title: Isothermal target and probe amplification assay for the real-time rapid detection of Staphylococcus aureus.
abstract: Staphylococcus aureus, the species most commonly associated with staphylococcal food poisoning, is one of the most prevalent causes of foodborne disease in Korea and other parts of the world, with much damage inflicted to the health of individuals and economic losses estimated at $120 million. To reduce food poisoning outbreaks by implementing prevention methods, rapid detection of S. aureus in foods is essential. Various types of detection methods for S. aureus are available. Although each method has advantages and disadvantages, high levels of sensitivity and specificity are key aspects of a robust detection method. Here, we describe a novel real-time isothermal target and probe amplification (iTPA) method that allows the rapid and simultaneous amplification of target DNA (the S. aureus nuc gene) and a fluorescence resonance energy transfer-based signal probe under isothermal conditions at 61 °C or detection of S. aureus in real time. The assay was able to specifically detect all 91 S. aureus strains tested without nonspecific detection of 51 non-S. aureus strains. The real-time iTPA assay detected S. aureus at an initial level of 10(1) CFU in overnight cultures of preenriched food samples (kiwi dressing, soybean milk, and custard cream). The advantage of this detection system is that it does not require a thermal cycler, reducing the cost of the real-time PCR and its footprint. Combined with a miniaturized fluorescence detector, this system can be developed into a simplified quantitative hand-held real-time device, which is often required. The iTPA assay was highly reliable and therefore may be used as a rapid and sensitive means of identifying S. aureus in foods.
authors.affiliation: Food Research and Development Center, Samsung Welstory, Inc., 2442-1 Yonggu-daero, Giheunggu, YongIn-si, Gyeonggi-do, South Korea.
authors.forename: Hyewon
authors.initials: H
authors.lastname: Shin
authors.affiliation: DxGene, Inc., 306 Ace Technotower 5th, 197-22 Guro-3-dong, Guro-gu, Seoul, South Korea.
authors.forename: Minhwan
authors.initials: M
authors.lastname: Kim
authors.affiliation: Department of Food Science and Nutrition, Dong-A University, 550 Nakdong-daero, Sahagu, Busan, South Korea.
authors.forename: Eunju
authors.initials: E
authors.lastname: Yoon
authors.affiliation: Food Research and Development Center, Samsung Welstory, Inc., 2442-1 Yonggu-daero, Giheunggu, YongIn-si, Gyeonggi-do, South Korea.
authors.forename: Gyoungwon
authors.initials: G
authors.lastname: Kang
authors.affiliation: DxGene, Inc., 306 Ace Technotower 5th, 197-22 Guro-3-dong, Guro-gu, Seoul, South Korea.
authors.forename: Seungyu
authors.initials: S
authors.lastname: Kim
authors.affiliation: Food Research and Development Center, Samsung Welstory, Inc., 2442-1 Yonggu-daero, Giheunggu, YongIn-si, Gyeonggi-do, South Korea.
authors.forename: Aelee
authors.initials: A
authors.lastname: Song
authors.affiliation: Food Research and Development Center, Samsung Welstory, Inc., 2442-1 Yonggu-daero, Giheunggu, YongIn-si, Gyeonggi-do, South Korea. jsoon.kim@samsung.com.
authors.forename: Jeongsoon
authors.initials: J
authors.lastname: Kim
mesh_terms: D015169:Colony Count, Microbial; D003470:Culture Media; D017931:DNA Primers; D004269:DNA, Bacterial / Q000302:isolation & purification*; D004926:Escherichia coli / Q000302:isolation & purification; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D008089:Listeria monocytogenes / Q000302:isolation & purification; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D056910:Republic of Korea; D012475:Salmonella / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D013202:Staphylococcal Food Poisoning; D013211:Staphylococcus aureus / Q000302:isolation & purification*; D015008:Yersinia enterocolitica / Q000302:isolation & purification
chemical_list: D003470:Culture Media; D017931:DNA Primers; D004269:DNA, Bacterial