Abstract
Pulsed-Field Gel Electrophoresis (PFGE) is a molecular subtyping method with high discriminatory power, reproducibility, and epidemiological concordance for the subtyping of Listeria monocytogenes and other bacteria. PFGE uses rare-cutting restriction enzymes (macrorestriction) that cut the genomic DNA, usually resulting in 6–25 DNA fragments ranging between 30 and 600 kb. Bacterial cells are immobilized in agarose plugs and subsequently lysed to release genomic DNA, which is then subjected to DNA digestion. AscI and ApaI restriction enzymes are typically used for L. monocytogenes. Electrophoresis using an alternating electric field direction results in a DNA banding pattern, or fingerprint, which is used to classify isolates into different pulsotypes. PFGE is currently the CDC’s gold standard method for epidemiological studies in foodborne outbreaks.
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References
Wiedmann M (2002) Molecular subtyping methods for Listeria monocytogenes. J AOAC Int 85:524–531
Graves LM, Hunter SB, Ong AR, Schoonmaker-Bopp D, Hise K, Kornstein L, Dewitt WE, Hayes PS, Dunne E, Mead P, Swaminathan B (2005) Microbiological aspects of the investigation that traced the 1998 outbreak of listeriosis in the United States to contaminated hot dogs and establishment of molecular subtyping-based surveillance for Listeria monocytogenes in the PulseNet network. J Clin Microbiol 43:2350–2355
Swaminathan B, Barrett TJ, Hunter SB, Tauxe RV, Force CPT (2001) PulseNet: the molecular subtyping network for foodborne bacterial disease surveillance, United States. Emerg Infect Dis 7:382–389
Scallan E, Hoekstra RM, Angulo FJ, Tauxe RV, Widdowson M-A, Roy SL, Jones JL, Griffin PM (2011) Foodborne illness acquired in the United States–major pathogens. Emerg Infect Dis 17:7–15
Murphy M, Corcoran D, Buckley JF, O’Mahony M, Whyte P, Fanning S (2007) Development and application of multiple-locus variable number of tandem repeat analysis (MLVA) to subtype a collection of Listeria monocytogenes. Int J Food Microbiol 115:187–194
PulseNet USA (2009) One-day (24–48 h) standardized laboratory protocol for molecular sub-typing of Listeria monocytogenes by Pulsed-Field Gel Electrophoresis (PFGE). http://www.pulsenetinternational.org/assets/PulseNet/uploads/pfge/PNL04_ListeriaPFGEProtocol.pdf
Fox EM, DeLappe N, Garvey P, Cormican M, Leonard N, Jordan K (2012) PFGE analysis of Listeria monocytogenes isolates of clinical, animal, food and environmental origin from Ireland. J Med Microbiol 61:540–547
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Luque-Sastre, L., Fanning, S., Fox, E.M. (2015). Pulsed-Field Gel Electrophoresis for Listeria monocytogenes . In: Jordan, K., Dalmasso, M. (eds) Pulse Field Gel Electrophoresis. Methods in Molecular Biology, vol 1301. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2599-5_5
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DOI: https://doi.org/10.1007/978-1-4939-2599-5_5
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2598-8
Online ISBN: 978-1-4939-2599-5
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