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1.
Clin Microbiol Infect ; 21(6): 570.e1-4, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25749563

RESUMO

To assess the distribution of phylogroups and O25/ST131 in the Netherlands, we performed a real-time polymerase chain reaction (PCR) on a collection of 108 wild-type Escherichia coli (WT-EC) and 134 extended-spectrum ß-lactamase-producing E. coli (ESBL-EC). Phylogroup B2 was predominant, but ESBL-EC were less likely to belong to this phylogroup (48.5%) than were WT-EC (66.7%; p = 0.005). In WT-EC, phylogroups B2 and D seem to be more virulent, having a higher prevalence among midstream urine isolates and blood culture isolates, than in catheter-related urine isolates (83.3% and 87.9% vs. 61.9%; p 0.048). O25/ST131 is associated with ESBL production, being almost absent among phylogroup B2 WT-EC (61.5% vs. 5.6%; p < 0.001).


Assuntos
Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/microbiologia , Escherichia coli/classificação , Escherichia coli/genética , Genótipo , Adulto , Idoso , Idoso de 80 Anos ou mais , Escherichia coli/isolamento & purificação , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Países Baixos/epidemiologia , Prevalência , Estudos Prospectivos , Reação em Cadeia da Polimerase em Tempo Real , Estudos Retrospectivos , Sorogrupo , beta-Lactamases/metabolismo
2.
Clin Microbiol Infect ; 19(7): 662-5, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-22971154

RESUMO

Correct detection of extended-spectrum beta-lactamases (ESBLs) is crucial for infection control and antibiotic choice. We performed a study to determine the cost-effectiveness of phenotypical testing, which can be inaccurate, and genotypical tests, which are considered to be more reliable but also more expensive. All patients that had been in isolation in the Amphia hospital because of the detection of ESBL according to the ESBL Etest were included in the survey. All strains were retested using the double disk confirmation test (DDCT) and a genotypical method. This was a commercially available microarray (Check-Points). Discordant results were confirmed by PCR and sequencing. In total 174 patients were included. In 24 of 174 (14%) patients, ESBL carriage could not be confirmed with the microarray. This was verified with PCR and sequencing. The mean duration of isolation was 15 days, adding up to a total number of isolation days of 2571. False-positive results according to the microarray resulted in a total of 279 days of unnecessary isolation for the Etest and 151 days for the DDCT. Using Etest to detect the presence of ESBL results in a false-positive outcome in 14% of the cases. This results in unnecessary isolation of patients, which can be omitted by using a genotypic method.


Assuntos
Bactérias/enzimologia , Técnicas Bacteriológicas/economia , Técnicas Bacteriológicas/métodos , Técnicas de Diagnóstico Molecular/economia , Técnicas de Diagnóstico Molecular/métodos , beta-Lactamases/análise , beta-Lactamases/genética , Bactérias/isolamento & purificação , Infecções Bacterianas/microbiologia , Análise Custo-Benefício , Erros de Diagnóstico , Genótipo , Hospitais , Humanos , Fenótipo
3.
Clin Microbiol Infect ; 16(6): 704-10, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-19624518

RESUMO

We developed a dermatophyte-specific single-tube real-time PCR assay based on internal transcribed sequences. This assay allows the rapid detection and identification of 11 clinically relevant species within the three dermatophyte genera Trichophyton, Microsporum and Epidermophyton in nail, skin and hair samples within a few hours. Analysis of 145 clinical samples (107 nail, 36 skin scale, and two hair) by both real-time PCR and a PCR-reverse line blot (PCR-RLB) assay described earlier revealed that 133 of the 145 samples had concordant real-time PCR and PCR-RLB detection results (83 positive, 49 negative, and one inhibited). Six samples were positive by real-time PCR and negative by PCR-RLB, and two were negative by real-time PCR and positive by PCR-RLB. Four samples demonstrated inhibition in one of the two PCR assays. Only one of 83 positive samples had discordant identification results between both assays (Trichophyton verrucosum and Trichophyton erinacei by real-time PCR and Trichophyton erinacei by PCR-RLB). Dermatophytes present in seven positive samples that were incompletely identified as Trichophyton sp. by PCR-RLB were identified to the species level by real-time PCR as Trichophyton interdigitale and Trichophyton rubrum in six cases and one case, respectively. One hundred and twenty of 145 samples were also analysed by conventional dermatophyte culture and by direct microscopy. Our single-tube real-time PCR assay proved to be suitable for direct detection and identification of dermatophytes in nail, skin and hair samples with minimal total assay time (4 h after overnight lysis) and hands-on time, without the need for post-PCR analysis, and with good sensitivity and specificity.


Assuntos
Arthrodermataceae/classificação , Arthrodermataceae/isolamento & purificação , Dermatomicoses/diagnóstico , Dermatomicoses/microbiologia , Micologia/métodos , Reação em Cadeia da Polimerase/métodos , Arthrodermataceae/genética , DNA Fúngico/genética , DNA Espaçador Ribossômico/genética , Cabelo/microbiologia , Humanos , Unhas/microbiologia , Sensibilidade e Especificidade , Pele/microbiologia
4.
Eur J Clin Microbiol Infect Dis ; 28(10): 1223-6, 2009 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-19562391

RESUMO

Nasal carriage is an important risk factor for the development of post-operative infections with Staphylococcus aureus and pre-operative treatment with mupirocin in carriers reduces the post-operative infection rate. Therefore, it is important to identify nasal carriage rapidly. Two polymerase chain reaction (PCR) techniques were compared to conventional culture in surgical patients. In 404 consecutive patients, nasal swabs were taken for pre-operative screening for the nasal carriage of S. aureus. The performance of the Roche Staphylococcus Kit on Lightcycler (Roche; RSA) and the Becton Dickinson (San Diego, CA) GeneOhm StaphSR assay on Smartcycler (Cepheid; BDSA) were compared with semi-quantitative culture. The sensitivity for culture, RSA and BDSA compared to the gold standard was 98.2, 82.0 and 85.6%, respectively, and the specificity was 100, 98.3 and 99.3%, respectively. The lower sensitivity of both PCR techniques was associated with samples with low bacterial loads. The RSA and BDSA were similar in performance and are suitable for the pre-operative identification of nasal carriers of S. aureus.


Assuntos
Portador Sadio/diagnóstico , Contagem de Colônia Microbiana/métodos , Cavidade Nasal/microbiologia , Reação em Cadeia da Polimerase/métodos , Staphylococcus aureus/isolamento & purificação , Portador Sadio/microbiologia , Humanos , Complicações Pós-Operatórias/epidemiologia , Complicações Pós-Operatórias/prevenção & controle , Valor Preditivo dos Testes , Cuidados Pré-Operatórios , Estudos Prospectivos , Sensibilidade e Especificidade , Infecções Estafilocócicas/epidemiologia , Infecções Estafilocócicas/prevenção & controle
5.
Clin Microbiol Infect ; 14(8): 778-88, 2008 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-18727802

RESUMO

A dermatophyte-specific PCR-reverse line blot (PCR-RLB) assay based on internal transcribed sequences was developed. This assay allows the rapid detection and identification of nine clinically relevant species within the three dermatophyte genera Trichophyton, Microsporum and Epidermophyton in nail, skin and hair samples within 1 day. Analysis of 819 clinical samples (596 nail, 203 skin and 20 hair) revealed a positive PCR-RLB result in 93.6% of 172 culture-positive and microscopy-positive samples. PCR-RLB was superior to culture and direct microscopy, in both detection and species identification. Comparison of identification results of 208 PCR-positive and culture-positive clinical samples showed five discrepancies (2.4%) between PCR-RLB identification and classical microscopic/biochemical identification of isolates. Comparison of PCR-RLB identification and classical identification of 98 other isolates (dermatophytes and non-dermatophytes) revealed 13 discrepancies (13.3%) and five incomplete identifications of Trichophyton spp. Sequence analysis of ITS1 regions of 23 samples with discrepant or incomplete identification results (four Centraalbureau voor Schimmelcultures dermatophyte strains, four clinical samples and 15 clinical isolates) confirmed identification results of PCR-RLB in 21 of the 23 analyzed samples. PCR-RLB proved to be extremely suitable for routine detection and identification of dermatophytes directly in nail, skin and hair samples because it is rapid, sensitive, specific and accurate.


Assuntos
Arthrodermataceae/classificação , Arthrodermataceae/isolamento & purificação , Dermatomicoses/diagnóstico , Cabelo/microbiologia , Unhas/microbiologia , Hibridização de Ácido Nucleico/métodos , Reação em Cadeia da Polimerase/métodos , Pele/microbiologia , Arthrodermataceae/genética , Primers do DNA , DNA Fúngico/análise , DNA Fúngico/isolamento & purificação , DNA Espaçador Ribossômico/análise , Dermatomicoses/microbiologia , Humanos , Técnicas de Tipagem Micológica , Sensibilidade e Especificidade , Fatores de Tempo
6.
Clin Microbiol Infect ; 13(6): 627-34, 2007 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-17378931

RESUMO

Cat-scratch disease (CSD), caused by Bartonella henselae infection, can mimic malignancy and can manifest atypically. Reliable serological testing is therefore of great clinical importance. The diagnostic performance of immunofluorescence assay (IFA) and ELISA was evaluated in a group of Dutch patients with proven CSD (clinical diagnosis confirmed by PCR). Sera of 51 CSD patients and 56 controls (patients with similar symptoms, but who were B. henselae PCR-negative and had an alternative confirmed diagnosis) were tested for anti-B. henselae IgM and IgG by IFA and ELISA. A commercially available IFA test for IgM had a sensitivity of 6%. In-house assays for IgM showed specificities of 93% (IFA) and 91% (ELISA), but with low sensitivities (53% and 65%, respectively). With a specificity of 82% (IFA) and 91% (ELISA), in-house IgG testing showed a significantly higher sensitivity in IFA (67%) than in ELISA (28%, p <0.01). Sensitivity was higher for genotype I (38-75%) than for genotype II (7-67%) infections, but this was only statistically significant for IgG ELISA (p <0.05). In conclusion, detection of IgM against B. henselae by in-house ELISA and IFA was highly specific for the diagnosis of CSD. The high seroprevalence in healthy individuals limits the clinical value of IgG detection for diagnosing CSD. Given the low sensitivity of the serological assays, negative serology does not rule out CSD and warrants further investigation, including PCR. Adding locally isolated (e.g., genotype II) B. henselae strains to future tests might improve the sensitivity.


Assuntos
Anticorpos Antibacterianos/sangue , Bartonella henselae/imunologia , Doença da Arranhadura de Gato/diagnóstico , Ensaio de Imunoadsorção Enzimática/métodos , Técnica Indireta de Fluorescência para Anticorpo/métodos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Doença da Arranhadura de Gato/microbiologia , Criança , Pré-Escolar , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Lactente , Pessoa de Meia-Idade , Países Baixos , Sensibilidade e Especificidade
7.
Clin Microbiol Infect ; 9(2): 135-9, 2003 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-12588334

RESUMO

The objective of this study was to compare four procedures for Chlamydia pneumoniae DNA extraction from vascular tissue. The NucliSens Kit, the QIAamp tissue DNA MiniKit, buffer-saturated phenol and the Geneclean II Kit were evaluated, based on the yield of recovered DNA, using PCR to detect C. pneumoniae in vascular tissue. The QIAamp tissue procedure had the highest detection level (0.004 inclusion-forming units/sample). All methods, except NucliSens (70 min), had a short handling time (30-40 min). Costs varied from 0.5 to 3.2 Euro.


Assuntos
Infecções por Chlamydia/patologia , Chlamydophila pneumoniae/genética , DNA Bacteriano/isolamento & purificação , Músculo Liso Vascular/microbiologia , Reação em Cadeia da Polimerase/métodos , DNA Bacteriano/química , DNA Bacteriano/genética , Humanos , Hibridização de Ácido Nucleico , Reação em Cadeia da Polimerase/economia
9.
Vet Q ; 19(sup1): 44-45, 1997 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-22047429
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