• Kankanit Pisamayarom Laboratory of Plant Transgenic Technology and Biosensor, Department of Botany, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand, Programs in Biotechnology, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand
  • Piyasak Chaumpluk Laboratory of Plant Transgenic Technology and Biosensor, Department of Botany, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand




Listeria Monocytogenes, Helicase Dependent Amplification (HDA), Blue Silver Nanoplates (Agnpls);, Frozen Seafood

Abstract [English]

Listeria monocytogenes, a foodborne pathogen, is considered as one of the major problems in food safety. With strong safety regulations, a monitoring measure is essential for protecting the health and safety of consumers. Thus, a reliable monitoring method is required. In this study, a rapid assay based on a combination of helicase dependent amplification (HDA) and DNA signal detection via nucleic acid hybridization in blue silver nanoplates (AgNPls) was established. The assay started directly after short term enrichment in terrific broth using cotton ball swapping technique on seafood surface. A HDA amplification of hly gene of L. monocytogenes at 65 °C allowed DNA signals to be increased, whereas the rendered DNA products were detected via nucleic acid hybridization with an oligonucleotide probe in AgNPls solution. The positive specimens induced blue silver nanoplates’ aggregation resulting in pale gray change to colorless, while the negative specimens showed the blue color of non-aggregated nanoplates. The method had a detection limit at 100 copies of L. monocytogenes DNA per 50 g of sample. This method was rapid, simple, did not require laboratory facilities and was suitable for field food safety monitoring


Download data is not yet available.


Acha, PN, Szyfres, B. Listeriosis. Zoonoses and Communicable Diseases Common to Man and Animals. (3rded.) Washington D.C.: Pan American Health Organization; 2003, 168-179.

Altschul, SF, Gish, W, Miller, W, Myers, EW, Lipman, DJ. Basic local alignment search tool, Journal of Molecular Biology, 215, 1990, 403-410. DOI: https://doi.org/10.1016/S0022-2836(05)80360-2

Bessesen, MT, Luo, QA, Rotbart, HA, Blaser, MJ, Ellison, RT. Detection of Listeria monocytogenes by using the polymerase chain reaction, Applied and Environmental Microbiology, 56(9), 1990, 2930-2932. DOI: https://doi.org/10.1128/AEM.56.9.2930-2932.1990

Bickley, J, Short, JK, McDowell, DG, Parkes, HC. Polymerase chain reaction (PCR) detection of Listeria monocytogenes in diluted milk and reversal of PCR inhibition caused by calcium ions, Letters in Applied Microbiology, 22, 1996, 153. DOI: https://doi.org/10.1111/j.1472-765X.1996.tb01131.x

Bloomfield, VA, Crothers, DM, Tinoco, I, Jr. Nuclei Acids: Structures, Properties, and Functions: Sausalito, CA: University Science Books; 2000.

Border, PM, Howard, JJ, Plastow, GS, Siggens, KW. Detection of Listeria species and Listeria monocytogenes using polymerase chain reaction, Letters in Applied Microbiology, 11, 1990, 158-162. DOI: https://doi.org/10.1111/j.1472-765X.1990.tb00149.x

Bortolussi, R. Listeriosis: a primer, Canadian Medical Association Journal, 179, 2008, 795-797. DOI: https://doi.org/10.1503/cmaj.081377

Chaumpluk, P, Chaiprasart, P. Fluorescence biosensor based on N-(2-Aminoethyl) glycine peptide nucleic acid for a simple and rapid detection of Escherichia coli in fresh-cut mango, Acta Horticulturae, 992, 2013, 551-560. DOI: https://doi.org/10.17660/ActaHortic.2013.992.68

Churchill, RLT, Lee, H, Hall, JC. Detection of Listeria monocytogenes and the toxin listeriolysin O in food, Journal of Microbiological Methods, 64, 2006, 141-170. DOI: https://doi.org/10.1016/j.mimet.2005.10.007

Cossart, P, Mengaud, J. Listeria monocytogenes: A model system for the molecular study of intracellular parasitism, Molecular Biology & Medicine, 6, 1989, 463-474.

de Valk, H, Vaillant, V, Jacquet, C, Rocourt, J, Le Querrec, F, Stainer, F, Quelquejeu, N, Pierre, O, Pierre, V, Desenclos, JC, Goulet, V. Two consecutive nationwide outbreaks of Listeriosis in France, October 1999-February 2000, American Journal of Epidemiology, 154(10), 2001, 944-950. DOI: https://doi.org/10.1093/aje/154.10.944

Donelly, CW. In: Ryser ET, Marth EH (Eds.). Listeria, Listeriosis, and Food safety, 2nd ed.; Marcel Dekker Inc., New York, N.Y.; 1991, 225-260.

Fawcett, T. An introduction to ROC analysis. Pattern Recogn Lett. 27, 2006, 861-874. DOI: https://doi.org/10.1016/j.patrec.2005.10.010

Fu, Z, Zhou, X, Xing, D. Rapid colorimetric gene-sensing of food pathogenic bacteria using biomodification-free gold nanoparticle, Sensors and Actuators B: Chemical, 182, 2013, 633. DOI: https://doi.org/10.1016/j.snb.2013.03.033

Furrer, B., Candrian, U, Hoefelein, C, Luethy, J. Detection and identification of Listeria monocytogenes in cooked sausage products and in milk by in vitro amplification of haemolysin gene fragments, Journal of Applied Microbiology, 70(5), 1991, 372-379. DOI: https://doi.org/10.1111/j.1365-2672.1991.tb02951.x

Giljohann, DA, Seferos, DS, Daniel, WL, Massich, MD, Patel, PC, Mirkin, CA. Gold nanoparticles for biology and medicine, Angewandte Chemie International Edition in English, 49(19), 2010, 3280-94. DOI: https://doi.org/10.1002/anie.200904359

Gravani, R. Listeria in food-processing facilities. In Ryser ET and Marth EH (ed.), Listeria, listeriosis, and food safety; Marcel Dekker, New York, N.Y.; 1999, 657-709.

Han, MS, Lytton-Jean, AK, Oh, BK, Heo, J, Mirkin, CA. Colorimetric screening of DNA binding molecules with gold nanoparticle probes; Angewandte Chemie International Edition in English, 45(11), 2006, 1807-1810. DOI: https://doi.org/10.1002/anie.200504277

Hunter, RJ. Foundations of Colloid Science; Oxford University Press, New York; 2001.

Huss, HH, Jorgensen, LV, Vogel, BF. Control options for Listeria monocytogenes in sea foods, International Journal of Food Microbiology, 62(3), 2000, 267-274. DOI: https://doi.org/10.1016/S0168-1605(00)00347-0

Jallewar, PK, Kalorey, DR, Kurkure, NV, Pandeb, VV, Barbuddhe, SB. Genotypic characterization of Listeria spp. isolated from fresh water fish, International Journal of Food Microbiology, 114, 2007, 120-123. DOI: https://doi.org/10.1016/j.ijfoodmicro.2006.09.034

Jørgensen, LV, Huss, HH. Prevalence and growth of Listeria monocytogenes in naturally contaminated seafood, International Journal of Food Microbiology, 42, 1998, 127. DOI: https://doi.org/10.1016/S0168-1605(98)00071-3

Kalorey, DK, Warke, SR, Kurkure, NV, Rawool, DB, Barbuddhe, SB. Listeria species in bovine raw milk: A large survey of Central India, Food Control, 19, 2008, 109-112. DOI: https://doi.org/10.1016/j.foodcont.2007.02.006

Kathariou, S. Foodborn outbreaks of listeriosis and epidemic-associated lineages of Listeria monocytogenes. In ME Torrence and RE Isaacson (ed.), Microbial food Safety in Animal Agriculture: Iowa State University Press, Ames, I; 2003, 243-256. DOI: https://doi.org/10.1002/9780470752616.ch25

Kreibig, U, Genzel, L. Optical absorption of small metallic particles, Surface Science, 1985, 156, A330-A331. DOI: https://doi.org/10.1016/0167-2584(85)90457-8

Kuribara, H, Shindo, Y, Matsuoka, T, Takubo, K, Futo, S, Aoki, N, Hirao, T, Akiyama, H, Goda, Y, Toyoda, M, Hino, A. Novel reference molecules for quantitation of genetically modified maize and soybean, Journal of AOAC International, 85, 2002, 1077-1089. DOI: https://doi.org/10.1093/jaoac/85.5.1077

Li, H, Rothberg, L. Colorimetric detection of DNA sequences based on electrostatic interactions with unmodified gold nanoparticles, Proceedings of the National Academy of Sciences, 101, 2004, 14036-14039. DOI: https://doi.org/10.1073/pnas.0406115101

Low, JC, Donachie, W. A review of Listeria monocytogenes and listeriosis, Veterinary Journal, 153(1), 1997, 9-29. DOI: https://doi.org/10.1016/S1090-0233(97)80005-6

Makino, SI, Kawamoto, K, Takeshi, K, Okada, Y, Yamasaki, M, Yamamoto, S, Igimi, S. An outbreak of food-borne listeriosis due to cheese in Japan, during 2001, International Journal of Food Microbiology, 104, 2005, 189-196. DOI: https://doi.org/10.1016/j.ijfoodmicro.2005.02.009

Nørrung, B, Andersen, JK, Schlundt, J. Incidence and control of Listeria monocytogenes in foods in Denmark, International Journal of Food Microbiology, 53(2-3), 1999, 195-203. DOI: https://doi.org/10.1016/S0168-1605(99)00157-9

Parnklang, T, Lertvachirapaiboon, C, Pienpinijtham, P, Wongravee, K, Thamma-charoen, C, Ekgasit, S. H2O2-triggered shape transformation of silver nanospheres to nanoprisms with controllable longitudinal LSPR wavelengths, Royal Society of Chemistry Advances, 3, 2013, 12886-12894. DOI: https://doi.org/10.1039/c3ra41486h

Portnoy, DA, Jacks, PS, Hinrichs, DJ. Role of hemolysin for the intracellular growth of Listeria monocytogenes, Journal of Experimental Medicine, 67, 1988, 1459–1471. DOI: https://doi.org/10.1084/jem.167.4.1459

Sambrook, J, Fritsch, EF, Maniatis, T. Molecular cloning: a laboratory manual: Cold Spring Harbor Laboratory Press; 1989.

Tham, W, Ericsson, H, Loncarevic, S, Unnerstad, H, Danielsson-Tham, ML. Lessons from an outbreak of listeriosis related to vacuum-packed gravad and cold-smoked fish, International Journal of Food Microbiology, 62, 2000, 173-175. DOI: https://doi.org/10.1016/S0168-1605(00)00332-9

Thomas, EJG, King, RK, Burchak, J, Gannon, VPJ. Sensitive and specific detection of Listeria monocytogenes in milk and ground beef with the polymerase chain reaction, Applied and Environmental Microbiology, 57, 1991, 2576-2580. DOI: https://doi.org/10.1128/AEM.57.9.2576-2580.1991

Thompson, DG, Enright, A, Faulds, K, Smith, WE, Graham, D. Ultrasensitive DNA detection using oligonucleotide-silver nanoparticle conjugates, Analytical Chemistry, 80, 2008, 2805-2810. DOI: https://doi.org/10.1021/ac702403w

Vincent, M, Xu, Y, Kong, H. Helicase-dependent isothermal DNA amplification, EMBO Reports, 5(8), 2004, 759-800. DOI: https://doi.org/10.1038/sj.embor.7400200

Yano, A, Ishimaru, R, Hujikata, R. Rapid and sensitive detection of heat-labile I and heat-stable I enterotoxin genes of enterotoxigenic Escherichia coli by Loop-Mediated Isothermal Amplification, Journal of Microbiological Methods, 68(22), 2007, 414-420. DOI: https://doi.org/10.1016/j.mimet.2006.09.024

Zhou, J, Ralston, J, Sedev, R, Beattie, DA. Functionalized gold nanoparticles: synthesis, structure and colloid stability, Journal of Colloid and Interface Science, 331, 2009, 251-262. DOI: https://doi.org/10.1016/j.jcis.2008.12.002




How to Cite

Pisamayarom, K., & Chaumpluk, P. (2017). RAPID LISTERIA MONOCYTOGENES ASSAY BASED ON HELICASE DEPENDENT AMPLIFICATION (HDA) AND NUCLEIC ACID HYBRIDIZATION IN BLUE SILVER NANOPLATES. International Journal of Research -GRANTHAALAYAH, 5(10), 322–335. https://doi.org/10.29121/granthaalayah.v5.i10.2017.2308