RESUMO
We describe the results of the Quality Control for Molecular Diagnostics 2013 Neisseria gonorrhoeae external quality assessment programme that included an N. gonorrhoeae strain harbouring an N. meningitidis porA gene which causes false-negative results in molecular diagnostic assays targeting the gonococcal porA pseudogene. Enhanced awareness of the international transmission of such gonococcal strains is needed to avoid false-negative results in both in-house and commercial molecular diagnostic assays used in laboratories worldwide, but particularly in Europe.
Assuntos
Gonorreia/diagnóstico , Ensaio de Proficiência Laboratorial , Neisseria gonorrhoeae/genética , Neisseria meningitidis/genética , Porinas/genética , Pseudogenes/genética , Controle de Qualidade , Europa (Continente) , Reações Falso-Negativas , Variação Genética , Gonorreia/genética , Gonorreia/microbiologia , Humanos , Dados de Sequência Molecular , Tipagem Molecular , Mutação , Neisseria gonorrhoeae/isolamento & purificação , Neisseria meningitidis/isolamento & purificação , Reação em Cadeia da Polimerase/métodos , Análise de Sequência de DNARESUMO
The cppB gene is often used as a target for detection of Neisseria gonorrhoeae by PCR. Using a coded panel of 500 DNA samples, we determined that the cppB gene is missing in 5.8% of N. gonorrhoeae strains, and therefore we consider the cppB gene to be an unsuitable target.
Assuntos
Proteínas da Membrana Bacteriana Externa/genética , Neisseria gonorrhoeae/isolamento & purificação , Reação em Cadeia da Polimerase/métodos , Humanos , Neisseria gonorrhoeae/genética , Reprodutibilidade dos TestesRESUMO
AIMS: In spite of increasing reuse of disposable catheters, there are few scientific data on potential viral transmission and infection after reuse. To determine the theoretical risk of virus transmission during reuse of catheters an in vitro study was performed using an RNA virus (echovirus-11) and a DNA virus (adenovirus-2). METHODS AND RESULTS: After deliberate contamination of the catheters, reprocessing and reuse of the cleaned and glutaraldehyde sterilized catheters was simulated. The presence of residual virus was determined by cell culture and by polymerase chain reaction (PCR). After the sterilization step, infectious enterovirus was detectable in one (10%) of the samples, whereas two (20%) contained detectable enterovirus RNA. After simulated reuse, enterovirus was cultured from one (10%) of the catheters, but no less than six (60%) of the samples were enterovirus PCR positive and one (10%) contained detectable adenovirus DNA. After sonification of the catheter tips no infectious virus could be detected, but enterovirus RNA was detected in two (20%) and adenovirus DNA in three (30%) of the samples. CONCLUSIONS: It has been clearly demonstrated in this in vitro study that, even after rigorous cleaning and sterilization, virus was still present in the catheter. Reuse of catheters, labelled for single-use only, is dangerous and should be prevented.
Assuntos
Cateterismo/instrumentação , Infecção Hospitalar/transmissão , Infecção Hospitalar/virologia , Contaminação de Equipamentos , Reutilização de Equipamento , Viroses/transmissão , Técnicas de Cultura de Células/métodos , Vírus de DNA/isolamento & purificação , Equipamentos Descartáveis , Humanos , Reação em Cadeia da Polimerase/métodos , Vírus de RNA/isolamento & purificação , Fatores de Risco , EsterilizaçãoRESUMO
The fully automated COBAS AMPLICOR CT/NG test for the detection of Chlamydia trachomatis was evaluated in a multicenter trial. Test performance was evaluated for 2,014 endocervical swab and 1,278 urine specimens obtained from women and for 373 urethral swab and 254 urine specimens obtained from men. Culture served as the reference test. Culture-negative, COBAS AMPLICOR-positive specimens that tested positive in a confirmatory PCR test for an alternative target sequence within the C. trachomatis major outer membrane protein gene were resolved as true positives. The overall prevalence of chlamydia was 4.3% in cervical swabs and 11.0% in urethral swabs from men. When the results for each specimen type were considered separately, the resolved sensitivities were 96.5% (83 of 86) for endocervical swab specimens, 95.1% (39 of 41) for urine specimens from women, 100.0% (41 of 41) for urethral swab specimens from men, and 94.4% (17 of 18) for urine specimens from men; the resolved specificities were 99.4% (1,912 of 1,924) for endocervical swab specimens, 99.8% (1,204 of 1,207) for urine specimens from women, 98. 5% (325 of 330) for urethral swab specimens from men, and 100.0% (236 of 236) for urine specimens from men. For the subset of patients from whom both swab and urine specimens were collected, the combined results for both specimen types were used to identify all infected patients. Using these combined reslts as criteria, the resolved sensitivities for the COBAS AMPLICOR test were 82.6% (38 of 46) for endocervical swab specimens, 84.4% (38 of 45) for urine specimens from women, 84.2% (16 of 19) for urethral swab specimens from men, and 89.5% (17 of 19) for urine specimens from men. In comparison, the sensitivity of culture was only 56.5% (26 of 46) for endocervical specimens and 63.2% (12 of 19) for urethral specimens from men. The internal control provided in the COBAS AMPLICOR test revealed that 2.9% of specimens were inhibitory when they were initially tested. Nevertheless, valid results were obtained for 99. 1% of specimens because 68.7% of the inhibitory specimens were not inhibitory when a second aliquot of the original sample was tested. Two additional COBAS AMPLICOR-positive specimens were detected by retesting inhibitory specimens. The COBAS AMPLICOR CT/NG test for the detection of C. trachomatis exhibited equally high sensitivities and specificities with both urogenital swab and urine specimens and, thus, is well-suited for use in screening.
Assuntos
Colo do Útero/microbiologia , Chlamydia trachomatis/isolamento & purificação , Reação em Cadeia da Polimerase/métodos , Uretra/microbiologia , Automação , Infecções por Chlamydia/microbiologia , Estudos de Avaliação como Assunto , Feminino , Humanos , Masculino , Kit de Reagentes para Diagnóstico , Sensibilidade e Especificidade , Urina/microbiologiaRESUMO
OBJECTIVE: To assess the accuracy of six commonly used diagnostic tests for Helicobacter pylori in a prospective study without using any specific test as the gold standard (the patient was regarded as H. pylori-infected if two or more tests, whatever their nature, were positive). METHODS: In 105 outpatients undergoing upper GI endoscopy, 62 without significant abnormalities, 28 with gastroesophageal reflux disease, 19 with peptic ulcer, one with erosive gastritis, and one with atrophic gastritis (some patients had more than one diagnosis), antral biopsy specimens were taken for culture, polymerase chain reaction, histological examination (hematoxylineosin and Giemsa stains), and rapid urease test. Corpus biopsy specimens were taken for histological examination. Serology (ELISA) and a 13C-urea breath test were also performed. Consistency of diagnosis between two pathologists was assessed by kappa statistics. RESULTS: Sensitivity and specificity, respectively, were as follows: culture, 98.4 and 100%; polymerase chain reaction, 96.7 and 100%; histological examination (antrum), 96 and 98.8%; histological examination (antrum + corpus), 98.4 and 98.8%; rapid urease test, 90.2 and 100%; 13C-urea breath test, 100 and 100%; and serological examination, 98.4 and 88.4% (95% in those who had not been previously treated for H. pylori). All H. pylori-positive cases were detected by culture and rapid urease test. In 86.4% of these cases all antral biopsy-based tests were positive. Agreement between pathologists was good, with a kappa coefficient around 0.90 for antral biopsy specimens. CONCLUSIONS: All antral biopsy-based tests, as well as the 13C-urea breath test, are accurate for the diagnosis of H. pylori infection. Sampling error is a problem of minor importance. The lower specificity of serological tests may be largely explained by previous treatment of H. pylori.
Assuntos
Infecções por Helicobacter/diagnóstico , Helicobacter pylori/isolamento & purificação , Biópsia , Testes Respiratórios , Ensaio de Imunoadsorção Enzimática , Estudos de Avaliação como Assunto , Feminino , Mucosa Gástrica/patologia , Refluxo Gastroesofágico/microbiologia , Humanos , Masculino , Pessoa de Meia-Idade , Úlcera Péptica/microbiologia , Reação em Cadeia da Polimerase , Estudos Prospectivos , Sensibilidade e EspecificidadeRESUMO
Eleven hundred and thirty-three clinical specimens submitted to the laboratory for diagnosis of respiratory virus infections were tested by direct immunofluorescence (DIF) for respiratory syncytial virus (RSV), by shell vial culture, and by conventional cell culture. The shell vial cultures were stained with 8 different monoclonal antibodies both 1 day and 3-7 days after inoculation. In order to limit the cost and the workload, mixtures of monoclonal antibodies were used. Coverslips with HEp-2 cells were incubated with a mixture of FITC-labeled monoclonal antibody to RSV and nonlabeled monoclonal antibody to adenovirus. When no RSV positive IF staining was observed after the first incubation step, the same coverslip was incubated once more with FITC-labeled anti-mouse antibody. A positive reaction at this stage indicated the presence of adenovirus. Similarly, cultures of tertiary monkey kidney cells were investigated with a mixture of two FITC-labeled monoclonals to the influenza viruses A and B and three nonlabeled monoclonals to the parainfluenza viruses 1, 2 and 3. If influenza virus or parainfluenza virus was detected, the exact type was determined by staining different parts of a duplicate coverslip. Shell vial cultures for cytomegalovirus (CMV) were always performed separately on human embryonic lung fibroblasts. Using this approach, we detected RSV (n = 248), CMV (n = 42), parainfluenza virus (n = 31), influenza virus (n = 28), and adenovirus (n = 6), in most cases after only one day of culture. For RSV, the sensitivity of the shell vial method was too low (74%) to allow omission of DIF (sensitivity 95%).(ABSTRACT TRUNCATED AT 250 WORDS)