Assuntos
Adenocarcinoma/genética , Aberrações Cromossômicas , Hibridização in Situ Fluorescente/métodos , Neoplasias Pancreáticas/genética , Adenocarcinoma/diagnóstico , Cromossomos Humanos Par 17/genética , Cromossomos Humanos Par 18/genética , Cromossomos Humanos Par 8/genética , Cromossomos Humanos Par 9/genética , Inibidor p16 de Quinase Dependente de Ciclina/genética , Humanos , Neoplasias Pancreáticas/diagnóstico , Proteínas Proto-Oncogênicas c-myc/genética , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Proteína Smad4/genética , Proteína Supressora de Tumor p53/genéticaRESUMO
A rapid procedure was developed for detection and identification of methicillin-resistant Staphylococcus aureus (MRSA) directly from sterile sites or mixed flora samples (e.g., nose or inguinal swabs). After a rapid conditioning of samples, the method consists of two main steps: (i) immunomagnetic enrichment in S. aureus and (ii) amplification-detection profile on DNA extracts using multiplex quantitative PCR (5'-exonuclease qPCR, TaqMan). The triplex qPCR assay measures simultaneously the following targets: (i) mecA gene, conferring methicillin resistance, common to both S. aureus and Staphylococcus epidermidis; (ii) femA gene from S. aureus; and (iii) femA gene from S. epidermidis. This quantitative approach allows discrimination of the origin of the measured mecA signal. qPCR data were calibrated using two reference strains (MRSA and methicillin-resistant S. epidermidis) processed in parallel to clinical samples. This 96-well format assay allowed analysis of 30 swab samples per run and detection of the presence of MRSA with exquisite sensitivity compared to optimal culture-based techniques. The complete protocol may provide results in less than 6 h (while standard procedure needs 2 to 3 days), thus allowing prompt and cost-effective implementation of contact precautions.
