B and tigecycline alone and combination with other antibiotics against carbapenem-resistant Enterobacter cloacae complex isolates, including high-risk clones.

The emergence of carbapenem-resistant Enterobacteriaceae (CRE) has become a significant problem for global public health. Currently, treatments program is minimal. This study aimed to evaluate the molecular mechanisms of carbapenem-resistant Enterobacter cloacae complex isolates (CREC) infections. Methods: Resistance genes were detected using PCR with specific primers. Multilocus sequence typing (MLST) was also performed. Furthermore, we evaluated the effects of polymyxin B (PMB) and tigecycline (TGC) antibiotics (Abs) alone and in combination with meropenem (MEM), amikacin (AMK), and levofloxacin (LEV) against CREC isolates. The results were then compared with in vitro synergy testing results obtained from time-kill assays (TKAs), and the microdilution checkerboard method. The synergistic efficiency of PMB + TGC was also evaluated. Abs use clinically achievable concentrations to determine the antibacterial effects of the Ab. Similar sequence type (ST) classifications had a comparably resistant phenotype; PMB-based combination therapy is better than TGC-based combination therapy. we found that the combination of PMB + AMK is promising for the treatment of AMK-sensitive CREC. The high-risk ST93 carrying the bla KPC-2 gene should be monitored.