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1.
Proc Natl Acad Sci U S A ; 120(51): e2314135120, 2023 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-38096417

RESUMO

Conjugative plasmids play a key role in the dissemination of antimicrobial resistance (AMR) genes across bacterial pathogens. AMR plasmids are widespread in clinical settings, but their distribution is not random, and certain associations between plasmids and bacterial clones are particularly successful. For example, the globally spread carbapenem resistance plasmid pOXA-48 can use a wide range of enterobacterial species as hosts, but it is usually associated with a small number of specific Klebsiella pneumoniae clones. These successful associations represent an important threat for hospitalized patients. However, knowledge remains limited about the factors determining AMR plasmid distribution in clinically relevant bacteria. Here, we combined in vitro and in vivo experimental approaches to analyze pOXA-48-associated AMR levels and conjugation dynamics in a collection of wild-type enterobacterial strains isolated from hospitalized patients. Our results revealed significant variability in these traits across different bacterial hosts, with Klebsiella spp. strains showing higher pOXA-48-mediated AMR and conjugation frequencies than Escherichia coli strains. Using experimentally determined parameters, we developed a simple mathematical model to interrogate the contribution of AMR levels and conjugation permissiveness to plasmid distribution in bacterial communities. The simulations revealed that a small subset of clones, combining high AMR levels and conjugation permissiveness, play a critical role in stabilizing the plasmid in different polyclonal microbial communities. These results help to explain the preferential association of plasmid pOXA-48 with K. pneumoniae clones in clinical settings. More generally, our study reveals that species- and strain-specific variability in plasmid-associated phenotypes shape AMR evolution in clinically relevant bacterial communities.


Assuntos
Antibacterianos , Permissividade , Humanos , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Farmacorresistência Bacteriana/genética , Plasmídeos/genética , Klebsiella pneumoniae/genética , Klebsiella/genética , Escherichia coli/genética , Bactérias/genética
2.
J Antimicrob Chemother ; 79(6): 1432-1440, 2024 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-38708553

RESUMO

OBJECTIVES: Despite the introduction of cystic fibrosis transmembrane conductance regulator (CFTR) modulators, Pseudomonas aeruginosa is still a major pathogen in people with cystic fibrosis (pwCF). We determine the activity of cefiderocol and comparators in a collection of 154 P. aeruginosa isolates recovered from pwCF during three multicentre studies performed in 17 Spanish hospitals in 2013, 2017 and 2021. METHODS: ISO broth microdilution was performed and MICs were interpreted with CLSI and EUCAST criteria. Mutation frequency and WGS were also performed. RESULTS: Overall, 21.4% were MDR, 20.8% XDR and 1.3% pandrug-resistant (PDR). Up to 17% of the isolates showed a hypermutator phenotype. Cefiderocol demonstrated excellent activity; only 13 isolates (8.4%) were cefiderocol resistant by EUCAST (none using CLSI). A high proportion of the isolates resistant to ceftolozane/tazobactam (71.4%), meropenem/vaborbactam (70.0%), imipenem/relebactam (68.0%) and ceftazidime/avibactam (55.6%) were susceptible to cefiderocol. Nine out of 13 cefiderocol-resistant isolates were hypermutators (P < 0.001). Eighty-three STs were detected, with ST98 being the most frequent. Only one isolate belonging to the ST175 high-risk clone carried blaVIM-2. Exclusive mutations affecting genes involved in membrane permeability, AmpC overexpression (L320P-AmpC) and efflux pump up-regulation were found in cefiderocol-resistant isolates (MIC = 4-8 mg/L). Cefiderocol resistance could also be associated with mutations in genes related to iron uptake (tonB-dependent receptors and pyochelin/pyoverdine biosynthesis). CONCLUSIONS: Our results position cefiderocol as a therapeutic option in pwCF infected with P. aeruginosa resistant to most recent ß-lactam/ß-lactamase inhibitor combinations.


Assuntos
Antibacterianos , Cefiderocol , Cefalosporinas , Fibrose Cística , Testes de Sensibilidade Microbiana , Infecções por Pseudomonas , Pseudomonas aeruginosa , Humanos , Fibrose Cística/microbiologia , Fibrose Cística/complicações , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/isolamento & purificação , Infecções por Pseudomonas/microbiologia , Espanha/epidemiologia , Antibacterianos/farmacologia , Cefalosporinas/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Adolescente , Adulto , Criança , Mutação , Tazobactam/farmacologia , Feminino , Masculino
3.
J Antimicrob Chemother ; 79(9): 2132-2141, 2024 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-38988305

RESUMO

BACKGROUND: Carbapenem-resistant Enterobacterales (CRE) bloodstream infections (BSIs) are a major threat to patients. To date, data on risk factors have been limited, with low internal and external validity. In this multicentre study, risk factors for CRE BSI were determined by comparison with two control groups: patients with carbapenem-susceptible Enterobacterales (CSE) BSI, and patients without Enterobacterales infection (uninfected patients). METHODS: A multicentre, case-control-control study was nested in a European prospective cohort study on CRE (EURECA). CRE BSI:CSE BSI matching was 1:1, CRE BSI:Uninfected patients matching was 1:3, based on hospital, ward and length of stay. Conditional logistic regression was applied. RESULTS: From March 2016 to November 2018, 73 CRE BSIs, 73 CSE BSIs and 219 uninfected patients were included from 18 European hospitals. For CRE versus CSE BSI, previous CRE colonization/infection [incidence rate ratio (IRR) 7.32; 95% CI 1.65-32.38) increased the risk. For CRE versus uninfected controls, independent risk factors included: older age (IRR 1.03; 95% CI 1.01-1.06), patient referral (long-term care facility: IRR 7.19; 95% CI 1.51-34.24; acute care hospital: IRR 5.26; 95% CI 1.61-17.11), previous colonization/infection with other MDR organisms (MDROs) (IRR 9.71; 95% CI 2.33-40.56), haemodialysis (IRR 8.59; 95% CI 1.82-40.53), invasive procedures (IRR 5.66; 95% CI 2.11-15.16), and ß-lactam/ß-lactamase inhibitor combinations (IRR 3.92; 95% CI 1.68-9.13) or third/fourth generation cephalosporin (IRR 2.75; 95% CI 1.06-7.11) exposure within 3 months before enrolment. CONCLUSIONS: Evidence of previous CRE colonization/infection was a major risk factor for carbapenem resistance among Enterobacterales BSI. Compared with uninfected patients, evidence of previous MDRO colonization/infection and healthcare exposure were important risk factors for CRE BSI. Targeted screening, infection prevention and antimicrobial stewardship should focus on these high-risk patients.


Assuntos
Antibacterianos , Bacteriemia , Enterobacteriáceas Resistentes a Carbapenêmicos , Carbapenêmicos , Infecções por Enterobacteriaceae , Humanos , Estudos de Casos e Controles , Fatores de Risco , Masculino , Infecções por Enterobacteriaceae/epidemiologia , Infecções por Enterobacteriaceae/microbiologia , Feminino , Idoso , Pessoa de Meia-Idade , Estudos Prospectivos , Bacteriemia/microbiologia , Bacteriemia/epidemiologia , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Carbapenêmicos/farmacologia , Enterobacteriáceas Resistentes a Carbapenêmicos/efeitos dos fármacos , Enterobacteriáceas Resistentes a Carbapenêmicos/isolamento & purificação , Europa (Continente)/epidemiologia , Idoso de 80 Anos ou mais
4.
Eur J Clin Microbiol Infect Dis ; 43(7): 1349-1353, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38780755

RESUMO

INTRODUCTION: Burkholderia cepacia complex (BCC) are non-fermenting Gram-negative bacteria that can chronically colonize the lungs of people with cystic fibrosis (pwCF), causing a severe and progressive respiratory failure, post-transplant complications and epidemic outbreaks. Therefore, rapid and accurate identification of these bacteria is relevant for pwCF, in order to facilitate early eradication and prevent chronic colonization. However, BCCs are often quite difficult to detect on culture media as they have a slow growth rate and can be hidden by other fast-growing microorganisms, including Pseudomonas aeruginosa and filamentous fungi. MATERIAL AND METHODS: We evaluated the sensitivity of CHROMagar™ B. cepacia agar using 11 isolates from a well-characterized BCC collection, using BCA agar (Oxoid, UK) as a gold standard. We also studied 180 clinical sputum samples to calculate positive (PPV) and negative (NPV) predictive values. Furthermore, we used three of the well-characterized BCC isolates to determine the limit of detection (LOD). RESULTS: Eleven isolates grew on CHROMagar™ B. cepacia at 37ºC after 48 h. The NPV and PPV of CHROMagar™ B. cepacia were 100% and 87.5%, respectively. The LOD of CHROMagar™ B. cepacia was around 1 × 103 CFU/ml, requiring a ten-fold dilution lower bacterial load than BCA for BCC detection. CONCLUSION: CHROMagar™ B. cepacia agar proved to have a very good sensitivity and specificity for the detection of clinical BCCs. Moreover, the chromogenic nature of the medium allowed us to clearly differentiate BCC from other Gram-negative species, filamentous fungi and yeasts, thereby facilitating the identification of contaminants.


Assuntos
Ágar , Técnicas Bacteriológicas , Infecções por Burkholderia , Complexo Burkholderia cepacia , Meios de Cultura , Fibrose Cística , Sensibilidade e Especificidade , Escarro , Humanos , Fibrose Cística/microbiologia , Fibrose Cística/complicações , Complexo Burkholderia cepacia/isolamento & purificação , Complexo Burkholderia cepacia/classificação , Escarro/microbiologia , Infecções por Burkholderia/microbiologia , Infecções por Burkholderia/diagnóstico , Meios de Cultura/química , Técnicas Bacteriológicas/métodos
5.
Eur J Clin Microbiol Infect Dis ; 43(3): 605-610, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38112967

RESUMO

FASTinov® developed a rapid antimicrobial susceptibility test that includes the purification of a bacterial suspension directly from positive blood cultures (BC). In order to streamline laboratory workflow, the use of the bacterial suspension obtained through FASTinov® sample prep was tested for identification (ID) by matrix absorption laser deionization-time of flight mass spectrometry (MALDI-TOF MS) (Bruker) in 364 positive BC, and its accuracy assessed comparing with the MALDI-TOF MS ID of the next-day subcultured colonies. FASTinov sample prep was highly reliable for rapid ID directly from BC with proportion of agreement of 94.9% for Gram-positive and 96.3% for Gram-negative bacteria.


Assuntos
Bacteriemia , Hemocultura , Humanos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Bactérias , Bactérias Gram-Negativas , Laboratórios , Bacteriemia/microbiologia
6.
Eur J Clin Microbiol Infect Dis ; 43(2): 279-296, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38041722

RESUMO

PURPOSE: To characterize the resistance mechanisms affecting the cefepime-taniborbactam combination in a collection of carbapenemase-producing Enterobacterales (CPE) and carbapenem-resistant Pseudomonas spp. (predominantly P. aeruginosa; CRPA) clinical isolates. METHODS: CPE (n = 247) and CRPA (n = 170) isolates were prospectively collected from patients admitted to 8 Spanish hospitals. Susceptibility to cefepime-taniborbactam and comparators was determined by broth microdilution. Cefepime-taniborbactam was the most active agent, inhibiting 97.6% of CPE and 67.1% of CRPA (MICs ≤ 8/4 mg/L). All isolates with cefepime-taniborbactam MIC > 8/4 mg/L (5 CPE and 52 CRPA) and a subset with MIC ≤ 8/4 mg/L (23 CPE and 24 CRPA) were characterized by whole genome sequencing. RESULTS: A reduced cefepime-taniborbactam activity was found in two KPC-ST307-Klebsiella pneumoniae isolates with altered porins [KPC-62-K. pneumoniae (OmpA, OmpR/EnvZ), KPC-150-K. pneumoniae (OmpK35, OmpK36)] and one each ST133-VIM-1-Enterobacter hormaechei with altered OmpD, OmpR, and OmpC; IMP-8-ST24-Enterobacter asburiae; and NDM-5-Escherichia coli with an YRIN-inserted PBP3 and a mutated PBP2. Among the P. aeruginosa (68/76), elevated cefepime-taniborbactam MICs were mostly associated with GES-5-ST235, OXA-2+VIM-2-ST235, and OXA-2+VIM-20-ST175 isolates also carrying mutations in PBP3, efflux pump (mexR, mexZ) and AmpC (mpl) regulators, and non-carbapenemase-ST175 isolates with AmpD-T139M and PBP3-R504C mutations. Overall, accumulation of these mutations was frequently detected among non-carbapenemase producers. CONCLUSIONS: The reduced cefepime-taniborbactam activity among the minority of isolates with elevated cefepime-taniborbactam MICs is not only due to IMP carbapenemases but also to the accumulation of multiple resistance mechanisms, including PBP and porin mutations in CPE and chromosomal mutations leading to efflux pumps up-regulation, AmpC overexpression, and PBP modifications in P. aeruginosa.


Assuntos
Antibacterianos , Proteínas de Bactérias , Ácidos Borínicos , Carbapenêmicos , Ácidos Carboxílicos , Humanos , Cefepima/farmacologia , Carbapenêmicos/farmacologia , Antibacterianos/farmacologia , Pseudomonas/genética , Espanha/epidemiologia , beta-Lactamases/genética , Pseudomonas aeruginosa/genética , Testes de Sensibilidade Microbiana
7.
Infection ; 2024 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-38703288

RESUMO

BACKGROUND: Community-acquired (CA) and healthcare-associated (HCA) infections caused by carbapenemase-producing Enterobacterales (CPE) are not well characterized. The objective was to provide detailed information about the clinical and molecular epidemiological features of nosocomial, HCA and CA infections caused by carbapenemase-producing Klebsiella pneumoniae (CP-Kp) and Escherichia coli (CP-Ec). METHODS: A prospective cohort study was performed in 59 Spanish hospitals from February to March 2019, including the first 10 consecutive patients from whom CP-Kp or CP-Ec were isolated. Patients were stratified according to acquisition type. A multivariate analysis was performed to identify the impact of acquisition type in 30-day mortality. RESULTS: Overall, 386 patients were included (363 [94%] with CP-Kp and 23 [6%] CP-Ec); in 296 patients (76.3%), the CPE was causing an infection. Acquisition was CA in 31 (8.0%) patients, HCA in 183 (47.4%) and nosocomial in 172 (48.3%). Among patients with a HCA acquisition, 100 (54.6%) had been previously admitted to hospital and 71 (38.8%) were nursing home residents. Urinary tract infections accounted for 19/23 (82.6%), 89/130 (68.5%) and 42/143 (29.4%) of CA, HCA and nosocomial infections, respectively. Overall, 68 infections (23%) were bacteremia (8.7%, 17.7% and 30.1% of CA, HCA and nosocomial, respectively). Mortality in infections was 28% (13%, 14.6% and 42.7% of CA, HCA and nosocomial, respectively). Nosocomial bloodstream infections were associated with increased odds for mortality (adjusted OR, 4.00; 95%CI 1.21-13.19). CONCLUSIONS: HCA and CA infections caused by CPE are frequent and clinically significant. This information may be useful for a better understanding of the epidemiology of CPE.

8.
Int J Mol Sci ; 25(14)2024 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-39063130

RESUMO

Carbapenemase-producing Enterobacterales are increasingly being recognized in nosocomial infections. The performance of a flow cytometry-based rapid assay for their detection and differentiation was evaluated. This is a disruptive phenotypic technology, phenotypic and growth-independent, that searches for the lesions produced by drugs acting on cells after a short incubation time. Overall, 180 Gram-negative bacteria were studied, and results were compared with those obtained molecularly by PCR and phenotypically by 'KPC, MBL and OXA-48 Confirm Kit'. This phenotypic method was used as reference for comparison purposes. Susceptibility to carbapenems (imipenem, meropenem, and ertapenem) was determined by standard broth microdilution. Overall, 112 isolates (62.2%) were carbapenemase producers, 41 KPCs, 36 MßLs, and 31 OXA-48, and 4 strains were KPC + MßL co-producers. Sixty-eight isolates were carbapenemase-negative. The percentage of agreement, sensitivity, and specificity were calculated according to ISO 20776-2:2021. The FASTinov assay showed 97.7% agreement with the reference method for carbapenemase detection. Discrepant flow cytometry results were obtained in four isolates compared with both reference and PCR results. The sensitivity and specificity of this new technology were 95.3% and 98.5%, respectively, for KPCs, 97.6% and 99.3% for MßLs, and 96.9% and 98% for OXA-48 detection. In conclusion, we describe a rapid flow cytometry assay with high accuracy for carbapenemase detection and the differentiation of various carbapenemases, which should impact clinical microbiology laboratories and patient management.


Assuntos
Proteínas de Bactérias , Citometria de Fluxo , Testes de Sensibilidade Microbiana , beta-Lactamases , beta-Lactamases/metabolismo , Citometria de Fluxo/métodos , Proteínas de Bactérias/metabolismo , Humanos , Testes de Sensibilidade Microbiana/métodos , Enterobacteriaceae/efeitos dos fármacos , Enterobacteriaceae/enzimologia , Antibacterianos/farmacologia , Sensibilidade e Especificidade , Carbapenêmicos/farmacologia
9.
Infect Immun ; 91(2): e0001223, 2023 02 16.
Artigo em Inglês | MEDLINE | ID: mdl-36722977

RESUMO

Colistin resistance is acquired by different lipopolysaccharide (LPS) modifications. We proposed to evaluate the of effect in vivo colistin resistance acquisition on the innate immune response. We used a pair of ST11 clone Klebsiella pneumoniae strains: an OXA-48, CTX-M-15 K. pneumoniae strain susceptible to colistin (CS-Kp) isolated from a urinary infection and its colistin-resistant variant (CR-Kp) from the same patient after prolonged treatment with colistin. No mutation of previously described genes for colistin resistance (pmrA, pmrB, mgrB, phoP/Q, arnA, arnC, arnT, ugdH, and crrAB) was found in the CR-Kp genome; however, LPS modifications were characterized by negative-ion matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. The strains were cocultured with human monocytes to determine their survival after phagocytosis and induction to apoptosis. Also, monocytes were stimulated with bacterial LPS to study cytokine and immune checkpoint production. The addition of 4-amino-4-deoxy-l-arabinose (Ara4N) to lipid A of CR-Kp accounted for the colistin resistance. CR-Kp survived significantly longer inside human monocytes after being phagocytosed than did the CS-Kp strain. In addition, LPS from CR-Kp induced both higher apoptosis in monocytes and higher levels of cytokine and immune checkpoint production than LPS from CS-Kp. Our data reveal a variable impact of colistin resistance on the innate immune system, depending on the responsible mechanism. Adding Ara4N to LPS in K. pneumoniae increases bacterial survival after phagocytosis and elicits a higher inflammatory response than its colistin-susceptible counterpart.


Assuntos
Colistina , Infecções por Klebsiella , Humanos , Colistina/farmacologia , Lipopolissacarídeos/farmacologia , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Imunidade Inata , Klebsiella pneumoniae , Citocinas , Infecções por Klebsiella/microbiologia , Farmacorresistência Bacteriana/genética , Testes de Sensibilidade Microbiana
10.
J Clin Microbiol ; 61(4): e0104922, 2023 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-37014210

RESUMO

The Enterobacter cloacae complex (ECC) encompasses heterogeneous clusters of species that have been associated with nosocomial outbreaks. These species may have different acquired antimicrobial resistance and virulence mechanisms, and their identification is challenging. This study aims to develop predictive models based on matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) profiles and machine learning for species-level identification. A total of 219 ECC and 118 Klebsiella aerogenes clinical isolates from three hospitals were included. The capability of the proposed method to differentiate the most common ECC species (Enterobacter asburiae, Enterobacter kobei, Enterobacter hormaechei, Enterobacter roggenkampii, Enterobacter ludwigii, and Enterobacter bugandensis) and K. aerogenes was demonstrated by applying unsupervised hierarchical clustering with principal-component analysis (PCA) preprocessing. We observed a distinctive clustering of E. hormaechei and K. aerogenes and a clear trend for the rest of the ECC species to be differentiated over the development data set. Thus, we developed supervised, nonlinear predictive models (support vector machine with radial basis function and random forest). The external validation of these models with protein spectra from two participating hospitals yielded 100% correct species-level assignment for E. asburiae, E. kobei, and E. roggenkampii and between 91.2% and 98.0% for the remaining ECC species; with data analyzed in the three participating centers, the accuracy was close to 100%. Similar results were obtained with the Mass Spectrometric Identification (MSI) database developed recently (https://msi.happy-dev.fr) except in the case of E. hormaechei, which was more accurately identified with the random forest algorithm. In short, MALDI-TOF MS combined with machine learning was demonstrated to be a rapid and accurate method for the differentiation of ECC species.


Assuntos
Algoritmos , Enterobacter cloacae , Humanos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos
11.
J Antimicrob Chemother ; 78(5): 1259-1264, 2023 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-36964710

RESUMO

OBJECTIVES: Ceftazidime/avibactam and cefiderocol are two of the latest antibiotics with activity against a wide variety of Gram-negatives, including carbapenem-resistant Enterobacterales. We sought to describe the phenotypic and genotypic characteristics of ceftazidime/avibactam- and cefiderocol-resistant KPC-Klebsiella pneumoniae (KPC-Kp) detected during an outbreak in 2020 in the medical ICU of our hospital. METHODS: We collected 11 KPC-Kp isolates (6 clinical; 5 surveillance samples) resistant to ceftazidime/avibactam and cefiderocol from four ICU patients (November 2020 to January 2021), without prior exposure to these agents. All patients had a decontamination regimen as part of the standard ICU infection prevention protocol. Additionally, one ceftazidime/avibactam- and cefiderocol-resistant KPC-Kp (June 2019) was retrospectively recovered. Antibiotic susceptibility was determined by broth microdilution. ß-Lactamases were characterized and confirmed. WGS was also performed. RESULTS: All KPC-Kp isolates (ceftazidime/avibactam MIC  ≥16/4 mg/L; cefiderocol MIC ≥4 mg/L) were KPC + CTX-M-15 producers and belonged to the ST307 high-risk-clone (ST307-HRC). KPC-62 (L168Q) was detected in all isolates involved in the 2020 outbreak, contained in January 2021. KPC-31 (D179Y) was identified in the KPC-Kp from 2019. Cloning experiments demonstrated that both blaKPC-62 and blaKPC-31 were responsible for ceftazidime/avibactam resistance (MIC >16 mg/L) and an increased cefiderocol MIC. Additionally, mutations in OmpA and EnvZ/OmpR porin proteins (in KPC-62-Kp) and in PBP2 (in KPC-31-Kp) were found and may be involved in cefiderocol resistance. CONCLUSIONS: The emergence of resistance to both ceftazidime/avibactam and cefiderocol in KPC-Kp-HRCs, together with the diversification of novel KPC enzymes displaying different antibiotic resistance phenotypes, is an epidemiological and clinical risk.


Assuntos
Ceftazidima , Infecções por Klebsiella , Humanos , Ceftazidima/farmacologia , Ceftazidima/uso terapêutico , Klebsiella pneumoniae , Espanha/epidemiologia , Estudos Retrospectivos , Infecções por Klebsiella/epidemiologia , Infecções por Klebsiella/tratamento farmacológico , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Hospitais Universitários , Cefiderocol
12.
J Antimicrob Chemother ; 78(9): 2291-2296, 2023 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-37533351

RESUMO

OBJECTIVES: To assess the microbiological characteristics of Escherichia coli causing healthcare-associated bacteraemia of urinary origin (HCA-BUO) in Spain (ITUBRAS-2 project), with particular focus on ESBL producers and isolates belonging to ST131 high-risk clone (HiRC). Clinical characteristics and outcomes associated with ST131 infection were investigated. METHODS: A total of 222 E. coli blood isolates were prospectively collected from patients with HCA-BUO from 12 tertiary-care hospitals in Spain (2017-19). Antimicrobial susceptibility and ESBL/carbapenemase production were determined. ST131 subtyping was performed. A subset of 115 isolates were selected for WGS to determine population structure, resistome and virulome. Clinical charts were reviewed. RESULTS: ESBL-producing E. coli prevalence was 30.6% (68/222). ST131 represented 29.7% (66/222) of E. coli isolates and accounted for the majority of ESBL producers (46/68, 67.6%). The C2/H30-Rx subclone accounted for most ST131 isolates (44/66) and was associated with CTX-M-15 (37/44) and OXA-1 enzymes (27/44). Cluster C1-M27 was identified in 4/10 isolates belonging to subclade C1/H30-R1 and associated with CTX-M-27. Additionally, ST131 isolates showed a high content of other acquired resistance genes, and clade C/ST131 isolates carried characteristic QRDR mutations. They were categorized as uropathogenic E. coli and had higher aggregate virulence scores. ST131 infection was associated with more complex patients, prior use of cephalosporins and inadequate empirical treatment but was not associated with worse clinical outcomes. CONCLUSIONS: ST131 HiRC is the main driver of ESBL-producing E. coli causing HCA-BUO in Spain, mainly associated with the expansion of subclade CTX-M-15-C2/H30-Rx and the emergence of CTX-M-27-C1/H30-R1 (Cluster C1-M27).


Assuntos
Bacteriemia , Infecções por Escherichia coli , Humanos , Escherichia coli , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/microbiologia , Espanha/epidemiologia , Epidemiologia Molecular , Genótipo , Bacteriemia/epidemiologia , beta-Lactamases/genética , Atenção à Saúde
13.
Antimicrob Agents Chemother ; 66(3): e0216121, 2022 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-35007130

RESUMO

Novel ß-lactam-ß-lactamase inhibitor combinations currently approved for clinical use are poorly active against metallo-ß-lactamase (MBL)-producing strains. We evaluated the in vitro activity of cefepime-taniborbactam (FTB [formerly cefepime-VNRX-5133]) and comparator agents against carbapenemase-producing Enterobacterales (n = 247) and carbapenem-resistant Pseudomonas species (n = 170) clinical isolates prospectively collected from different clinical origins in patients admitted to 8 Spanish hospitals. FTB was the most active agent in both Enterobacterales (97.6% MICFTB, ≤8/4 mg/L) and Pseudomonas (67.1% MICFTB, ≤8/4 mg/L) populations. The MICFTB was >8 mg/L in 6/247 (2.4%) Enterobacterales isolates (3 KPC-producing Klebsiella pneumoniae isolates, 1 VIM-producing Enterobacter cloacae isolate, 1 IMP-producing E. cloacae isolate, and 1 NDM-producing Escherichia coli isolate) and in 56/170 (32.9%) Pseudomonas isolates, 19 of them carbapenemase producers (15 producers of VIM, 2 of GES, 1 of GES+VIM, and 1 of GES+KPC). Against the Enterobacterales isolates with meropenem MICs of >2 mg/L (138/247), FTB was the most active agent against both serine-ß-lactamases (107/138) and MBL producers (31/138) (97.2 and 93.5% MICFTB, ≤8/4 mg/L, respectively), whereas the activity of comparators was reduced, particularly against the MBL producers (ceftazidime-avibactam, 94.4 and 12.9%, meropenem-vaborbactam, 85.0 and 64.5%, imipenem-relebactam, 76.6 and 9.7%, ceftolozane-tazobactam, 1.9 and 0%, and piperacillin-tazobactam, 0 and 0%, respectively). Among the meropenem-resistant Pseudomonas isolates (163/170; MIC, >2 mg/L), the activities of FTB against serine-ß-lactamase (35/163) and MBL (43/163) producers were 88.6 and 65.1%, respectively, whereas the susceptibilities of comparators were as follows: ceftazidime-avibactam, 88.5 and 16.0%, meropenem-vaborbactam, 8.5 and 7.0%, imipenem-relebactam, 2.9 and 2.3%, ceftolozane-tazobactam, 0 and 2.3%, and piperacillin-tazobactam, 0 and 0%, respectively. Microbiological results suggest FTB as a potential therapeutic option in patients infected with carbapenemase-producing Enterobacterales and carbapenem-resistant Pseudomonas isolates, including MBL producers.


Assuntos
Pseudomonas aeruginosa , beta-Lactamases , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Compostos Azabicíclicos/farmacologia , Proteínas de Bactérias , Ácidos Borínicos , Ácidos Carboxílicos , Cefepima/farmacologia , Humanos , Testes de Sensibilidade Microbiana , Espanha
14.
J Clin Microbiol ; 60(3): e0027621, 2022 03 16.
Artigo em Inglês | MEDLINE | ID: mdl-34346716

RESUMO

The European Committee on Antimicrobial Susceptibility Testing (EUCAST) is an international susceptibility testing committee, organized by the European Society for Clinical Microbiology and Infectious Diseases (ESCMID) and functioning as the breakpoint advisory committee of the European Medicines Agency (EMA). The original remit of EUCAST was to harmonize European clinical breakpoints, but very soon, the activities expanded beyond the borders of Europe and included newly licensed agents in Europe. Among the milestones were the aggregating of large numbers of MIC distributions, creating software to display these distributions, the EUCAST concept of identifying epidemiological cutoff values (ECOFF), and the development of a EUCAST disk diffusion method. The EUCAST Development Laboratory has played a critical role in the development of antimicrobial susceptibility testing (AST) methodology, including development work for novel antimicrobial agents and for rapid AST directly from blood culture bottles. EUCAST has several standing subcommittees, including for AST in fungi (AFST) and mycobacteria (AMST) and for microorganisms of veterinary interest (VetCAST), and ad hoc subcommittees on subjects such as anaerobic bacteria, MIC and zone diameter distributions and epidemiological cutoff values, the relationship between phenotypic and genotypic resistance, and expert rules and methods for the detection of resistance mechanisms. All EUCAST decisions are subjected to the EUCAST public consultation process, the only exception being breakpoints of novel antimicrobial agents where confidentiality agreements during the licensing process prevent public participation. EUCAST has recently revised the definitions of clinical susceptibility interpretive categories S, I, and R, acknowledging the intimate relationship between drug exposure and susceptibility reporting.


Assuntos
Anti-Infecciosos , Fungos , Antibacterianos/farmacologia , Europa (Continente) , Humanos , Testes de Sensibilidade Microbiana
15.
J Clin Microbiol ; 60(3): e0224521, 2022 03 16.
Artigo em Inglês | MEDLINE | ID: mdl-35107303

RESUMO

The emergence of Klebsiella pneumoniae isolates carrying novel blaKPC variants conferring ceftazidime-avibactam (CAZ/AVI) resistance is being increasingly reported. We evaluated the accuracy of phenotypic methods commonly used in routine clinical laboratories in the detection of novel K. pneumoniae carbapenemase (KPC) enzymes. Additionally, we characterized by whole-genome sequencing (WGS) the KPC-ST307-K. pneumoniae isolates recovered in our hospital before and after CAZ/AVI therapy. Rectal colonization or infection by carbapenem-resistant KPC-3 K. pneumoniae isolates (imipenem MIC, 16 mg/L; meropenem MIC, 8 to >16 mg/L) and CAZ/AVI-susceptible isolates (CAZ/AVI MIC, 1 to 2 mg/L) were first detected in three intensive care unit (ICU) patients admitted between March 2020 and July 2020. KPC K. pneumoniae isolates with increased CAZ/AVI MICs (8 to 32 mg/L) and carbapenem susceptibility (imipenem and meropenem MIC, <1 mg/L) were recovered within 6 to 24 days after CAZ/AVI treatment. WGS confirmed that all KPC K. pneumoniae isolates belonged to the sequence type 307 (ST307) high-risk clone and carried identical antimicrobial resistance genes and virulence factors. The presence of the novel blaKPC-46, blaKPC-66, and blaKPC-92 genes was confirmed in the K. pneumoniae isolates with increased CAZ/AVI MICs and restored carbapenem activity. KPC production was confirmed by immunochromatography, the eazyplex Superbug CRE system, and the Xpert Carba-R assay in all KPC K. pneumoniae isolates, but not in any isolate using chromogenic agar plates for carbapenemase producers (ChromID-CARBA), the KPC/MBL/OXA-48 Confirm kit, and the ß-CARBA test. Nevertheless, all grew in chromogenic agar plates for extended-spectrum ß-lactamase (ESBL) producers (ChromID-ESBL). We report the failure of the most common phenotypic methods used for the detection of novel KPC carbapenemases but not of rapid molecular or immunochromatography assays, thus highlighting their relevance in microbiology laboratories.


Assuntos
Infecções por Klebsiella , Klebsiella pneumoniae , Ágar , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Compostos Azabicíclicos , Proteínas de Bactérias/genética , Carbapenêmicos/uso terapêutico , Ceftazidima/farmacologia , Células Clonais , Combinação de Medicamentos , Humanos , Imipenem/uso terapêutico , Infecções por Klebsiella/tratamento farmacológico , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/genética , Meropeném , Testes de Sensibilidade Microbiana , beta-Lactamases/genética
16.
J Antimicrob Chemother ; 77(Suppl_1): i2-i9, 2022 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-36065724

RESUMO

BACKGROUND: In the late 1990s, as a response to rising antimicrobial resistance (AMR), an independent multinational, interdisciplinary group was formed specifically targeting primary care antibiotic prescribing for community-acquired respiratory tract infections (CA-RTIs). The group comprised senior clinicians from Canada, Israel, Spain, Sweden, UK and USA. The group's objectives were to provide recommendations for antibiotic stewardship in the community because, whilst it was widely accepted that inappropriate antibiotic use was contributing to AMR, it remained difficult to change prescribing behaviour. The group aimed to identify principles underlying appropriate antibiotic prescribing and guideline formulation to reduce morbidity from CA-RTIs, limit therapeutic failure and, importantly, curb AMR emergence. The group published a report in 2002, which has become known as the Consensus Principles. OBJECTIVES: (i) To consider the relevance of the Consensus Principles in 2022 by reviewing current global approaches to rising AMR. A wide range of factors, such as antibiotic overuse, most recently seen in COVID-19 patients, are still driving rising AMR even though there has been a high-level international response to the AMR threat; and (ii) as an introduction to this Supplement, which reports the findings of analyses of how AMR is being addressed in nine disparate countries (Brazil, India, Kuwait, Mexico, Pakistan, Russia, Saudi Arabia, Türkiye and Vietnam). Understanding how these initiatives are being pursued in different countries helps identify areas where more information is needed. CONCLUSIONS: Adherence to the Consensus Principles remains as important now as it was in 2002. Achieving appropriate antibiotic prescribing is a vital objective in order that the right patient receives the right antibiotics at the right time to ensure optimal clinical outcomes while at the same time helping to limit further increases in AMR.


Assuntos
Gestão de Antimicrobianos , COVID-19 , Infecções Comunitárias Adquiridas , Infecções Respiratórias , Antibacterianos/uso terapêutico , Infecções Comunitárias Adquiridas/tratamento farmacológico , Consenso , Humanos , Prescrição Inadequada , Infecções Respiratórias/tratamento farmacológico
17.
J Antimicrob Chemother ; 77(11): 3009-3015, 2022 10 28.
Artigo em Inglês | MEDLINE | ID: mdl-35971566

RESUMO

OBJECTIVES: To detect a potential hidden dissemination of the blaOXA-48 gene among Proteus mirabilis isolates obtained from a single centre. METHODS: P. mirabilis from diverse clinical samples presenting an ESBL phenotype or obtained from blood cultured from 2017 to 2019 were evaluated. Bacterial identification was performed using MALDI-TOF MS. MICs were determined using International Organization for Standardization (ISO) standard microdilution and interpreted following EUCAST guidelines. WGS was performed using both short- and long-read technologies and assemblies were done using Unicycler. Resistomes were assessed using the ResFinder database. SNPs were detected using the PATRIC bioinformatics platform. Cloning experiments were performed using the pCRII-TOPO cloning kit. RESULTS: Thirty-one out of 108 (28.7%) isolates were positive for blaOXA-48 and blaCTX-M-15. Twenty-nine out of 31 of the isolates were susceptible to temocillin, piperacillin/tazobactam, ertapenem and meropenem, whereas only 2/31 showed a resistance phenotype against these antibiotics. Both blaOXA-48 and blaCTX-M-15 genes were detected within the same chromosomally integrated new transposon in all isolates. The resistant isolates displayed a single mutation located in the putative promoter upstream of blaOXA-48. Cloning experiments confirmed that the mutation was responsible for the resistance phenotype. CONCLUSIONS: The presence of a chromosomal copy of blaOXA-48 did not confer resistance to carbapenems, but a single mutation in the promoter could lead to an increase in resistance. This study shows a hidden circulation of OXA-48-positive, but carbapenem- and piperacillin/tazobactam-susceptible, P. mirabilis isolates that can become resistant to ß-lactams after a single mutation.


Assuntos
Carbapenêmicos , Proteus mirabilis , Carbapenêmicos/farmacologia , Proteus mirabilis/genética , beta-Lactamases/genética , Testes de Sensibilidade Microbiana , Antibacterianos/farmacologia , Combinação Piperacilina e Tazobactam
18.
J Antimicrob Chemother ; 77(11): 3163-3172, 2022 10 28.
Artigo em Inglês | MEDLINE | ID: mdl-36059128

RESUMO

OBJECTIVES: To study the in vitro activity of imipenem/relebactam and comparators and the imipenem/relebactam resistance mechanisms in a Pseudomonas aeruginosa collection from Portugal (STEP, 2017-18) and Spain (SUPERIOR, 2016-17) surveillance studies. METHODS: P. aeruginosa isolates (n = 474) were prospectively recovered from complicated urinary tract (cUTI), complicated intra-abdominal (cIAI) and lower respiratory tract (LRTI) infections in 11 Portuguese and 8 Spanish ICUs. MICs were determined (ISO broth microdilution). All imipenem/relebactam-resistant P. aeruginosa isolates (n = 30) and a subset of imipenem/relebactam-susceptible strains (n = 32) were characterized by WGS. RESULTS: Imipenem/relebactam (93.7% susceptible), ceftazidime/avibactam (93.5% susceptible) and ceftolozane/tazobactam (93.2% susceptible) displayed comparable activity. The imipenem/relebactam resistance rate was 6.3% (Portugal 5.8%; Spain 8.9%). Relebactam restored imipenem susceptibility to 76.9% (103/134) of imipenem-resistant isolates, including MDR (82.1%; 32/39), XDR (68.8%; 53/77) and difficult-to-treat (DTR) isolates (67.2%; 45/67). Among sequenced strains, differences in population structure were detected depending on the country: clonal complex (CC)175 and CC309 in Spain and CC235, CC244, CC348 and CC253 in Portugal. Different carbapenemase gene distributions were also found: VIM-20 (n = 3), VIM-1 (n = 2), VIM-2 (n = 1) and VIM-36 (n = 1) in Spain and GES-13 (n = 13), VIM-2 (n = 3) and KPC-3 (n = 2) in Portugal. GES-13-CC235 (n = 13) and VIM type-CC175 (n = 5) associations were predominant in Portugal and Spain, respectively. Imipenem/relebactam showed activity against KPC-3 strains (2/2), but was inactive against all GES-13 producers and most of the VIM producers (8/10). Mutations in genes affecting porin inactivation, efflux pump overexpression and LPS modification might also be involved in imipenem/relebactam resistance. CONCLUSIONS: Microbiological results reinforce imipenem/relebactam as a potential option to treat cUTI, cIAI and LRTI caused by MDR/XDR P. aeruginosa isolates, except for GES-13 and VIM producers.


Assuntos
Infecções por Pseudomonas , Infecções Respiratórias , Humanos , Pseudomonas aeruginosa/genética , Portugal , Infecções por Pseudomonas/microbiologia , Espanha , Compostos Azabicíclicos/farmacologia , Imipenem/farmacologia , Antibacterianos/farmacologia , Testes de Sensibilidade Microbiana , Unidades de Terapia Intensiva , Infecções Respiratórias/microbiologia
19.
BMC Microbiol ; 22(1): 129, 2022 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-35549675

RESUMO

BACKGROUND: The potential pathogenic role of Stenotrophomonas maltophilia in lung disease and in particular in cystic fibrosis is unclear. To develop further understanding of the biology of this taxa, the taxonomic position, antibiotic resistance and virulence factors of S. maltophilia isolates from patients with chronic lung disease were studied. RESULTS: A total of 111 isolates recovered between 2003 and 2016 from respiratory samples from patients in five different countries were included. Based on a cut-off of 95%, analysis of average nucleotide identity by BLAST (ANIb) showed that the 111 isolates identified as S. maltophilia by Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF/MS) belonged to S. maltophilia (n = 65), S. pavanii (n = 6) and 13 putative novel species (n = 40), which each included 1-5 isolates; these groupings coincided with the results of the 16S rDNA analysis, and the L1 and L2 ß-lactamase Neighbor-Joining phylogeny. Chromosomally encoded aminoglycoside resistance was identified in all S. maltophilia and S. pavani isolates, while acquired antibiotic resistance genes were present in only a few isolates. Nevertheless, phenotypic resistance levels against commonly used antibiotics, determined by standard broth microbroth dilution, were high. Although putative virulence genes were present in all isolates, the percentage of positive isolates varied. The Xps II secretion system responsible for the secretion of the StmPr1-3 proteases was mainly limited to isolates identified as S. maltophilia based on ANIb, but no correlation with phenotypic expression of protease activity was found. The RPF two-component quorum sensing system involved in virulence and antibiotic resistance expression has two main variants with one variant lacking 190 amino acids in the sensing region. CONCLUSIONS: The putative novel Stenotrophomonas species recovered from patient samples and identified by MALDI-TOF/MS as S. maltophilia, differed from S. maltophilia in resistance and virulence genes, and therefore possibly in pathogenicity. Revision of the Stenotrophomonas taxonomy is needed in order to reliably identify strains within the genus and elucidate the role of the different species in disease.


Assuntos
Fibrose Cística , Infecções por Bactérias Gram-Negativas , Infecções Respiratórias , Stenotrophomonas maltophilia , Antibacterianos/farmacologia , Resistência Microbiana a Medicamentos , Humanos , Stenotrophomonas , Fatores de Virulência/genética
20.
Sens Actuators B Chem ; 369: 132217, 2022 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-35755181

RESUMO

The development of DNA-sensing platforms based on new synthetized Methylene Blue functionalized carbon nanodots combined with different shape gold nanostructures (AuNs), as a new pathway to develop a selective and sensitive methodology for SARS-CoV-2 detection is presented. A mixture of gold nanoparticles and gold nanotriangles have been synthetized to modify disposable electrodes that act as an enhanced nanostructured electrochemical surface for DNA probe immobilization. On the other hand, modified carbon nanodots prepared a la carte to contain Methylene Blue (MB-CDs) are used as electrochemical indicators of the hybridization event. These MB-CDs, due to their structure, are able to interact differently with double and single-stranded DNA molecules. Based on this strategy, target sequences of the SARS-CoV-2 virus have been detected in a straightforward way and rapidly with a detection limit of 2.00 aM. Moreover, this platform allows the detection of the SARS-CoV-2 sequence in the presence of other viruses, and also a single nucleotide polymorphism (SNPs). The developed approach has been tested directly on RNA obtained from nasopharyngeal samples from COVID-19 patients, avoiding any amplification process. The results agree well with those obtained by RT-qPCR or reverse transcription quantitative polymerase chain reaction technique.

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