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1.
Arch Virol ; 164(11): 2783-2787, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31414285

RESUMO

Several types of Escherichia coli O-antigens form highly effective shields protecting the bacterial cell surface and preventing bacteriophages from interacting directly with their secondary (terminal) receptors. However, it is not clear if O-antigens of various types (O-serotypes) differ in their anti-phage protection efficacy. Here, we describe a new E. coli strain, F5, which has an E. coli O28ab-related O-antigen. Although the amount of O-antigen produced by this strain is comparable to that produced by other E. coli strains we tested, it appears to give the cells significantly lower protection against phage attack than other O-antigen types, such as the O-polysaccharide of E. coli F17, which we studied earlier.


Assuntos
Bacteriófagos/metabolismo , Escherichia coli/metabolismo , Antígenos O/metabolismo , Ligação Viral , Escherichia coli/classificação , Escherichia coli/genética , Antígenos O/genética
2.
J Med Microbiol ; 68(7): 1033-1041, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31199225

RESUMO

PURPOSE: Enteropathogens are frequently associated with diarrheal disease. Knowledge of their etiology and epidemiology is essential for the prevention and control of the sickness. This study describes the microbiological and epidemiological features of diarrheal disease in 197 symptomatic and 223 asymptomatic under-five-year-old children from southeastern Brazil, between January 2015 and September 2016. METHODS: Isolation of Escherichia coli, Salmonella, Shigella and Campylobacter was realized by culture. E. coli strains were screened by multiplex PCR, PFGE and O:H serotyping. Antimicrobial susceptibility testing was also performed. RESULTS: Most of the 127 enteropathogens isolated were diarrheagenic E. coli (96.1 %), with predominance of several serotypes of enteropathogenic E. coli (EPEC) and enteroaggregative E. coli (EAEC). Age, sex, rotavirus vaccination, recent use of antibiotics and previous contact with pets, were factors that revealed no significant effects on the probability of infection by the predominant pathogens. Even so, higher incomes could be related to a lesser chance of testing positive for EPEC. Evidence of possible EAEC clonal spread was detected, as well as genetic similarity among strains from both symptomatic and asymptomatic children. Resistance to antimicrobial agents was more pronounced among EAEC than EPEC. CONCLUSION: The occurrence of genetically similar diarrheagenic E. coli in both groups of children, likewise resistant to these agents, underscores the importance of establishing strategies for the prevention of outbreaks, especially among low-income households.


Assuntos
Diarreia/epidemiologia , Diarreia/microbiologia , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/microbiologia , Escherichia coli/genética , Brasil/epidemiologia , Pré-Escolar , Diarreia/economia , Escherichia coli/classificação , Infecções por Escherichia coli/economia , Fezes/microbiologia , Feminino , Humanos , Renda , Lactente , Masculino
3.
Chem Commun (Camb) ; 55(51): 7358-7361, 2019 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-31172143

RESUMO

Here, we describe a simple mix-and-read method for the detection of specific bacterial strains that uses a DNAzyme and a molecular beacon to generate a signal. We have greatly improved upon the previously described DNAzyme-based bacteria detection method by eliminating a tedious preparation step while maintaining detection sensitivity.


Assuntos
Extratos Celulares/análise , DNA Bacteriano/análise , DNA Catalítico/metabolismo , DNA de Cadeia Simples/análise , Escherichia coli/classificação , Técnicas Biossensoriais , Corantes Fluorescentes/química , Limite de Detecção
4.
Int J Food Microbiol ; 305: 108244, 2019 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-31202150

RESUMO

This study examined the prevalence and phenotypic and genotypic antibiotic resistance patterns of Staphylococcus aureus in sushi from 20 Danish outlets. Microbial quality of sushi products and food inspector ranking of outlets were assessed and results for thirteen of the outlets were compared with findings from a previous study in 2012. Inspector rankings were similar in the two studies. The mesophilic aerobic counts were slightly lower (p = 0.0296) in 2017 than in 2012 with average values of the 13 shops of 5.2 log CFU/g and 5.7 log CFU/g, respectively. In both studies E. coli was only found in the products from outlets that did not have consistently superior rankings. On the other hand prevalence and average counts of Staphylococcus spp. were slightly higher in 2017 (p = 0.0286) but no methicillin-resistant S. aureus (MRSA) were observed in the present study. Methicillin-sensitive S. aureus (MSSA) were, however, isolated from 18.7% of sushi products with an average count below 2 log CFU/g. Based on spa and multilocus sequence typing (MLST), isolates belonged to clonal complex CC7 (t2016), CC20 (t7836), CC45 (t065, t127, t362), CC88 (t1998) and CC398 (t164, t331, t1451). The Panton-Valentine leukocidin (PVL)-encoding gene lukF was detected only in isolates of the t065 spa-type whereas the scn gene from the ΦSa3 prophage was detected in 76.5% of the isolates, supporting that the majority of isolates were of likely human origin. Thirty-six isolates (70.6%) were resistant to at least one of the antibiotic compounds tested. Antibiotic resistance genes that confer resistance to ß-lactams (blaZ) and macrolides (ermC) were detected in 33.3% and 9.8% of isolates, respectively. The tet(K) gene that encode tetracycline resistance was only found in a t7836 strain. Overall, this study indicates that S. aureus in sushi products in Denmark do not represent a major food safety hazard due to, firstly, the low temperature and limited time of storage of product may prevent significant growth and production of toxic levels of enterotoxin of this species. Secondly, the S. aureus isolates obtained did not include MRSA variants and none of them encoded PVL that constitute one of the virulence factors in pathogenesis. Several MSSA isolates contained however genes encoding antibiotic resistance, which emphasize the potential role of foods as vehicles for transmission of such variants.


Assuntos
Farmacorresistência Bacteriana , Produtos Pesqueiros/microbiologia , Inspeção de Alimentos/normas , Animais , Antibacterianos/farmacologia , Dinamarca , Escherichia coli/classificação , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Peixes , Humanos , Meticilina/farmacologia , Staphylococcus aureus Resistente à Meticilina/classificação , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Testes de Sensibilidade Microbiana , Tipagem de Sequências Multilocus , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/classificação , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/genética , Staphylococcus aureus/isolamento & purificação , Fatores de Virulência/genética , beta-Lactamas/farmacologia
5.
BMC Res Notes ; 12(1): 244, 2019 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-31036061

RESUMO

OBJECTIVES: Diabetic foot ulcers (DFUs) often lead to hospital admissions, amputations and deaths; however, there is no up-to-date information on microbial isolates from DFUs and no mention of utilization of molecular techniques in Sub-Saharan Africa. We conducted a cross-sectional study among 83 adult patients at a tertiary hospital in Kenya over 12 months. The study aimed to isolate, identify bacteria, their antibiotic susceptibility patterns in active DFUs, and to compare standard microbiological methods versus a real-time PCR commercial kit in the detection of Staphylococcus aureus DNA and methicillin-resistant S. aureus (MRSA) DNA. RESULTS: Eighty swabs (94%) were culture-positive; 29% were Gram-positive and 65% were Gram-negative. The main organisms isolated were S. aureus (16%), Escherichia coli (15%), Proteus mirabilis (11%), Klebsiella pneumoniae (7%) and Pseudomonas aeruginosa (7%). The bacterial isolates showed resistance to commonly used antibiotics such as ampicillin, amoxicillin, cefepime, ceftazidime, cefuroxime, clindamycin, erythromycin, piperacillin-tazobactam, tetracycline and trimethoprim-sulphamethoxazole (TMPSMX). Thirty-one percent of the S. aureus isolated and 40% of the Gram-negatives were multi-drug resistant organisms (MDROs). There was a high prevalence of nosocomial bacteria. MRSA were not identified using culture methods but were identified using PCR. PCR was more sensitive but less specific than culture-based methods to identify S. aureus.


Assuntos
Antibacterianos/uso terapêutico , Infecções Bacterianas/diagnóstico , Pé Diabético/diagnóstico , Farmacorresistência Bacteriana Múltipla , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Staphylococcus aureus/efeitos dos fármacos , Infecções Bacterianas/tratamento farmacológico , Infecções Bacterianas/epidemiologia , Infecções Bacterianas/microbiologia , Técnicas de Tipagem Bacteriana , Cefalosporinas/uso terapêutico , Clindamicina/uso terapêutico , Estudos Transversais , Pé Diabético/tratamento farmacológico , Pé Diabético/epidemiologia , Pé Diabético/microbiologia , Escherichia coli/classificação , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Humanos , Quênia/epidemiologia , Klebsiella pneumoniae/classificação , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/isolamento & purificação , Macrolídeos/uso terapêutico , Staphylococcus aureus Resistente à Meticilina/classificação , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Testes de Sensibilidade Microbiana , Penicilinas/uso terapêutico , Proteus mirabilis/classificação , Proteus mirabilis/efeitos dos fármacos , Proteus mirabilis/genética , Proteus mirabilis/isolamento & purificação , Pseudomonas aeruginosa/classificação , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real , Staphylococcus aureus/classificação , Staphylococcus aureus/genética , Staphylococcus aureus/isolamento & purificação , Sulfanilamidas/uso terapêutico
6.
Microb Pathog ; 132: 355-361, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31096003

RESUMO

It is important to evaluate the antimicrobial resistance pattern by genotypic and phenotypic methods in epidemiological studies in order to control antimicrobial resistance and to improve the outcome of the treatments. Four-hundred and thirty clinical mastitis samples were collected from 14 dairy herds in five different cities. The antimicrobial susceptibility test was performed using agar disk diffusion for 70 identified Escherichia coli isolates. The antimicrobial resistance genes including strA, strB, aadA, sulI, sulII, sulIII, ampC were detected by PCR method. Phylogenic groups were determined by Clermont's multiplex PCR method, and RAPD typing was performed on all isolates. Most isolates were resistant to lincomicin and streptomycin, whereas sulfa-trimethoprim has the lowest resistance rate. Moreover, ampC, aadA and sul2 genes had the highest frequency (92.85%, 38.57%, and 32.85% respectively). 20% of all the isolates carried strA and strB genes, and 11.42% of the isolates had sul1 gene and 10% of the isolates had the less frequent sul3 gene. Of the total of 70 E. coli isolates, 26 (37.14%), 20 (28.5%), 17 (24.2%), 8 (11.4%) isolates belonged to B1, A, B2 and D phylogenic groups respectively. strA, strB, sul2and aadA resistance genes had the highest percentage in A phylogenic groups. Based on RAPD-PCR method, E. coli isolates were classified in four clusters. The result showed a high phenotypic and genotypic E. coli resistance to the current antimicrobials with a similar pattern in different cities; also the majority of E. coli isolates belonged to B1 group which mainly contains the commensal E. coli isolates.


Assuntos
Farmacorresistência Bacteriana Múltipla/genética , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Genótipo , Mastite Bovina/microbiologia , Animais , Antibacterianos/farmacologia , Bovinos , Escherichia coli/classificação , Escherichia coli/efeitos dos fármacos , Infecções por Escherichia coli/veterinária , Proteínas de Escherichia coli/genética , Feminino , Testes de Sensibilidade Microbiana/veterinária , Filogenia , Reação em Cadeia da Polimerase , Técnica de Amplificação ao Acaso de DNA Polimórfico , Estreptomicina/farmacologia
7.
Vet Ital ; 55(1): 35-46, 2019 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-30951180

RESUMO

The objective of the study was to determine the virulence and antimicrobial resistance traits of 100 fecal E. coli strains isolated from clinically healthy chickens in Algeria. Most of isolates belonged to phylogroups A (45%) and B1 (37%) and showed a great diversity in DNA profiles. The genes fimH, tsh, entB, iutA, irp2, fyuA, iroN, sitA, etsA, etsB, eitA, iss, traT, ompT, hlyF, vat, ibeA, cvaA, cvaB5', cvaB3', cvaC, cma and cbi were detected. Combinations of virulence genes defined 67 virulence profiles. High resistance rates (62­97%) were noted for amoxicillin, amoxicillin­clavulanic acid, cefazolin, fluoroquinolones, tetracycline, trimethoprim, sulfonamides and sulfamethoxazole/ trimethoprim, and 93% of strains were multidrug­resistant. Combinations of resistance phenotypes defined 59 resistance patterns. The genes blaTEM, blaSHV, blaCTX­M­1, tetA, tetB, qnrB, qnrS1, sul1, sul2, sul3, dfrA1, dfrA7, dfrA12 and dfrA14 were identified and class 1 integrons were detected in 49% of isolates. A rate of 37% of strains was resistant to mercury, with the presence of merA gene. The study reports the presence in the avian strains isolated from fecal swabs of virulence genes of plasmid origin characteristic of ExPEC strains associated with high resistance to first­line antibiotics and class 1 integrons, this augurs a risk for human and animal health.


Assuntos
Galinhas , Farmacorresistência Bacteriana/genética , Infecções por Escherichia coli/veterinária , Escherichia coli/isolamento & purificação , Perfil Genético , Doenças das Aves Domésticas/epidemiologia , Argélia/epidemiologia , Animais , Anti-Infecciosos/farmacologia , Escherichia coli/classificação , Escherichia coli/genética , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/microbiologia , Fezes/microbiologia , Filogenia , Doenças das Aves Domésticas/microbiologia , Virulência/genética
8.
Eur J Clin Microbiol Infect Dis ; 38(5): 945-949, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30877486

RESUMO

Our aim was to evaluate the prevalence of fosfomycin-resistant strains among ESBL-producing Escherichia coli isolates recovered from community patients in Switzerland. A total of 1225 ESBL-producing E. coli isolates were collected between 2012 and 2013 from a private and community laboratory. Fosfomycin resistance was assessed by using the novel rapid fosfomycin/E. coli NP test and agar dilution method. Resistant isolates were further investigated for acquired resistance genes fosA1-7 by PCR and sequencing. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) were performed to evaluate the clonal relationship among fosA3-carrying isolates. Out of the 1225 ESBL-producing E. coli isolates analyzed in this study, 1208 were fosfomycin susceptible while 17 were fosfomycin resistant. No discrepancy was observed between the rapid fosfomycin/E. coli NP test and the agar dilution method taken as the gold standard. Five out of the 17 resistant isolates carried a fosA-like gene. No clonal relationship was observed among those isolates. Here, the prevalence of fosfomycin resistance among ESBL-producing E. coli isolates in the community is reported for the first time in Switzerland, being ca. 1.4%. Among the five isolates carrying a fosA gene, four encoded the FosA3 enzyme, being the most prevalent fosfomycin-resistant determinant. An excellent correlation was observed between minimum inhibitory concentration-based susceptibility categorization and results of the rapid fosfomycin/E. coli NP test, further indicating the excellent sensitivity and specificity of this recently developed rapid test whose results are obtained in less than 2 h.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana Múltipla , Infecções por Escherichia coli/microbiologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/enzimologia , Fosfomicina/farmacologia , beta-Lactamases/biossíntese , DNA Bacteriano/genética , Farmacorresistência Bacteriana Múltipla/genética , Escherichia coli/classificação , Escherichia coli/genética , Infecções por Escherichia coli/epidemiologia , Proteínas de Escherichia coli/genética , Humanos , Testes de Sensibilidade Microbiana , Tipagem Molecular , Prevalência , Análise de Sequência de DNA , Suíça/epidemiologia , beta-Lactamases/genética
9.
Nat Commun ; 10(1): 1360, 2019 03 25.
Artigo em Inglês | MEDLINE | ID: mdl-30911012

RESUMO

TonB-dependent transporters (TBDTs) are ubiquitous outer membrane ß-barrel proteins that import nutrients and bacteriocins across the outer membrane in a proton motive force-dependent manner, by directly connecting to the ExbB/ExbD/TonB system in the inner membrane. Here, we show that the TBDT Oar in Myxococcus xanthus is required for secretion of a protein, protease PopC, to the extracellular milieu. PopC accumulates in the periplasm before secretion across the outer membrane, and the proton motive force has a role in secretion to the extracellular milieu. Reconstitution experiments in Escherichia coli demonstrate that secretion of PopC across the outer membrane not only depends on Oar but also on the ExbB/ExbD/TonB system. Our results indicate that TBDTs and the ExbB/ExbD/TonB system may have roles not only in import processes but also in secretion of proteins.


Assuntos
Proteínas de Bactérias/genética , Membrana Celular/metabolismo , Regulação Bacteriana da Expressão Gênica , Proteínas de Membrana/genética , Myxococcus xanthus/genética , Peptídeo Hidrolases/genética , Proteínas de Bactérias/metabolismo , Transporte Biológico , Escherichia coli/classificação , Escherichia coli/genética , Escherichia coli/metabolismo , Teste de Complementação Genética , Isoenzimas/genética , Isoenzimas/metabolismo , Proteínas de Membrana/metabolismo , Myxococcus xanthus/classificação , Myxococcus xanthus/metabolismo , Peptídeo Hidrolases/metabolismo , Periplasma/metabolismo , Filogenia , Força Próton-Motriz
10.
BMC Genomics ; 20(1): 212, 2019 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-30866824

RESUMO

BACKGROUND: Escherichia coli is an important pathogen that causes diarrhea in both humans and animals. To determine the relationships between putative virulence factors and pathotypes or host taxa, many molecular studies on diarrhea-associated E. coli have been reported. However, little is known regarding genome-wide variation of E. coli from animal hosts. In this study, we performed whole genome sequencing of 127 E. coli isolates from sheep and swine with diarrhea in China. We compared isolates to explore the phylogenomic relatedness based on host origin. We explored the relationships of putative virulence factors across host taxa and pathotypes. Antimicrobial resistance was also tested. RESULTS: The E. coli genomes in this study were diverse with clear differences in the SNP, MLST, and O serotypes. Seven putative virulence factors (VFs) were prevalent (> 95%) across the isolates, including Hcp, csgC, dsdA, feoB, fepA, guaA, and malX. Sixteen putative VFs showed significantly different distributions (P < 0.05) in strains from sheep and swine and were primarily adhesion- and toxin-related genes. Some putative VFs were co-occurrent in some specific pathotypes and O serotypes. The distribution of 4525 accessory genes of the 127 strains significantly differed (P < 0.05) between isolates obtained from the two animal species. The 127 animal isolates sequenced in this study were each classified into one of five pathotypes: EAEC, ETEC, STEC, DAEC, and EPEC, with 66.9% of isolates belonging to EAEC. Analysis of stx subtypes and a minimum spanning tree based on MLST revealed that STEC isolates from sheep and EAEC isolates from sheep and swine have low potential to infect humans. Antibiotic resistance analysis showed that the E. coli isolates were highly resistant to ampicillin and doxycycline. Isolates from southeast China were more resistant to antibiotics than isolates from northwest China. Additionally, the plasmid-mediated colist in resistance gene mcr-1 was detected in 15 isolates, including 4 from sheep in Qinghai and 11 from swine in Jiangsu. CONCLUSIONS: Our study provides insight into the genomes of E. coli isolated from animal sources. Distinguishable differences between swine and sheep isolates at the genomic level provides a baseline for future investigations of animal E. coli pathogens.


Assuntos
Animais Domésticos/microbiologia , Diarreia/microbiologia , Infecções por Escherichia coli/veterinária , Escherichia coli/classificação , Genômica/métodos , Animais , Técnicas de Tipagem Bacteriana , China , Diarreia/veterinária , Farmacorresistência Bacteriana , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Escherichia coli/patogenicidade , Proteínas de Escherichia coli/genética , Tipagem de Sequências Multilocus , Filogenia , Polimorfismo de Nucleotídeo Único , Análise de Sequência de DNA/métodos , Sorogrupo , Ovinos/microbiologia , Suínos/microbiologia , Fatores de Virulência/genética
11.
Nat Commun ; 10(1): 935, 2019 02 25.
Artigo em Inglês | MEDLINE | ID: mdl-30804335

RESUMO

Synthetic biology aims to design and construct bacterial genomes harboring the minimum number of genes required for self-replicable life. However, the genome-reduced bacteria often show impaired growth under laboratory conditions that cannot be understood based on the removed genes. The unexpected phenotypes highlight our limited understanding of bacterial genomes. Here, we deploy adaptive laboratory evolution (ALE) to re-optimize growth performance of a genome-reduced strain. The basis for suboptimal growth is the imbalanced metabolism that is rewired during ALE. The metabolic rewiring is globally orchestrated by mutations in rpoD altering promoter binding of RNA polymerase. Lastly, the evolved strain has no translational buffering capacity, enabling effective translation of abundant mRNAs. Multi-omic analysis of the evolved strain reveals transcriptome- and translatome-wide remodeling that orchestrate metabolism and growth. These results reveal that failure of prediction may not be associated with understanding individual genes, but rather from insufficient understanding of the strain's systems biology.


Assuntos
Escherichia coli/genética , Genoma Bacteriano , Escherichia coli/classificação , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Evolução Molecular , Regulação Bacteriana da Expressão Gênica , Tamanho do Genoma , Mutação , Fenótipo , Transcriptoma
12.
Clin Microbiol Rev ; 32(2)2019 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-30700432

RESUMO

New Delhi metallo-ß-lactamase (NDM) is a metallo-ß-lactamase able to hydrolyze almost all ß-lactams. Twenty-four NDM variants have been identified in >60 species of 11 bacterial families, and several variants have enhanced carbapenemase activity. Klebsiella pneumoniae and Escherichia coli are the predominant carriers of bla NDM, with certain sequence types (STs) (for K. pneumoniae, ST11, ST14, ST15, or ST147; for E. coli, ST167, ST410, or ST617) being the most prevalent. NDM-positive strains have been identified worldwide, with the highest prevalence in the Indian subcontinent, the Middle East, and the Balkans. Most bla NDM-carrying plasmids belong to limited replicon types (IncX3, IncFII, or IncC). Commonly used phenotypic tests cannot specifically identify NDM. Lateral flow immunoassays specifically detect NDM, and molecular approaches remain the reference methods for detecting bla NDM Polymyxins combined with other agents remain the mainstream options of antimicrobial treatment. Compounds able to inhibit NDM have been found, but none have been approved for clinical use. Outbreaks caused by NDM-positive strains have been reported worldwide, attributable to sources such as contaminated devices. Evidence-based guidelines on prevention and control of carbapenem-resistant Gram-negative bacteria are available, although none are specific for NDM-positive strains. NDM will remain a severe challenge in health care settings, and more studies on appropriate countermeasures are required.


Assuntos
Escherichia coli/enzimologia , Klebsiella pneumoniae/enzimologia , beta-Lactamases/genética , beta-Lactamases/metabolismo , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Península Balcânica , Escherichia coli/classificação , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Medicina Baseada em Evidências , Variação Genética , Humanos , Índia , Klebsiella pneumoniae/classificação , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/genética , Oriente Médio , Filogeografia , Inibidores de beta-Lactamases/farmacologia
13.
Microb Genom ; 5(2)2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30672731

RESUMO

Avian pathogenic Escherichia coli (APEC) cause widespread economic losses in poultry production and are potential zoonotic pathogens. Genome sequences of 95 APEC from commercial poultry operations in four Australian states that carried the class 1 integrase gene intI1, a proxy for multiple drug resistance (MDR), were characterized. Sequence types ST117 (22/95), ST350 (10/95), ST429 and ST57 (each 9/95), ST95 (8/95) and ST973 (7/95) dominated, while 24 STs were represented by one or two strains. FII and FIB repA genes were the predominant (each 93/95, 98 %) plasmid incompatibility groups identified, but those of B/O/K/Z (25/95, 26 %) and I1 (24/95, 25 %) were also identified frequently. Virulence-associated genes (VAGs) carried by ColV and ColBM virulence plasmids, including those encoding protectins [iss (91/95, 96 %), ompT (91/95, 96 %) and traT (90/95, 95 %)], iron-acquisition systems [sitA (88/95, 93 %), etsA (87/95, 92 %), iroN (84/95, 89 %) and iucD/iutA (84/95, 89 %)] and the putative avian haemolysin hylF (91/95, 96 %), featured prominently. Notably, mobile resistance genes conferring resistance to fluoroquinolones, colistin, extended-spectrum ß-lactams and carbapenems were not detected in the genomes of these 95 APEC but carriage of the sulphonamide resistance gene, sul1 (59/95, 63 %), the trimethoprim resistance gene cassettes dfrA5 (48/95, 50 %) and dfrA1 (25/95, 27 %), the tetracycline resistance determinant tet(A) (51/95, 55 %) and the ampicillin resistance genes blaTEM-1A/B/C (48/95, 52 %) was common. IS26 (77/95, 81 %), an insertion element known to capture and mobilize a wide spectrum of antimicrobial resistance genes, was also frequently identified. These studies provide a baseline snapshot of drug-resistant APEC in Australia and their role in the carriage of ColV-like virulence plasmids.


Assuntos
Farmacorresistência Bacteriana Múltipla/genética , Infecções por Escherichia coli/microbiologia , Infecções por Escherichia coli/veterinária , Escherichia coli/classificação , Escherichia coli/genética , Doenças das Aves Domésticas/microbiologia , Animais , Austrália , Toxinas Bacterianas/genética , Elementos de DNA Transponíveis , DNA Bacteriano/genética , Escherichia coli/patogenicidade , Proteínas de Escherichia coli/genética , Genoma Bacteriano , Integrases/genética , Plasmídeos , Análise de Sequência de DNA/métodos , Virulência/genética , Fatores de Virulência/genética , Sequenciamento Completo do Genoma
14.
BMC Microbiol ; 19(1): 17, 2019 01 17.
Artigo em Inglês | MEDLINE | ID: mdl-30654756

RESUMO

BACKGROUND: To describe the temporal dynamics, molecular characterization, clinical and ex vivo virulence of emerging O1:K1 neonatal meningitis Escherichia coli (NMEC) strains of Sequence Type complex (STc) 95 in France. The national reference center collected NMEC strains and performed whole genome sequencing (WGS) of O1:K1 STc95 NMEC strains for phylogenetic and virulence genes content analysis. Data on the clinical and biological features of patients were also collected. Ex vivo virulence was assessed using the Dictyostelium discoideum amoeba model. RESULTS: Among 250 NMEC strains collected between 1998 and 2015, 38 belonged to O1:K1 STc95. This clonal complex was the most frequently collected after 2004, representing up to 25% of NMEC strains in France. Phylogenetic analysis demonstrated that most (74%) belonged to a cluster designated D-1, characterized by the adhesin FimH30. There is no clinical data to suggest that this cluster is more pathogenic than its counterparts, although it is highly predominant and harbors a large repertoire of extraintestinal virulence factors, including a pS88-like plasmid. Ex vivo virulence model showed that this cluster was generally less virulent than STc95 reference strains of O45S88:H7 and O18:H7 serotypes. However, the model showed differences between several subclones, although they harbor the same known virulence determinants. CONCLUSIONS: The emerging clonal group O1:K1 STc95 of NMEC strains is mainly composed of a cluster with many virulence factors but of only moderate virulence. Whether its emergence is due to its ability to colonize the gut thanks to FimH30 or pS88-like plasmid remains to be determined.


Assuntos
Escherichia coli/genética , Genoma Bacteriano/genética , Doenças do Recém-Nascido/microbiologia , Meningite devida a Escherichia coli/microbiologia , Sequenciamento Completo do Genoma , Escherichia coli/classificação , Escherichia coli/patogenicidade , França , Humanos , Recém-Nascido , Modelos Genéticos , Filogenia , Virulência/genética
15.
Clin Microbiol Infect ; 25(7): 839-844, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30648603

RESUMO

OBJECTIVES: Distinguishing between urinary tract infection (UTI) and asymptomatic bacteriuria (ABU) is difficult in the geriatric population since specific symptoms are often lacking. Escherichia coli is the most frequent UTI pathogen in this population but also a common urine colonizer. We hypothesized that detecting E. coli phylogroups B2 or D, which were previously associated with virulent strains responsible for extra-intestinal infections outside elderly patients, could help in distinguishing UTI from ABU. METHODS: Consecutive cases of E. coli bacteriuria diagnosed in hospitalized patients >75 years old during 3 months were investigated for E. coli phylogroups. Multiplex PCR was used to search for several virulence genes as previously described. Characteristics of UTI and ABU cases, assessed retrospectively according to definitions and geriatric expertise, were compared. RESULTS: Out of 233 bacteriuria cases, 60 were assessed to be UTI and 163 to be ABU, with 10 cases unclassified. E. coli strains belonging to the phylogroups B2 and D were significantly more frequent in UTI (48/60, 80%) than in ABU (101/163, 62%) by univariate and multivariate analyses (OR 3.05, 1.44-6.86, p 0.005). Out of all the host and bacterial characteristics studied, falls (p 0.032), comorbidities (p 0.041), and altered autonomy evaluated by a low activity of daily living score (p 0.027) were also associated with UTI using univariate and multivariate analysis. CONCLUSIONS: Determination of the E. coli phylogroup, in addition to some host characteristics, can help to distinguish UTI from ABU in elderly patients with bacteriuria. If this hypothesis is confirmed by prospective studies, then inappropriate use of antibiotics may be reduced in ABU cases.


Assuntos
Infecções Assintomáticas , Bacteriúria/microbiologia , Infecções por Escherichia coli/diagnóstico , Escherichia coli/classificação , Infecções Urinárias/microbiologia , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/farmacologia , Bacteriúria/diagnóstico , Escherichia coli/efeitos dos fármacos , Escherichia coli/isolamento & purificação , Proteínas de Escherichia coli/genética , Feminino , Humanos , Masculino , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase Multiplex , Estudos Prospectivos , Estudos Retrospectivos , Infecções Urinárias/diagnóstico , Virulência , Fatores de Virulência/genética
16.
Nat Protoc ; 14(2): 415-440, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30635653

RESUMO

The construction of genome-wide mutant collections has enabled high-throughput, high-dimensional quantitative characterization of gene and chemical function, particularly via genetic and chemical-genetic interaction experiments. As the throughput of such experiments increases with improvements in sequencing technology and sample multiplexing, appropriate tools must be developed to handle the large volume of data produced. Here, we describe how to apply our approach to high-throughput, fitness-based profiling of pooled mutant yeast collections using the BEAN-counter software pipeline (Barcoded Experiment Analysis for Next-generation sequencing) for analysis. The software has also successfully processed data from Schizosaccharomyces pombe, Escherichia coli, and Zymomonas mobilis mutant collections. We provide general recommendations for the design of large-scale, multiplexed barcode sequencing experiments. The procedure outlined here was used to score interactions for ~4 million chemical-by-mutant combinations in our recently published chemical-genetic interaction screen of nearly 14,000 chemical compounds across seven diverse compound collections. Here we selected a representative subset of these data on which to demonstrate our analysis pipeline. BEAN-counter is open source, written in Python, and freely available for academic use. Users should be proficient at the command line; advanced users who wish to analyze larger datasets with hundreds or more conditions should also be familiar with concepts in analysis of high-throughput biological data. BEAN-counter encapsulates the knowledge we have accumulated from, and successfully applied to, our multiplexed, pooled barcode sequencing experiments. This protocol will be useful to those interested in generating their own high-dimensional, quantitative characterizations of gene or chemical function in a high-throughput manner.


Assuntos
Interação Gene-Ambiente , Genoma Bacteriano , Genoma Fúngico , Saccharomyces cerevisiae/genética , Bibliotecas de Moléculas Pequenas/farmacologia , Software , Código de Barras de DNA Taxonômico/métodos , DNA Bacteriano/genética , DNA Bacteriano/metabolismo , DNA Fúngico/genética , DNA Fúngico/metabolismo , Escherichia coli/classificação , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Mutação , Saccharomyces cerevisiae/classificação , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/metabolismo , Schizosaccharomyces/classificação , Schizosaccharomyces/efeitos dos fármacos , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Zymomonas/classificação , Zymomonas/efeitos dos fármacos , Zymomonas/genética , Zymomonas/metabolismo
17.
Cell Host Microbe ; 25(2): 301-312.e5, 2019 02 13.
Artigo em Inglês | MEDLINE | ID: mdl-30683582

RESUMO

Crohn's disease (CD) is an inflammatory bowel disease influenced by bacteria. Adherent-invasive E. coli (AIEC) is associated with CD, yet the adaptations facilitating AIEC gut colonization are unknown. AIEC isolates exhibit high genetic diversity, suggesting strains evolve independently across different gut environments. We tracked the adaptive evolution of AIEC in a murine model of chronic colonization across multiple hosts and transmission events. We detected evolved lineages that outcompeted the ancestral strain in the host through independent mechanisms. One lineage was hypermotile because of a mobile insertion sequence upstream of the master flagellar regulator, flhDC, which enhanced AIEC invasion and establishment of a mucosal niche. Another lineage outcompeted the ancestral strain through improved use of acetate, a short-chain fatty acid in the gut. The presence of hypermotile and acetate-consuming lineages discriminated E. coli isolated from CD patients from healthy controls, suggesting an evolutionary trajectory that distinguishes AIEC from commensal E. coli.


Assuntos
Adaptação Biológica , Doença de Crohn/microbiologia , Infecções por Escherichia coli/microbiologia , Escherichia coli/classificação , Escherichia coli/isolamento & purificação , Trato Gastrointestinal/microbiologia , Variação Genética , Animais , Células CACO-2 , Modelos Animais de Doenças , Transmissão de Doença Infecciosa , Escherichia coli/genética , Infecções por Escherichia coli/transmissão , Feminino , Humanos , Camundongos Endogâmicos C57BL
19.
J Med Microbiol ; 68(2): 169-177, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30624176

RESUMO

PURPOSE: In infectious disease therapy, administration of adequate antimicrobial agents is essential for preventing the emergence and spread of resistant bacteria. However, conventional antimicrobial susceptibility testing (AST), based on bacterial growth, is time consuming; therefore, a rapid, simple assay is needed for the timely selection of appropriate antibiotics in clinical laboratories. Here, we established a simple, cost-effective, time-saving and highly sensitive AST assay based on loop-mediated isothermal amplification (LAMP). METHODOLOGY: The targeted bacteria were cultivated for a short period with or without antibiotic before the LAMP reaction. The time to detect a positive reaction with LAMP was used to generate a threshold time (Tt) value, and subtraction of the Tt value for an antibiotic-free sample from the Tt value in an antibiotic-exposed sample generated the ΔTt value, which was used as a marker of antimicrobial susceptibility. The ΔTt value generated using the LAMP-based assay simply and quickly detected antimicrobial resistance in clinical Escherichia coli isolates. RESULTS: Detection of susceptibility to levofloxacin using the ΔTt value perfectly matched with the results of the conventional assay. In addition, the sensitivity and specificity for the detection of ampicillin, trimethoprim-sulfamethoxazole and fosfomycin resistance were 100 %, 93.8 %, 100 % and 80.0 %, 93.3 %, 97.6 %, respectively. CONCLUSION: These results showed that this LAMP-based AST has high sensitivity and specificity for detecting resistant strains and a significant time advantage compared with the conventional method.


Assuntos
Anti-Infecciosos/farmacologia , Farmacorresistência Bacteriana , Escherichia coli/efeitos dos fármacos , Testes de Sensibilidade Microbiana/métodos , DNA Bacteriano/química , DNA Ribossômico/química , Escherichia coli/classificação , Escherichia coli/genética , Infecções por Escherichia coli/microbiologia , Humanos , Técnicas de Diluição do Indicador , Testes de Sensibilidade Microbiana/normas , Fenótipo , RNA Ribossômico 16S/genética , Sensibilidade e Especificidade , Infecções Urinárias/microbiologia , Urina/microbiologia , beta-Lactamases/genética
20.
J Med Microbiol ; 68(2): 178-187, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30624177

RESUMO

PURPOSE: Escherichia coli is a leading cause of bloodstream infection (BSI) in hospitals and communities. METHODOLOGY: We conducted a retrospective study in 2015 to evaluate the clinical features and microbiological characteristics of E. coli BSI acquired in the hospital and community. RESULTS: A total of 100 patients with E. coli BSI were enrolled, among whom 60 % had hospital-onset (HO) BSI while 40 % had community-onset (CO) BSI. Patients with HO BSI had higher percentages of haematological disorders, immunosuppression conditions, underwent surgery within 2 weeks and had a higher 30-day mortality. The prevalences of multidrug-resistant and extended-spectrum ß-lactamase-producing strains were 81 and 60 %, respectively. Resistance percentages to ampicillin, ampicillin-sulbactam, cefazolin, ceftriaxone, ciprofloxacin and levofloxacin were greater than 50 %. Of the 43 different sequence types (STs) identified, ST131 (15.3 %) was the most common. The serum agglutination rate was 52 % in which 13 O and 11 H serogroups were observed. Among the 36 detected virulence factor (VF) genes, IutA (66 %) and traT (61 %) were the most predominant. papA, papC and papEF were different between the CO and HO BSI groups. VF scores were high (mean >7) in the frequently detected ST95, ST1193 and ST131. CONCLUSION: This study revealed that the clinical features of HO and CO E. coli BSI were different. STs and serotypes showed a great diversity in this region while VF genes of the isolates varied between clones.


Assuntos
Bacteriemia/microbiologia , Infecções Comunitárias Adquiridas/microbiologia , Infecção Hospitalar/microbiologia , Infecções por Escherichia coli/microbiologia , Escherichia coli/classificação , Adulto , Idoso , Idoso de 80 Anos ou mais , Testes de Aglutinação , Escherichia coli/genética , Escherichia coli/imunologia , Escherichia coli/patogenicidade , Humanos , Masculino , Pessoa de Meia-Idade , Tipagem de Sequências Multilocus , Filogenia , Estudos Retrospectivos , Sorogrupo , Fatores de Virulência/genética
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