RESUMO
The objective of this study was to develop a DNA sequencing assay that examines sensitively and reliably all conserved domains of the reverse transcriptase-encoding region of the HBV genome for antiviral resistance-associated mutations while simultaneously producing ample information for precise genotyping and determination of HBsAg mutation. This assay was used to examine 1000 de-identified HBV DNA positive samples with known viral loads from a broad-based, unselected patient population from across the United States. Of these, 946 were assayed successfully. Antiviral resistance-associated mutations were identified in 104 samples. The escape mutation sG145R in the surface antigen was identified in 0.8% of patient samples. Infections with genotypes A, B, C, D, E, F, G and H were observed in 36.6%, 19.6%, 21.7%, 13.5%, 3.6%, 0.7%, 2.2%, and 0.5% of patient samples respectively. Fifteen samples (1.6%) appeared to harbor infections with multiple genotypes as shown by the presence of double peaks throughout sequence electropherograms. The limit of detection of this assay was approximately 150IU/mL.
Assuntos
Farmacorresistência Viral/genética , Antígenos de Superfície da Hepatite B/genética , Vírus da Hepatite B/efeitos dos fármacos , Vírus da Hepatite B/genética , Mutação , Análise de Sequência de DNA/métodos , Antivirais/farmacologia , DNA Viral/química , Genótipo , Vírus da Hepatite B/classificação , Humanos , Filogenia , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Carga ViralAssuntos
Sequência de Bases , Sondas de DNA/genética , DNA Espaçador Ribossômico/genética , Variação Genética , Mycobacterium chelonae/classificação , Micobactérias não Tuberculosas/classificação , Mycobacterium chelonae/genética , Micobactérias não Tuberculosas/genética , Reação em Cadeia da Polimerase/métodos , RNA Ribossômico 16S/genética , RNA Ribossômico 23S/genética , Sensibilidade e EspecificidadeRESUMO
The DiversiLab System, which includes microfluidics-based detection, reagent kits, and software for data processing and analysis, is an automated method using repetitive sequence-based PCR (rep-PCR) for microbial strain typing. To assess the reliability of the DiversiLab System for strain characterization of Staphylococcus aureus, we tested clinical isolates sent to ARUP Laboratories for typing and compared results to those of pulsed field electrophoresis (PFGE) aided by the cluster analysis provided by BioNumerics software. spa typing was performed when the results of these two methods for an outbreak were not concordant. The study included 89 S. aureus isolates (65 mecA positive, 24 mecA negative) from 19 outbreaks (2 to 11 isolates/outbreak). The DiversiLab and PFGE-BioNumerics results were concordant for 15 of the 19 outbreaks. For the remaining four outbreaks, there was partial concordance between the two methods. spa typing results were the same as or more similar to rep-PCR results for three of those outbreaks and were more similar to PFGE results for one. With regard to performance, the DiversiLab system was considerably less labor intensive than PFGE and provided results in less than 24 h, compared with 2 to 3 days for PFGE. Additionally, the Web-based DiversiLab software provides standardized comparisons among isolates almost instantaneously and generates user-friendly, customized reports.
