Detection and Identification of Six Foodborne Bacteria by Two-tube Multiplex Real Time PCR and Melting Curve Analysis / 生物医学与环境科学(英文)
Biomedical and Environmental Sciences
;
(12): 770-778, 2014.
Artigo
em Inglês
| WPRIM
| ID: wpr-270541
ABSTRACT
<p><b>OBJECTIVE</b>This study is aimed to develop a two-tube melting curve-based multiplex real time PCR assay (MCMRT-PCR) for the simultaneous detection of six common foodborne pathogenic bacteria (diarrhoeagenic Escherichia coli, Salmonella, and Shigella in tube 1, Staphylococcus aureus, Vibrio parahaemolyticus, and Listeria monocytogenes in tube 2).</p><p><b>METHODS</b>A two-tube MCMRT-PCR assay was performed on 7900HT Fast Real-Time PCR System (Applied Biosystems, USA). Amplification by PCR was optimized to obtain high efficiency. The sensitivity and specificity of assays were investigated.</p><p><b>RESULTS</b>The detection limit of optimized MCMRT-PCR assay was 3.9×102 CFU/mL for S. aureus, 4.4×102 CFU/mL for L. monocytogenes, 3.0×102 CFU/mL for Salmonella, 2.5×102 CFU/mL for Shigella, 2.1×102 CFU/mL for V. parahaemolyticus, and 1.2×102 CFU/mL for E. coli. The feasibility of MCMRT-PCR was further evaluated using artificially contaminated milk, the sensitivity was at the level of 105 CFU/mL.</p><p><b>CONCLUSION</b>A two-tube MCMRT-PCR assay using six primer sets was developed for detection of multiple pathogens. Our findings demonstrates that the proposed two-tube assay is reliable, useful and rapid for simultaneous detection of six foodborne pathogenic bacteria with an intended application in provincial Centers for Diseases Control and Prevention (CDC).</p>
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Bactérias
/
Sensibilidade e Especificidade
/
Leite
/
Reação em Cadeia da Polimerase Multiplex
/
Reação em Cadeia da Polimerase em Tempo Real
/
Microbiologia de Alimentos
/
Genética
/
Métodos
/
Microbiologia
Tipo de estudo:
Estudo diagnóstico
/
Estudo prognóstico
Limite:
Animais
Idioma:
Inglês
Revista:
Biomedical and Environmental Sciences
Ano de publicação:
2014
Tipo de documento:
Artigo
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