RESUMO
The genomic comparison of two Klebsiella michiganensis clinical isolates recovered from the same patient, one resistant to piperacillin-tazobactam and intermediate to cefotaxime, the other resistant to ceftazidime but susceptible to piperacillin-tazobactam, revealed one mutation in the blaOXY-1-24 gene accounting for a L169M substitution in the Ω loop. Cloning experiment in Escherichia coli demonstrated the contribution of this mutation to the hydrolysis spectrum extension towards ceftazidime and cefepime, whereas the resistance to piperacillin-tazobactam was reduced. To the best of our knowledge, this study shows for the first time that ceftazidime resistance can occur in vivo from OXY-1 precursor by structural alteration.
Assuntos
Antibacterianos , Cefiderocol , Cefalosporinas , Infecções por Klebsiella , Klebsiella pneumoniae , Testes de Sensibilidade Microbiana , beta-Lactamases , Humanos , Antibacterianos/farmacologia , beta-Lactamases/genética , Cefalosporinas/farmacologia , Cromossomos Bacterianos/genética , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/enzimologia , Klebsiella pneumoniae/isolamento & purificaçãoRESUMO
OBJECTIVES: To identify the genetic change responsible for resistance to penicillins, extended-spectrum cephalosporins (ESCs), aminoglycosides and ciprofloxacin in a Serratia marcescens clinical isolate recovered from a pancreatic abscess 6â weeks after a WT strain was isolated from the same patient. The impact on the fitness was also assessed. METHODS: The genomes of both S. marcescens isolates were sequenced using Illumina technology, assembled, annotated and compared with each other. PCR amplification followed by Sanger sequencing was carried out to confirm the mutation. Complementation of the resistant isolate with a recombinant plasmid harbouring the WT gene was performed. The growth rates were measured for both isolates in LB medium. RESULTS: Comparative genomic analysis disclosed only one frameshift mutation (690delG) in the cpxA gene, which codes for the histidine kinase of a two-component system (TCS). This change introduced a premature termination codon, leading to the truncated CpxA_HatR variant that contained 234 amino acids instead of 464. Complementation, which consisted of transfer of the WT cpxA into the resistant S. marcescens derivative, restored completely its susceptibility to ESCs, aminoglycosides and ciprofloxacin, thus confirming the contribution of the CpxA_HatR variant to resistance. Growth analysis showed that the fitness of the resistant isolate was unchanged. CONCLUSIONS: This study shows for the first time that constitutive activation of the Cpx pathway can per se confer resistance to ESCs and ciprofloxacin, in addition to the aminoglycoside resistance usually described. It sheds new light on the role of altered TCSs in fostering bacterial survival.
Assuntos
Mutação da Fase de Leitura , Serratia marcescens , Aminoglicosídeos , Antibacterianos/farmacologia , Ciprofloxacina/farmacologia , Resistência a Medicamentos , HumanosRESUMO
The Proteus mirabilis PmirS clinical isolate, which was susceptible to imipenem (0.5 µg/mL) and amikacin (1 µg/mL), was recovered from a bronchial aspirate of a patient who recently underwent lung transplantation. The P. mirabilis PmirR clinical isolate, which exhibited resistance to imipenem (16 µg/mL) and amikacin (24 µg/mL), was isolated 3 weeks later from the same patient and the same specimen type. Using short-read sequencing technology, these isolates appeared to be genetically identical except the cpxA gene of the PmirR isolate that was mutated leading to the His-208-Pro substitution. The structural alteration was localized in the histidine kinase, adenylate cyclase, methyl accepting protein, phosphatase (HAMP) domain, which is involved in the signal transduction between the sensor kinase and the regulator response of the CpxA/CpxR two-component system (TCS). No significant defect in the growth rate was found between the PmirS and PmirR isolates. This study suggests that alteration in CpxA might confer imipenem and amikacin resistance in P. mirabilis. This study brings new evidence that the TCS alteration could provide an adaptive capacity in a clinical context by conferring antibiotic resistance without fitness cost.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Proteínas Quinases/genética , Proteus mirabilis/efeitos dos fármacos , Proteus mirabilis/genética , Amicacina/farmacologia , Humanos , Imipenem/farmacologia , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-IdadeRESUMO
Alteration in two-component systems (TCSs), which are signal transduction pathways in prokaryotes, can result in antibiotic resistance. Recently, it has been shown that the overexpression, using a multicopy cloning vector, of the dcuR, rcsB, and yehT genes, which code for the response regulator (RR) part of TCSs, enhanced the minimal inhibitory concentrations (MICs) of carbapenems in Escherichia coli K-12 derivative KAM3. Herein, the contribution to carbapenem resistance of the DcuS/DcuR, RcsC/RcsB, and YehU/YehT TCSs was assessed in E. coli K-12 derivative BW25113 (A phylogroup) and 536 (B2 phylogroup) recipient strains in combination with extended-spectrum ß-lactamase that exhibit a weak carbapenemase activity. The genes encoding both the sensor kinase (SK) and the RR, on the one hand, and the genes encoding the SK only, on the other hand, of these regulating pathways were disrupted. Subsequently, the mutants and their parental strains were transformed by a recombinant plasmid encoding the CTX-M-15 gene, before testing their susceptibility to carbapenems and their fitness. Results showed a trade-off between enhanced MICs for ertapenem, which remained above the clinical resistance breakpoint, and decreased growth rate, specifically for the 536 strain SK mutants. In conclusion, mutations in dcuS/dcuR, rcsC/rcsB, and yehU/yehT genes may be a pivotal first-step event in the development of carbapenem resistance.
Assuntos
Carbapenêmicos/farmacologia , Escherichia coli K12/efeitos dos fármacos , Escherichia coli K12/metabolismo , Proteínas de Escherichia coli/metabolismo , Antibacterianos/farmacologia , Proteínas de Bactérias/metabolismo , Proteínas de Ligação a DNA/metabolismo , Resistência Microbiana a Medicamentos/fisiologia , Ertapenem/farmacologia , Infecções por Escherichia coli/tratamento farmacológico , Infecções por Escherichia coli/microbiologia , Humanos , Testes de Sensibilidade Microbiana/métodos , Complexos Multienzimáticos/metabolismo , Fosfoproteínas Fosfatases/metabolismo , Proteínas Quinases/metabolismo , Fatores de Transcrição/metabolismo , beta-Lactamases/metabolismoRESUMO
Objectives: The whole-genome sequence (WGS) of Klebsiella pneumoniae KP3771 isolate was characterized. This strain was recovered from the urine sample of an 80-year-old man hospitalized in an intensive care unit of the University Hospital Tahar Sfar in Tunisia. Materials and Methods: WGS using a MiSeq platform was used. The assembled genome was subjected to several software analyses. Results: K. pneumoniae KP3771 was resistant to all antibiotics but colistin and tigecycline. WGS analysis found 18 transmissible genes encoding resistance markers, including blaNDM-1 and blaCTX-M-15 genes, which were carried by four plasmids belonging to the Inc Ib, IIk, and R groups. Three families of genes encoding virulence factors were detected, including adhesins (fimH, fimA, fimB, fimC, mrkD, Kpn, and ycfM), siderophores (enterobactin, aerobactin, and yersiniabactin siderophores), and protectin/invasin (traT). The strain was assigned to the sequence type 147. Conclusions: This study describes the genome of a carbapenemase-producing K. pneumoniae clinical isolate recovered in Tunisia. Bacteria WGS has become the reference technology to address epidemiological issues; this high level of information is particularly well suited to enrich epidemiological workflows' output.
Assuntos
Farmacorresistência Bacteriana Múltipla/genética , Genoma Bacteriano , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/genética , beta-Lactamases/genética , Adesinas Bacterianas/genética , Adesinas Bacterianas/metabolismo , Antibacterianos/farmacologia , Colistina/farmacologia , Feminino , Expressão Gênica , Hospitais , Humanos , Infecções por Klebsiella/tratamento farmacológico , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/isolamento & purificação , Masculino , Testes de Sensibilidade Microbiana , Plasmídeos/química , Plasmídeos/metabolismo , Sideróforos/biossíntese , Tigeciclina/farmacologia , Tunísia/epidemiologia , Fatores de Virulência/genética , Fatores de Virulência/metabolismo , Sequenciamento Completo do Genoma , beta-Lactamases/metabolismoRESUMO
Neisseria meningitidis sequence type 11 is an emerging cause of urethritis. We demonstrate by using whole-genome sequencing orogenital transmission of a N. meningitidis sequence type 11 isolate causing urethritis in a monogamous couple of men who have sex with men. These results suggest dissemination of this clonal complex among low-risk patients.
Assuntos
Antibacterianos/administração & dosagem , Doenças Transmissíveis Emergentes/transmissão , Infecções Meningocócicas/transmissão , Neisseria meningitidis/genética , Infecções Sexualmente Transmissíveis/transmissão , Uretrite/microbiologia , Doença Aguda , Azitromicina/administração & dosagem , Ceftriaxona/administração & dosagem , Doenças Transmissíveis Emergentes/tratamento farmacológico , Doenças Transmissíveis Emergentes/microbiologia , Humanos , Injeções Intramusculares , Masculino , Infecções Meningocócicas/tratamento farmacológico , Infecções Meningocócicas/microbiologia , Neisseria meningitidis/isolamento & purificação , Minorias Sexuais e de Gênero , Infecções Sexualmente Transmissíveis/tratamento farmacológico , Infecções Sexualmente Transmissíveis/microbiologia , Resultado do Tratamento , Uretrite/tratamento farmacológico , Sequenciamento Completo do Genoma , Adulto JovemRESUMO
This study was conducted to investigate extended-spectrum-ß-lactamase (ESBL) producing Enterobacteriaceae isolates from the Center of Maternity and Neonatology of Monastir, Tunisia. Fourty-six strains out of 283 were found to produce ESBL: Klebsiella pneumoniae (n = 37), Escherichia coli (n = 6), Enterobacter cloacae (n = 2), and Citrobacter freundi (n = 1). Genotyping analysis, using ERIC2 and RAPD, showed that strains were clonally unrelated. PCR amplification followed by sequencing revealed that all strains produced CTX-M-15. This enzyme was co-produced with TEM and SHV determinants in 34 and 36 strains respectively. The blaCTXM-15 gene was bracked by ISEcp1 and/or IS26 in 42 out of the 46 ESBL positive strains. The quinolone resistance determinants were associated to the ESBL producing isolates: we identified the qnrB1 gene in six isolates and the aac(6')-Ib-cr gene in five isolates. This epidemiological study shows the widespread of CTX-M-15 and qnr determinants among enterobacterial isolates from neonates hospitalized at the center of Maternity and Neonatology of Monastir suggesting either mother portage or horizontal transmission.
Assuntos
Enterobacteriaceae/genética , Enterobacteriaceae/isolamento & purificação , Genótipo , Plasmídeos/genética , beta-Lactamases/genética , Proteínas de Bactérias/genética , Citrobacter freundii/efeitos dos fármacos , Citrobacter freundii/genética , Infecção Hospitalar/microbiologia , Transmissão de Doença Infecciosa , Farmacorresistência Bacteriana/efeitos dos fármacos , Farmacorresistência Bacteriana/genética , Enterobacter cloacae/efeitos dos fármacos , Enterobacter cloacae/genética , Enterobacteriaceae/efeitos dos fármacos , Infecções por Enterobacteriaceae/microbiologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Humanos , Recém-Nascido , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/genética , Testes de Sensibilidade Microbiana , Epidemiologia Molecular , Quinolonas/farmacologia , Estudos Retrospectivos , Centros de Atenção Terciária , TunísiaAssuntos
Proteínas de Bactérias/análise , Bivalves/microbiologia , Escherichia coli/enzimologia , Escherichia coli/isolamento & purificação , Microbiologia de Alimentos , beta-Lactamases/análise , Animais , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Carbapenêmicos/farmacologia , Escherichia coli/genética , Genoma Bacteriano , Testes de Sensibilidade Microbiana , Tunísia , Sequenciamento Completo do Genoma , beta-Lactamases/genéticaRESUMO
In this study, an ertapenem-nonsusceptible Escherichia coli isolate was investigated to determine the genetic basis for its carbapenem resistance phenotype. This clinical strain was recovered from a patient that received, 1 year previously, ertapenem to treat a cholangitis due to a carbapenem-susceptible extended-spectrum-ß-lactamase (ESBL)-producing E. coli isolate. Whole-genome sequencing of these strains was performed using Illumina and single-molecule real-time sequencing technologies. It revealed that they belonged to the ST131 clonal group, had the predicted O25b:H4 serotype, and produced the CTX-M-15 and TEM-1 ß-lactamases. One nucleotide substitution was identified between these strains. It affected the ompR gene, which codes for a regulatory protein involved in the control of OmpC/OmpF porin expression, creating a Gly-63-Val substitution. The role of OmpR alteration was confirmed by a complementation experiment that fully restored the susceptibility to ertapenem of the clinical isolate. A modeling study showed that the Gly-63-Val change displaced the histidine-kinase phosphorylation site. SDS-PAGE analysis revealed that the ertapenem-nonsusceptible E. coli strain had a decreased expression of OmpC/OmpF porins. No significant defect in the growth rate or in the resistance to Dictyostelium discoideum amoeba phagocytosis was found in the ertapenem-nonsusceptible E. coli isolate compared to its susceptible parental strain. Our report demonstrates for the first time that ertapenem resistance may emerge clinically from ESBL-producing E. coli due to mutations that modulate the OmpR activity.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Escherichia coli , Transativadores/genética , beta-Lactamases/genética , beta-Lactamas/farmacologia , Enterobacteriáceas Resistentes a Carbapenêmicos/genética , Colangite/microbiologia , Dictyostelium/metabolismo , Dictyostelium/microbiologia , Farmacorresistência Bacteriana/genética , Ertapenem , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Infecções por Escherichia coli/tratamento farmacológico , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Fagocitose/fisiologia , Porinas/biossíntese , beta-Lactamases/metabolismoRESUMO
An interregional surveillance program was conducted in the northwestern part of France to determine the prevalence of carbapenem-nonsusceptible Enterobacteriaceae (CNSE) isolates and their susceptibility to ceftazidime-avibactam and aztreonam-avibactam combinations. Nonduplicate CNSE clinical isolates were prospectively collected from six hospitals between June 2012 and November 2013. MICs of ceftazidime and aztreonam, alone or combined with a fixed concentration of avibactam (4 µg/ml), and those of carbapenems (comparator agents) were determined. MICs of ertapenem in combination with phenylalanine arginine-naphthylamide dihydrochloride (PAßN) were also determined to assess active efflux. Genes encoding carbapenemases, plasmid-mediated AmpC enzymes, extended-spectrum ß-lactamases (ESBLs), and major outer membrane proteins (OMPs) were amplified and sequenced. OMPs were also extracted for SDS-PAGE analysis. Among the 139 CNSE isolates, mainly Enterobacter spp. and Klebsiella pneumoniae, 123 (88.4%) were ertapenem nonsusceptible, 12 (8.6%) exhibited reduced susceptibility to all carbapenems, and 4 Proteeae isolates (2.9%) were resistant to imipenem. Carbapenemase production was detected in only two isolates (producing OXA-48 and IMI-3). In contrast, OMP deficiency, in association with AmpCs and/or ESBLs (mainly CTX-M-9, SHV-12, and CTX-M-15), was largely identified among CNSE isolates. The ceftazidime-avibactam and aztreonam-avibactam combinations exhibited potent activity against CNSE isolates (MIC50/MIC90, 1/1 µg/ml and 0.5/0.5 µg/ml, respectively) compared to that of ceftazidime and aztreonam alone (MIC50/MIC90, 512/512 µg/ml and 128/512 µg/ml, respectively). This study reveals the in vitro activity of ceftazidime-avibactam and aztreonam-avibactam combinations against a large collection of porin-deficient enterobacterial isolates that are representative of the CNSE recovered in the northern part of France.
Assuntos
Antibacterianos/farmacologia , Compostos Azabicíclicos/farmacologia , Aztreonam/farmacologia , Ceftazidima/farmacologia , Infecções por Enterobacteriaceae/epidemiologia , Enterobacteriaceae/genética , beta-Lactamases/genética , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Dipeptídeos/metabolismo , Combinação de Medicamentos , Farmacorresistência Bacteriana Múltipla/genética , Enterobacteriaceae/efeitos dos fármacos , Enterobacteriaceae/isolamento & purificação , Enterobacteriaceae/metabolismo , Infecções por Enterobacteriaceae/tratamento farmacológico , Infecções por Enterobacteriaceae/microbiologia , Monitoramento Epidemiológico , Ertapenem , França/epidemiologia , Expressão Gênica , Humanos , Testes de Sensibilidade Microbiana , Plasmídeos/química , Plasmídeos/metabolismo , Prevalência , Estudos Prospectivos , beta-Lactamases/metabolismo , beta-Lactamas/farmacologiaRESUMO
OBJECTIVES: Resistance to all ß-lactams is emerging among Pseudomonas aeruginosa (PA) clinical isolates. Aztreonam and cefepime act synergistically in vitro against AmpC overproducing PA isolates. The objective of this study was to evaluate the clinical efficacy of this treatment in ICU patients infected with multidrug-resistant PA. MATERIAL AND METHODS: Retrospective study (2 years, 2 ICUs) in a tertiary university hospital. Inclusion criteria were proven infection with evidence of a bacterial strain of PA resistant to all ß-lactams and treated with the association of at least aztreonam plus cefepime. Treatment was considered effective for pneumonia using CPIS scores at the end of treatment and for other infections, using the SOFA score and signs of infection improvement at the end of treatment. Infectious episodes were classified as cure or failure. RESULTS: Thirteen patients were included (10 nosocomial pneumonia, 3 nosocomial intra-abdominal infections). The median [25th-75th percentiles] admission SAPS2 score was 54 [51-69] and the median SOFA score at the beginning of infection was 7 [4-8]. The median CPIS scores for pneumonia at the beginning and end of treatment were 9 [7-10.5] and 2 [0.75-5.5]. The duration of treatment with the combination of aztreonam plus cefepime was 14 days [9.5-16]. Nine episodes were classified as cures and 4 as failures, indicating a clinical efficacy of 69.2%. Overall mortality was 38.5%. DISCUSSION: These data suggest that the association of cefepime plus aztreonam could be an attractive alternative in the treatment of infections with multidrug-resistant PA to all ß-lactams with a clinical efficacy rate of 69%.
Assuntos
Antibacterianos/uso terapêutico , Aztreonam/uso terapêutico , Cefalosporinas/uso terapêutico , Infecção Hospitalar/tratamento farmacológico , Infecção Hospitalar/microbiologia , Infecções por Pseudomonas/tratamento farmacológico , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas de Bactérias/genética , Cefepima , Cuidados Críticos , Combinação de Medicamentos , Farmacorresistência Bacteriana Múltipla , Feminino , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Projetos Piloto , Pneumonia Bacteriana/tratamento farmacológico , Pneumonia Bacteriana/microbiologia , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/genética , Estudos Retrospectivos , Resistência beta-Lactâmica , beta-Lactamases/genéticaRESUMO
A retrospective epidemiological survey was carried out to determine the prevalence of carbapenemase producers among enterobacterial clinical isolates recovered in the center of maternity and neonatology of Monastir (Tunisia). PCR screening identified 1 OXA-48 and 2 OXA-204 producers, which coexpressed the CTX-M-15 or the CMY-4 ß-lactamases. PCR mapping showed that the bla(OXA-48) gene was carried by a Tn1999.2 transposon, whereas the bla(OXA-204) gene was part of the Tn2016 transposon-like structure. The OXA-48- or OXA-204-producing Klebsiella pneumoniae clinical isolates and the OXA-204-expressing Escherichia coli clinical isolate belonged to the widespread sequence types ST11, ST101, and ST617, respectively. The OXA-204 enzyme, which is a point derivative of the OXA-48 carbapenemase, had hitherto been reported in 2013 from K. pneumoniae isolate. Our study shows for the first time the dissemination of this resistance marker in E. coli strain. The coproduction of OXA-204 with CTX-M-15 and CMY-4 enzymes may potentiate the risk of multiresistance and may enhance the risk of dissemination.
Assuntos
Infecções por Enterobacteriaceae/epidemiologia , Infecções por Enterobacteriaceae/microbiologia , Escherichia coli/enzimologia , Klebsiella pneumoniae/enzimologia , beta-Lactamases/metabolismo , Elementos de DNA Transponíveis , DNA Bacteriano/química , DNA Bacteriano/genética , Estudos Epidemiológicos , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Ordem dos Genes , Humanos , Recém-Nascido , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/isolamento & purificação , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Estudos Retrospectivos , Análise de Sequência de DNA , Tunísia/epidemiologia , beta-Lactamases/genéticaRESUMO
A Cedecea davisae isolate, which was intermediate or resistant to third-generation cephalosporins and carbapenems, was recovered from a urine sample. Susceptibility testing, isoelectric focusing, and analysis of outer membrane proteins showed that AmpC ß-lactamase expression combined with porin deficiency accounted for the carbapenem resistance. A cloning experiment followed by phenotypic and enzymatic characterization identified a novel class C enzyme that was phylogenetically and biochemically close to the chromosome-borne ß-lactamases of the genera Enterobacter and Citrobacter.
Assuntos
Proteínas de Bactérias/genética , Carbapenêmicos/farmacologia , Enterobacteriaceae/efeitos dos fármacos , Enterobacteriaceae/enzimologia , beta-Lactamases/genética , Idoso , Sequência de Aminoácidos , Proteínas da Membrana Bacteriana Externa/genética , Proteínas de Bactérias/biossíntese , Farmacorresistência Bacteriana/genética , Enterobacteriaceae/genética , Feminino , Humanos , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Filogenia , Porinas/genética , beta-Lactamases/biossínteseRESUMO
In a series of 82 Staphylococcus strains isolated from culture, 100% were identified as Staphylococcus aureus by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS); 99.9% (77/82) of them were resistant to benzylpenicillin, oxacillin, and cefoxitin, and 6.1% (5/82) were susceptible to methicillin. Xpert MRSA/SA assay results were concordant with the phenotypic results in 76.8% (63/82) of cases and discordant in 23.2% (19/82) of cases. The MRSA/SA ELITe MGB kit results were concordant with phenotypic results in 100% of the cases. When comparing the Xpert MRSA/SA assay results with the MRSA/SA ELITe MGB kit results, 78% (64/82) of the cases were concordant, while 22% (18/82) of the cases were discordant. No statistically significant differences were observed between the two techniques. The PCR protocol that was used to validate the results of these two methods gave the following results: 49 were conventional methicillin-resistant S. aureus (MRSA) isolates (mecA positive and mecALGA251 negative), and 25 were phenotypic MRSA isolates (mecA negative and mecALGA251 positive).
Assuntos
Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Testes de Sensibilidade Microbiana/métodos , Técnicas de Diagnóstico Molecular/métodos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Infecções Estafilocócicas/diagnóstico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas de Bactérias/genética , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Proteínas de Ligação às Penicilinas , Infecções Estafilocócicas/microbiologia , Adulto JovemRESUMO
Only a few plasmid-borne AmpC (pAmpC) ß-lactamases, such as CMY-2, can account for carbapenem resistance in Enterobacteriaceae in combination with outer membrane impermeability. The aim of this study was to elucidate the contribution of Asn-346, which is well conserved among carbapenem-hydrolyzing pAmpCs, to the hydrolysis spectrum of CMY-2. Site-directed mutagenesis experiments were carried out to replace Asn-346 with glycine, alanine, valine, glutamate, aspartate, serine, threonine, glutamine, tyrosine, isoleucine, lysine, and histidine. The recombinant plasmids were transferred into wild-type and porin-deficient Escherichia coli strains. Asn-346 replacement reduced significantly the MICs of all ß-lactams, except the Asn-346-Ile substitution that increased the MICs of cephalosporins, whereas it decreased those of carbapenems. The biochemical characterization, along with a molecular modeling study, showed that the size and the polarity of the side chain at position 346 assisted substrate binding and turnover. This study shows for the first time that the amino acid at position 346 contributes to the ß-lactamase activity of cephalosporinases. Asparagine and isoleucine residues, which are well conserved at position 346 among AmpC-type enzymes, modulate their hydrolysis spectrum in an opposing sense. Ile-346 confers higher level of cephalosporins resistance, whereas Asn-346 confers carbapenem resistance in combination with outer membrane impermeability.
Assuntos
Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Escherichia coli/enzimologia , beta-Lactamases/química , beta-Lactamases/metabolismo , Asparagina/genética , Proteínas de Bactérias/genética , Carbapenêmicos/farmacologia , Escherichia coli/química , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Cinética , Mutagênese Sítio-Dirigida , beta-Lactamases/genéticaRESUMO
'Leptotrichia amnionii' is an underestimated fastidious inhabitant of the vaginal flora that can cause upper genital tract infections when predisposing factors are present. We describe here what is believed to be the first reported case of early onset meningitis due to 'L. amnionii' in a neonate with intrauterine growth retardation. The outcome was favourable after cefotaxime treatment.
Assuntos
Infecções por Fusobacteriaceae/microbiologia , Leptotrichia/isolamento & purificação , Meningites Bacterianas/microbiologia , Antibacterianos/administração & dosagem , Antibacterianos/uso terapêutico , Cefotaxima/administração & dosagem , Cefotaxima/uso terapêutico , Farmacorresistência Bacteriana , Quimioterapia Combinada , Feminino , Retardo do Crescimento Fetal , Infecções por Fusobacteriaceae/tratamento farmacológico , Humanos , Recém-Nascido , Leptotrichia/classificação , Leptotrichia/genética , Meningites Bacterianas/tratamento farmacológico , Dados de Sequência Molecular , Netilmicina/administração & dosagem , Netilmicina/uso terapêutico , Gravidez , Complicações Infecciosas na Gravidez/microbiologia , Adulto JovemRESUMO
The Xpert MRSA/SA BC assay was examined prospectively in patients with staphylococcal bacteremia including 6 patients with blood culture bottles inoculated with biological fluid (synovial fluid in 4 cases and peritoneal fluid in 2 cases). Among the 56 Staphylococci species isolated, 80.3% were coagulase-negative staphylococci (CoNS) and 19.7% were S. aureus. Methicillin susceptibility test results showed that 77.8% of isolates were methicillin-resistant CoNS (MRCoNS) and 22.2% of isolates were methicillin-susceptible CoNS (MSCoNS). Of 11 S. aureus isolates, 63.7% were methicillin-susceptible S. aureus (MSSA) and 36.3% were methicillin-resistant S. aureus (MRSA). Xpert MRSA/SA BC results showed that genotypic results were in concordance with phenotypic results in 94.6% of cases versus 5.4% of discordant cases. Of these 3 discordant cases, 1 S. aureus isolate had an MRSA phenotype and an SPA(+)mec(+)SCCmec(-) genotype and another S. aureus isolate was phenotypically MSSA and genotypically SPA(+)mec(+)SCCmec(-), and 1 S. epidermidis isolate was phenotypically MSCoNS and genotypically SPA(-)mec(+)SCCmec(-).