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1.
BMC Biol ; 17(1): 76, 2019 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-31533707

RESUMO

BACKGROUND: The selection pressure exercised by antibiotic drugs is an important consideration for the wise stewardship of antimicrobial treatment programs. Treatment decisions are currently based on crude assumptions, and there is an urgent need to develop a more quantitative knowledge base that can enable predictions of the impact of individual antibiotics on the human gut microbiome and resistome. RESULTS: Using shotgun metagenomics, we quantified changes in the gut microbiome in two cohorts of hematological patients receiving prophylactic antibiotics; one cohort was treated with ciprofloxacin in a hospital in Tübingen and the other with cotrimoxazole in a hospital in Cologne. Analyzing this rich longitudinal dataset, we found that gut microbiome diversity was reduced in both treatment cohorts to a similar extent, while effects on the gut resistome differed. We observed a sharp increase in the relative abundance of sulfonamide antibiotic resistance genes (ARGs) by 148.1% per cumulative defined daily dose of cotrimoxazole in the Cologne cohort, but not in the Tübingen cohort treated with ciprofloxacin. Through multivariate modeling, we found that factors such as individual baseline microbiome, resistome, and plasmid diversity; liver/kidney function; and concurrent medication, especially virostatic agents, influence resistome alterations. Strikingly, we observed different effects on the plasmidome in the two treatment groups. There was a substantial increase in the abundance of ARG-carrying plasmids in the cohort treated with cotrimoxazole, but not in the cohort treated with ciprofloxacin, indicating that cotrimoxazole might contribute more efficiently to the spread of resistance. CONCLUSIONS: Our study represents a step forward in developing the capability to predict the effect of individual antimicrobials on the human microbiome and resistome. Our results indicate that to achieve this, integration of the individual baseline microbiome, resistome, and mobilome status as well as additional individual patient factors will be required. Such personalized predictions may in the future increase patient safety and reduce the spread of resistance. TRIAL REGISTRATION: ClinicalTrials.gov, NCT02058888 . Registered February 10 2014.

2.
J Antimicrob Chemother ; 74(7): 2065-2074, 2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-31220256

RESUMO

OBJECTIVES: We assessed the efficacy and safety of an oral antimicrobial regimen for short- and long-term intestinal eradication of ESBL-producing Escherichia coli and Klebsiella pneumoniae (ESBL-EC/KP) in immunocompromised patients. METHODS: We performed a randomized (2:1), double-blind multicentre Phase II study in four haematology-oncology departments. Patients colonized with ESBL-EC/KP received a 7 day antimicrobial regimen of oral colistin (2 × 106 IU 4×/day), gentamicin (80 mg 4×/day) and fosfomycin (three administrations of 3 g every 72 h), or placebo. Faecal, throat and urine specimens were collected on day 0, 6 ± 2, 11 ± 2, 28 ± 4 and 42 ± 4 after treatment initiation, and the quantitative burden of ESBL-EC/KP, resistance genes and changes in intestinal microbiota were analysed. Clinicaltrials.gov: NCT01931592. RESULTS: As the manufacture of colistin powder was suspended worldwide, the study was terminated prematurely. Overall, 29 (18 verum/11 placebo) out of 47 patients were enrolled. The short-term intestinal eradication was marginal at day 6 (verum group 15/18, 83.3% versus placebo 2/11, 18.2%; relative risk 4.58, 95% CI 1.29-16.33; Fisher's exact test P = 0.001) and not evident at later timepoints. Quantitative analysis showed a significant decrease of intestinal ESBL-EC/KP burden on day 6. Sustained intestinal eradication (day 28 + 42) was not achieved (verum, 38.9% versus placebo, 27.3%; P = 0.299). In the verum group, mcr-1 genes were detected in two faecal samples collected after treatment. Microbiome analysis showed a significant decrease in alpha diversity and a shift in beta diversity. CONCLUSIONS: In this prematurely terminated study of a 7 day oral antimicrobial eradication regimen, short-term ESBL-EC/KP suppression was marginal, while an altered intestinal microbiota composition was clearly apparent.

3.
Sci Rep ; 9(1): 6406, 2019 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-30992476

RESUMO

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

4.
Artigo em Inglês | MEDLINE | ID: mdl-30782988

RESUMO

Vancomycin-resistant Enterococcus faecium (VREfm) is a frequent cause of nosocomial outbreaks. In the second half of 2015, a sharp increase in the incidence of VREfm was observed at our university medical center. Next-generation sequencing (NGS) was used to analyze the first isolates of VREfm recovered from patients between 2010 and 2016 (n = 773) in order to decipher epidemiological change, outbreak dynamics, and possible transmission routes. VREfm isolates were analyzed using whole-genome sequencing followed by sequence type extraction and phylogenetic analysis. We examined epidemiological data, room occupancy data, and patient transferals and calculated an intensity score for patient-to-patient contact. Phylogenetic analysis revealed the presence of 38 NGS clusters and 110 single clones. The increase of VREfm was caused mainly by the expansion of two newly introduced NGS clusters, comprising VanB-type strains determined by multilocus sequence typing (MLST) as sequence type 80 (ST80) and ST117. By combining phylogenetic information with epidemiological data, intrahospital transmission could be demonstrated, however to a lesser extent than initially expected based solely on epidemiological data. The outbreak clones were continuously imported from other hospitals, suggesting a change in the epidemiological situation at a regional scale. By tracking intrahospital patient transferals, two major axes could be identified that contributed to the spread of VREfm within the hospital. NGS-based outbreak analysis revealed a dramatic change in the local and regional epidemiology of VREfm, emphasizing the role of health care networks in the spread of VREfm.

5.
Sci Rep ; 8(1): 13767, 2018 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-30213965

RESUMO

High throughput sequencing has been proposed as a one-stop solution for diagnostics and molecular typing directly from patient samples, allowing timely and appropriate implementation of measures for treatment, infection prevention and control. However, it is unclear how the variety of available methods impacts the end results. We applied shotgun metagenomics on diverse types of patient samples using three different methods to deplete human DNA prior to DNA extraction. Libraries were prepared and sequenced with Illumina chemistry. Data was analyzed using methods likely to be available in clinical microbiology laboratories using genomics. The results of microbial identification were compared to standard culture-based microbiological methods. On average, 75% of the reads corresponded to human DNA, being a major determinant in the analysis outcome. None of the kits was clearly superior suggesting that the initial ratio between host and microbial DNA or other sample characteristics were the major determinants of the proportion of microbial reads. Most pathogens identified by culture were also identified through metagenomics, but substantial differences were noted between the taxonomic classification tools. In two cases the high number of human reads resulted in insufficient sequencing depth of bacterial DNA for identification. In three samples, we could infer the probable multilocus sequence type of the most abundant species. The tools and databases used for taxonomic classification and antimicrobial resistance identification had a key impact on the results, recommending that efforts need to be aimed at standardization of the analysis methods if metagenomics is to be used routinely in clinical microbiology.

6.
J Clin Microbiol ; 56(11)2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30135233

RESUMO

Klebsiella pneumoniae and related species are frequent causes of nosocomial infections and outbreaks. Therefore, quick and reliable strain typing is crucial for the detection of transmission routes in the hospital. The aim of this study was to evaluate Fourier transform infrared spectroscopy (FTIR) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) as rapid methods for typing clinical Klebsiella isolates in comparison to whole-genome sequencing (WGS), which was considered the gold standard for typing and identification. Here, 68 clinical Klebsiella strains were analyzed by WGS, FTIR, and MALDI-TOF MS. FTIR showed high discriminatory power in comparison to the WGS reference, whereas MALDI-TOF MS exhibited a low ability to type the isolates. MALDI-TOF mass spectra were further analyzed for peaks that showed high specificity for different Klebsiella species. Phylogenetic analysis revealed that the Klebsiella isolates comprised three different species: K. pneumoniae, K. variicola, and K. quasipneumoniae Genome analysis showed that MALDI-TOF MS can be used to distinguish K. pneumoniae from K. variicola due to shifts of certain mass peaks. The peaks were tentatively identified as three ribosomal proteins (S15p, L28p, L31p) and one stress response protein (YjbJ), which exhibit amino acid differences between the two species. Overall, FTIR has high discriminatory power to recognize the clonal relationship of isolates, thus representing a valuable tool for rapid outbreak analysis and for the detection of transmission events due to fast turnaround times and low costs per sample. Furthermore, specific amino acid substitutions allow the discrimination of K. pneumoniae and K. variicola by MALDI-TOF MS.

7.
Artigo em Inglês | MEDLINE | ID: mdl-30603083

RESUMO

Background: Infections caused by third generation cephalosporin-resistant Enterobacteriaceae (3GCREB) are an increasing healthcare problem. We aim to describe the 3GCREB infection incidence and compare it to prevalence upon admission. In addition, we aim to describe infections caused by 3GCREB, which are also carbapenem resistant (CRE). Methods: In 2014-2015, we performed prospective 3GCREB surveillance in clinically relevant patient specimens (screening specimens excluded). Infections counted as hospital-acquired (HAI) when the 3GCREB was detected after the third day following admission, otherwise as community-acquired infection (CAI). Results: Of 578,420 hospitalized patients under surveillance, 3367 had a 3GCREB infection (0.58%). We observed a similar 3GCREB CAI and HAI incidence (0.28 and 0.31 per 100 patients, respectively). The most frequent pathogen was 3GCR E. coli, in CAI and HAI (0.15 and 0.12 per 100 patients). We observed a CRE CAI incidence of 0.006 and a HAI incidence of 0.008 per 100 patients (0.014 per 1000 patient days). Conclusions: Comparing the known 3GCREB admission prevalence of the participating hospitals (9.5%) with the percentage of patients with a 3GCREB infection (0.58%), we conclude the prevalence of 3GCREB in university hospitals to be about 16 times higher than suggested when only patients with 3GCREB infections are considered. Moreover, we find the HAI and CAI incidence caused by CRE in Germany to be relatively low.


Assuntos
Infecções Comunitárias Adquiridas/epidemiologia , Infecção Hospitalar/epidemiologia , Infecções por Enterobacteriaceae/epidemiologia , Idoso , Cefalosporinas , Farmacorresistência Bacteriana , Enterobacteriaceae/isolamento & purificação , Feminino , Alemanha/epidemiologia , Hospitais Universitários , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Admissão do Paciente , Estudos Prospectivos
8.
BMC Genomics ; 18(1): 859, 2017 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-29126393

RESUMO

BACKGROUND: Pseudomonas putida is a Gram-negative, non-fermenting bacterium frequently encountered in various environmental niches. P. putida rarely causes disease in humans, though serious infections and outbreaks have been reported from time to time. Some have suggested that P. putida functions as an exchange platform for antibiotic resistance genes (ARG), and thus represents a serious concern in the spread of ARGs to more pathogenic organisms within a hospital. Though poorly understood, the frequency of ARG exchange between P. putida and the more virulent Pseudomonas aeruginosa and its clinical relevance are particularly important for designing efficient infection control strategies, such as deciding whether high-risk patients colonized with a multidrug resistant but typically low pathogenic P. putida strain should be contact isolated or not. RESULTS: In this study, 21,373 screening samples (stool, rectal and throat swab) were examined to determine the presence of P. putida in a high-risk group of haemato-oncology patients during a 28-month period. A total of 89 P. putida group strains were isolated from 85 patients, with 41 of 89 (46.1%) strains harbouring the metallo-beta-lactamase gene bla VIM. These 41 clinical isolates, plus 18 bla VIM positive environmental P. putida isolates, and 17 bla VIM positive P. aeruginosa isolates, were characterized by whole genome sequencing (WGS). We constructed a maximum-likelihood tree to separate the 59 bla VIM positive P. putida group strains into eight distinct phylogenetic clusters. Bla VIM-1 was present in 6 clusters while bla VIM-2 was detected in 4 clusters. Five P. putida group strains contained both, bla VIM-1 and bla VIM-2 genes. In contrast, all P. aeruginosa strains belonged to a single genetic cluster and contained the same ARGs. Apart from bla VIM-2 and sul genes, no other ARGs were shared between P. aeruginosa and P. putida. Furthermore, the bla VIM-2 gene in P. aeruginosa was predicted to be only chromosomally located. CONCLUSION: These data provide evidence that no exchange of comprehensive ARG harbouring mobile genetic elements had occurred between P. aeruginosa and P. putida group strains during the study period, thus eliminating the need to implement enhanced infection control measures for high-risk patients colonized with a bla VIM positiv P. putida group strains in our clinical setting.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana/genética , Meio Ambiente , Transferência Genética Horizontal , Genômica , Pseudomonas aeruginosa/genética , Pseudomonas putida/genética , Humanos , Filogenia , Pseudomonas putida/efeitos dos fármacos , Pseudomonas putida/fisiologia
9.
Artigo em Inglês | MEDLINE | ID: mdl-28630206

RESUMO

Colistin is a last-resort antibiotic commonly used against multidrug-resistant strains of Pseudomonas aeruginosa To investigate the potential for in situ evolution of resistance against colistin and to map the molecular targets of colistin resistance, we exposed two P. aeruginosa isolates to colistin using a continuous-culture device known as a morbidostat. As a result, colistin resistance reproducibly increased 10-fold within 10 days and 100-fold within 20 days, along with highly stereotypic yet strain-specific mutation patterns. The majority of mutations hit the pmrAB two-component signaling system and genes involved in lipopolysaccharide (LPS) synthesis, including lpxC, pmrE, and migA We tracked the frequencies of all arising mutations by whole-genome deep sequencing every 3 to 4 days to obtain a detailed picture of the dynamics of resistance evolution, including competition and displacement among multiple resistant subpopulations. In 7 out of 18 cultures, we observed mutations in mutS along with a mutator phenotype that seemed to facilitate resistance evolution.


Assuntos
Colistina/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/genética , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Humanos , Testes de Sensibilidade Microbiana/métodos , Mutação/genética , Infecções por Pseudomonas/tratamento farmacológico , Infecções por Pseudomonas/microbiologia
10.
Artigo em Inglês | MEDLINE | ID: mdl-28413631

RESUMO

To assess the scope of infection control measures for multidrug-resistant bacteria in high-risk settings, a survey among university hospitals was conducted. Fourteen professionals from 8 sites participated. Reported policies varied largely with respect to the types of wards conducting screening, sample types used for screening and implementation of contact precautions. This variability among sites highlights the need for an evidence-based consensus of current infection control policies.

11.
J Clin Microbiol ; 55(7): 2116-2126, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28446572

RESUMO

Bloodstream infections (BSI) are an important cause of morbidity and mortality. Increasing rates of antimicrobial-resistant pathogens limit treatment options, prompting an empirical use of broad-range antibiotics. Fast and reliable diagnostic tools are needed to provide adequate therapy in a timely manner and to enable a de-escalation of treatment. The Accelerate Pheno system (Accelerate Diagnostics, USA) is a fully automated test system that performs both identification and antimicrobial susceptibility testing (AST) directly from positive blood cultures within approximately 7 h. In total, 115 episodes of BSI with Gram-negative bacteria were included in our study and compared to conventional culture-based methods. The Accelerate Pheno system correctly identified 88.7% (102 of 115) of all BSI episodes and 97.1% (102 of 105) of isolates that are covered by the system's identification panel. The Accelerate Pheno system generated an AST result for 91.3% (95 of 104) samples in which the Accelerate Pheno system identified a Gram-negative pathogen. The overall category agreement between the Accelerate Pheno system and culture-based AST was 96.4%, the rates for minor discrepancies 1.4%, major discrepancies 2.3%, and very major discrepancies 1.0%. Of note, ceftriaxone, piperacillin-tazobactam, and carbapenem resistance was correctly detected in blood culture specimens with extended-spectrum beta-lactamase-producing Escherichia coli (n = 7) and multidrug-resistant Pseudomonas aeruginosa (n = 3) strains. The utilization of the Accelerate Pheno system reduced the time to result for identification by 27.49 h (P < 0.0001) and for AST by 40.39 h (P < 0.0001) compared to culture-based methods in our laboratory setting. In conclusion, the Accelerate Pheno system provided fast, reliable results while significantly improving turnaround time in blood culture diagnostics of Gram-negative BSI.


Assuntos
Técnicas Bacteriológicas/métodos , Sangue/microbiologia , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Negativas/isolamento & purificação , Infecções por Bactérias Gram-Negativas/diagnóstico , Sepse/diagnóstico , Automação Laboratorial/métodos , Bactérias Gram-Negativas/classificação , Humanos , Sensibilidade e Especificidade , Fatores de Tempo
12.
J Mol Med (Berl) ; 95(1): 41-51, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27766372

RESUMO

The increasing threat of antimicrobial resistance poses one of the greatest challenges to modern medicine. The collection of all antimicrobial resistance genes carried by various microorganisms in the human body is called the human resistome and represents the source of resistance in pathogens that can eventually cause life-threatening and untreatable infections. A deep understanding of the human resistome and its multilateral interaction with various environments is necessary for developing proper measures that can efficiently reduce the spread of resistance. However, the human resistome and its evolution still remain, for the most part, a mystery to researchers. Metagenomics, particularly in combination with next-generation-sequencing technology, provides a powerful methodological approach for studying the human microbiome as well as the pathogenome, the virolume and especially the resistome. We summarize below current knowledge on how the human resistome is shaped and discuss how metagenomics can be employed to improve our understanding of these complex processes, particularly as regards a rapid translation of new findings into clinical diagnostics, infection control and public health.


Assuntos
Resistência Microbiana a Medicamentos , Metagenoma , Metagenômica , Microbiota/efeitos dos fármacos , Microbiota/genética , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Infecções Bacterianas/diagnóstico , Infecções Bacterianas/tratamento farmacológico , Infecções Bacterianas/microbiologia , Humanos , Controle de Infecções , Metagenômica/métodos , Pesquisa Médica Translacional
13.
Diagn Microbiol Infect Dis ; 87(1): 71-73, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27769572

RESUMO

As part of the multicenter Antibiotic Therapy Optimisation Study-the largest study on the prevalence of third-generation cephalosporin-resistant Enterobacteriaceae carriage upon hospital admission-minimum inhibitory concentration values were generated for ampicillin/sulbactam, amoxicillin/clavulanic acid, piperacillin/tazobactam, mecillinam, mecillinam/clavulanic acid, and temocillin against third-generation cephalosporin-resistant Escherichia coli, Klebsiella species and Enterobacter species.


Assuntos
Antibacterianos/farmacologia , Infecções por Enterobacteriaceae/microbiologia , Enterobacteriaceae/efeitos dos fármacos , Penicilinas/farmacologia , Resistência beta-Lactâmica , Inibidores de beta-Lactamases/farmacologia , Cefalosporinas/farmacologia , Testes Diagnósticos de Rotina , Enterobacteriaceae/isolamento & purificação , Humanos , Testes de Sensibilidade Microbiana
14.
Int J Antimicrob Agents ; 49(2): 239-242, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-27939093

RESUMO

As part of the multicentre Antibiotic Therapy Optimisation Study (ATHOS), minimum inhibitory concentrations (MICs) were determined for cephalosporins alone and in combination with the ß-lactamase inhibitors tazobactam, clavulanic acid and avibactam against third-generation cephalosporin-resistant Escherichia coli, Klebsiella spp. and Enterobacter spp. isolates collected in German hospitals. MIC50/90 values were 0.25-4 mg/L for cefepime/tazobactam, 0.25-2 mg/L for ceftazidime/avibactam, 0.125-0.5 mg/L for ceftaroline/avibactam, 0.5-4 mg/L for cefpodoxime/clavulanic acid and 0.25-1 mg/L for aztreonam/avibactam, depending on the underlying resistance mechanism and organism. Based on in vitro testing, ß-lactam antibiotics play an important role in the treatment of infections due to ß-lactamase-producing organisms.


Assuntos
Antibacterianos/farmacologia , Compostos Azabicíclicos/farmacologia , Aztreonam/farmacologia , Cefalosporinas/farmacologia , Infecções por Enterobacteriaceae/microbiologia , Enterobacteriaceae/efeitos dos fármacos , Inibidores de beta-Lactamases/farmacologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Resistência às Cefalosporinas , Ácido Clavulânico/farmacologia , Enterobacteriaceae/isolamento & purificação , Feminino , Alemanha , Hospitais , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Ácido Penicilânico/análogos & derivados , Ácido Penicilânico/farmacologia , Tazobactam , Adulto Jovem
15.
Artigo em Inglês | MEDLINE | ID: mdl-27956426

RESUMO

The metallo-beta-lactamase GIM-1 has been found in various bacterial host species nearly exclusively in western Germany. However, not much is known about the epidemiology of GIM-1-positive Serratia marcescens Here we report on a surprisingly protracted regional dissemination. In-hospital transmission was investigated by using conventional epidemiological tools to identify spatiotemporal links. Strain typing was performed using pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing (WGS). Bayesian phylogeny was used to infer the time axis of the observed occurrence. Thirteen S. marcescens strains from 10 patients from 6 different German hospitals were investigated. Suspected in-hospital transmissions were confirmed by molecular typing at a higher resolution by WGS than by PFGE. A detailed sequence analysis demonstrated the spread of one predominant strain variant but also provided evidence for transfer of the blaGIM-1 gene cassette between different strains. A Bayesian phylogenetic analysis showed that the most recent common ancestor of the identified clonal cluster could be dated back to April 1993 (95% highest posterior density interval, January 1973 to March 2003) and that this strain might have already harbored the blaGIM-1 at that time and, therewith, years before the first detection of this resistance gene in clinical specimens. This study shows a long-standing clonal and plasmid-mediated expansion of GIM-1-producing S. marcescens that might have gone unnoticed in the absence of a standardized and effective molecular screening for carbapenemases. The systematic and early detection of resistance is thus highly advisable, especially for the prevention of potentially long-term dissemination that may progress beyond control.


Assuntos
Infecção Hospitalar/transmissão , Genoma Bacteriano , Filogenia , Infecções por Serratia/transmissão , Serratia marcescens/genética , Resistência beta-Lactâmica/genética , beta-Lactamases/genética , Antibacterianos/farmacologia , Técnicas de Tipagem Bacteriana , Teorema de Bayes , Células Clonais , Infecção Hospitalar/tratamento farmacológico , Infecção Hospitalar/epidemiologia , Infecção Hospitalar/microbiologia , Eletroforese em Gel de Campo Pulsado , Expressão Gênica , Genótipo , Alemanha , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Testes de Sensibilidade Microbiana , Epidemiologia Molecular , Plasmídeos/química , Plasmídeos/metabolismo , Infecções por Serratia/tratamento farmacológico , Infecções por Serratia/epidemiologia , Infecções por Serratia/microbiologia , Serratia marcescens/classificação , Serratia marcescens/efeitos dos fármacos , Serratia marcescens/crescimento & desenvolvimento , beta-Lactamases/metabolismo
16.
J Biotechnol ; 250: 45-50, 2017 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-27984120

RESUMO

One important question in microbiome analysis is how to assess the homogeneity of the microbial composition in a given environment, with respect to a given analysis method. Do different microbial samples taken from the same environment follow the same taxonomic distribution of organisms, or the same distribution of functions? Here we provide a non-parametric statistical "triangulation test" to address this type of question. The test requires that multiple replicates are available for each of the biological samples, and it is based on three-way computational comparisons of samples. To illustrate the application of the test, we collected three biological samples taken from different locations in one piece of human stool, each represented by three replicates, and analyzed them using MEGAN. (Despite its name, the triangulation test does not require that the number of biological samples or replicates be three.) The triangulation test rejects the null hypothesis that the three biological samples exhibit the same distribution of taxa or function (error probability ≤0.05), indicating that the microbial composition of the investigated human stool is not homogenous on a macroscopic scale, suggesting that pooling material from multiple locations is a reasonable practice. We provide an implementation of the test in our open source program MEGAN Community Edition.


Assuntos
Algoritmos , Bactérias/genética , Bactérias/isolamento & purificação , Técnicas de Tipagem Bacteriana/métodos , Interpretação Estatística de Dados , Microbiota/genética , Análise de Sequência de DNA/métodos , Bactérias/classificação , Simulação por Computador , Fezes/microbiologia , Ensaios de Triagem em Larga Escala/métodos , Humanos , Modelos Estatísticos , Reprodutibilidade dos Testes , Tamanho da Amostra , Sensibilidade e Especificidade
17.
Antimicrob Agents Chemother ; 59(12): 7335-45, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26369961

RESUMO

The human gut forms a dynamic reservoir of antibiotic resistance genes (ARGs). Treatment with antimicrobial agents has a significant impact on the intestinal resistome and leads to enhanced horizontal transfer and selection of resistance. We have monitored the development of intestinal ARGs over a 6-day course of ciprofloxacin (Cp) treatment in two healthy individuals by using sequenced-based metagenomics and different ARG quantification methods. Fixed- and random-effect models were applied to determine the change in ARG abundance per defined daily dose of Cp as an expression of the respective selection pressure. Among various shifts in the composition of the intestinal resistome, we found in one individual a strong positive selection for class D beta-lactamases which were partly located on a mobile genetic element. Furthermore, a trend to a negative selection has been observed with class A beta-lactamases (-2.66 hits per million sample reads/defined daily dose; P = 0.06). By 4 weeks after the end of treatment, the composition of ARGs returned toward their initial state but to a different degree in both subjects. We present here a novel analysis algorithm for the determination of antibiotic selection pressure which can be applied in clinical settings to compare therapeutic regimens regarding their effect on the intestinal resistome. This information is of critical importance for clinicians to choose antimicrobial agents with a low selective force on their patients' intestinal ARGs, likely resulting in a diminished spread of resistance and a reduced burden of hospital-acquired infections with multidrug-resistant pathogens.


Assuntos
Antibacterianos/farmacologia , Resistência Microbiana a Medicamentos/efeitos dos fármacos , Resistência Microbiana a Medicamentos/genética , Microbioma Gastrointestinal/efeitos dos fármacos , Metagenômica/métodos , Adulto , Algoritmos , Biodiversidade , Ciprofloxacino/farmacologia , Microbioma Gastrointestinal/genética , Humanos , Masculino , Reação em Cadeia da Polimerase em Tempo Real/métodos , Seleção Genética/efeitos dos fármacos , beta-Lactamases/genética
18.
Expert Rev Anti Infect Ther ; 13(9): 1159-70, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26153817

RESUMO

Pseudomonas aeruginosa is a Gram-negative human pathogen with extensively drug-resistant (XDR) strains emerging in hospitals across the globe. This systematic review is focused on the worldwide prevalence of XDR P. aeruginosa (XDR-PA) and on the risk factors associated with its colonization and infection, based on literature available through PubMed, Web of Science and BioMed Central databases. An overview of surveillance systems is provided as well as a synopsis on the prevalence of XDR-PA, showing an increase in recent reports. Risk factors independently associated with XDR-PA colonization or infections are described in four groups with reference to antimicrobial therapy, medical devices as well as patient- and hospital environment-related factors.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Infecções por Pseudomonas/tratamento farmacológico , Infecções por Pseudomonas/epidemiologia , Pseudomonas aeruginosa/efeitos dos fármacos , Animais , Antibacterianos/uso terapêutico , Contagem de Colônia Microbiana/métodos , Farmacorresistência Bacteriana Múltipla/fisiologia , Humanos , Prevalência , Infecções por Pseudomonas/diagnóstico , Pseudomonas aeruginosa/fisiologia , Fatores de Risco
19.
J Antimicrob Chemother ; 70(5): 1322-30, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25583750

RESUMO

OBJECTIVES: Here we report on a long-term outbreak from 2009 to 2012 with an XDR Pseudomonas aeruginosa on two wards at a university hospital in southern Germany. METHODS: Whole-genome sequencing was performed on the outbreak isolates and a core genome was constructed for molecular epidemiological analysis. We applied a time-place-sequence algorithm to improve estimation of transmission probabilities. RESULTS: By using conventional infection control methods we identified 49 P. aeruginosa strains, including eight environmental isolates that belonged to ST308 (by MLST) and carried the metallo-ß-lactamase IMP-8. Phylogenetic analysis on the basis of a non-recombinant core genome that contained 22 outbreak-specific SNPs revealed a pattern of four dominant clades with a strong phylogeographic structure and allowed us to determine the potential temporal origin of the outbreak to July 2008, 1 year before the index case was diagnosed. Superspreaders at the root of clades exhibited a high number of probable and predicted transmissions, indicating their exceptional position in the outbreak. CONCLUSIONS: Our results suggest that the initial expansion of dominant sublineages was driven by a few superspreaders, while environmental contamination seemed to sustain the outbreak for a long period despite regular environmental control measures.


Assuntos
Surtos de Doenças , Farmacorresistência Bacteriana Múltipla , Infecções por Pseudomonas/epidemiologia , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/classificação , Pseudomonas aeruginosa/efeitos dos fármacos , Transmissão de Doença Infecciosa , Microbiologia Ambiental , Estudos Epidemiológicos , Genoma Bacteriano , Alemanha/epidemiologia , Hospitais Universitários , Humanos , Epidemiologia Molecular , Tipagem Molecular , Infecções por Pseudomonas/transmissão , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/isolamento & purificação , Análise de Sequência de DNA , Análise Espaço-Temporal
20.
BMC Infect Dis ; 14: 650, 2014 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-25490897

RESUMO

BACKGROUND: This study aimed to investigate risk factors for colonisation with extensively drug-resistant P. aeruginosa (XDR-PA) in immunocompromised patients and to build a clinical risk score (CRS) based on these results. METHODS: We conducted a matched case-control study with 31 cases and 93 controls (1:3). Cases were colonised with XDR-PA during hospitalisation. Independent risk factors were determined using a three step conditional logistic regression procedure. A CRS was built with respect to the corresponding risk fraction of each risk factor, and its discriminatory power was estimated by receiver operating characteristic (ROC) analysis. RESULTS: The presence of a central venous catheter (OR 7.41, P = 0.0008), the presence of a urinary catheter (OR 21.04, P < 0.0001), CRP > 10 mg/dl (OR 7.36, P = 0.0015), and ciprofloxacin administration (OR 5.53, P = 0.025) were independent risk factors. The CRS exhibited a high discriminatory power, defining a high risk population with an approximately fourteen times greater risk for XDR-PA colonisation. CONCLUSIONS: Unnecessary use of antibiotics, particularly ciprofloxacin should be avoided, and a high standard of infection control measures must be achieved when using medical devices. A CRS can be used for adaptation of the active screening culture policy to the local setting.


Assuntos
Infecção Hospitalar/etiologia , Farmacorresistência Bacteriana Múltipla , Hospedeiro Imunocomprometido , Infecções por Pseudomonas/etiologia , Pseudomonas aeruginosa/isolamento & purificação , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Infecção Hospitalar/diagnóstico , Infecção Hospitalar/imunologia , Feminino , Doenças Hematológicas/complicações , Doenças Hematológicas/terapia , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Infecções por Pseudomonas/diagnóstico , Infecções por Pseudomonas/imunologia , Fatores de Risco
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