Abstract
Meat adulteration has been a great concern in the food industry worldwide. Due to its lower cost, chicken is generally used as an adulterant in several meat products. The main objective of this research was to develop a novel detection platform based on colourimetric loop-mediated isothermal amplification (LAMP) for authenticating chicken content in raw and processed meat products. Conditions for the colourimetric LAMP were thus optimised. Neutral red, a pH-sensitive indicator, was introduced into the LAMP reactions to help distinguish the positive/negative outcomes. LAMP reaction containing amplified LAMP amplicons changed its colour from yellow to pink/magenta, while the reaction without amplified DNA products remained in its original yellow colour. This single-step colourimetric LAMP was also validated for its high specificity towards chicken DNA without any cross-reactivity with the DNA from other types of meat. The limits of detection (LOD) for chicken DNA and chicken in binary meat admixtures were 1 pg and 0.01% (w/w), respectively. From 33 different commercial processed food samples, the assay confirmed chicken content in 14 declared chicken-containing products and in 8 products without the declaration of chicken content, while the remaining 11 meat samples without the declaration of chicken content showed no detectable chicken DNA. Furthermore, without the requirement of DNA purification, the direct colourimetric LAMP assay is capable of accurate authentication of chicken content in raw meat matrices and commercial processed food samples, making it a valuable analytical tool to facilitate on-site food authentication, as well as in low-resource laboratory settings.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data Availability
All data generated or analysed during this study are included in this published article and its supplementary information file.
References
Ahmed MU, Hasan Q, Mosharraf Hossain M, Saito M, Tamiya E (2010) Meat species identification based on the loop mediated isothermal amplification and electrochemical DNA sensor. Food Control 21(5):599–605. https://doi.org/10.1016/j.foodcont.2009.09.001
Ali ME, Hashim U, Mustafa S, Che Man YB, Islam KN (2012) Gold nanoparticle sensor for the visual detection of pork adulteration in meatball formulation. J Nanomater 2012:103607. https://doi.org/10.1155/2012/103607
Alikord M, Momtaz H, Keramat J, Kadivar M, Rad AH (2018) Species identification and animal authentication in meat products: a review. J Food Meas Charact 12(1):145–155. https://doi.org/10.1007/s11694-017-9625-z
Ballin NZ, Vogensen FK, Karlsson AH (2009) Species determination – can we detect and quantify meat adulteration? Meat Sci 83(2):165–174. https://doi.org/10.1016/j.meatsci.2009.06.003
Barakat H, El-Garhy HAS, Moustafa MMA (2014) Detection of pork adulteration in processed meat by species-specific PCR-QIAxcel procedure based on D-loop and cytb genes. Appl Microbiol Biotechnol 98(23):9805–9816. https://doi.org/10.1007/s00253-014-6084-x
Cawthorn D-M, Steinman HA, Hoffman LC (2013) A high incidence of species substitution and mislabelling detected in meat products sold in South Africa. Food Control 32(2):440–449. https://doi.org/10.1016/j.foodcont.2013.01.008
Cho A-R, Dong H-J, Cho S (2014) Meat species identification using loop-mediated isothermal amplification assay targeting species-specific mitochondrial DNA. Korean J Food Sci Anim Resour 34(6):799–807. https://doi.org/10.5851/kosfa.2014.34.6.799
Ckumdee J, Kaewphinit T, Chansiri K, Santiwatanakul S (2016) Development of Au-nanoprobes combined with loop-mediated isothermal amplification for detection of isoniazid resistance in Mycobacterium tuberculosis. J Chem 2016:3474396. https://doi.org/10.1155/2016/3474396
Di Pinto A, Bottaro M, Bonerba E, Bozzo G, Ceci E, Marchetti P, Mottola A, Tantillo G (2015) Occurrence of mislabeling in meat products using DNA-based assay. J Food Sci Technol 52(4):2479–2484. https://doi.org/10.1007/s13197-014-1552-y
Duan Y, Zhang X, Ge C, Wang Y, Cao J, Jia X, Wang J, Zhou M (2014) Development and application of loop-mediated isothermal amplification for detection of the F167Y mutation of carbendazim-resistant isolates in Fusarium graminearum. Sci Rep 4(1):7094. https://doi.org/10.1038/srep07094
Farag M, Alagawany M, El-Hack M, Tiwari R, Dhama K (2015) Identification of different animal species in meat and meat products - trends and advances. Adv Anim Vet Sci 3:334–346. https://doi.org/10.14737/journal.aavs/2015/3.6.334.346
Fujimura S, Kawano S, Koga H, Takeda H, Kadowaki M, Ishibashi T (1995) Identification of taste-active components in the chicken meat extract by omission test-involvement of glutamic acid, IMP and potassium lon. Nihon Chikusan Gakkaiho 66:43–51
Fujimura T, Matsumoto T, Tanabe S, Morimatsu F (2008) Specific discrimination of chicken DNA from other poultry DNA in processed foods using the polymerase chain reaction. Biosci Biotechnol Biochem 72(3):909–913. https://doi.org/10.1271/bbb.70662
Goto M, Honda E, Ogura A, Nomoto A, Hanaki K-I (2009) Colorimetric detection of loop-mediated isothermal amplification reaction by using hydroxy naphthol blue. Biotechniques 46(3):167–172. https://doi.org/10.2144/000113072
Izadpanah M, Mohebali N, Elyasi Gorji Z, Farzaneh P, Vakhshiteh F, Shahzadeh Fazeli SA (2018) Simple and fast multiplex PCR method for detection of species origin in meat products. J Food Sci Technol 55(2):698–703. https://doi.org/10.1007/s13197-017-2980-2
Johnson R (2014) Food fraud and “Economically motivated adulteration” of food and food ingredients. Congr Res Serv:1–40. https://sgp.fas.org/crs/misc/R43358.pdf
Kane DE, Hellberg RS (2016) Identification of species in ground meat products sold on the U.S. commercial market using DNA-based methods. Food Control 59:158–163. https://doi.org/10.1016/j.foodcont.2015.05.020
Karabasanavar NS, Singh SP, Kumar D, Shebannavar SN (2013) Development and application of highly specific PCR for detection of chicken (Gallus gallus) meat adulteration. Eur Food Res Technol 236(1):129–134. https://doi.org/10.1007/s00217-012-1868-7
Karami A, Gill P, Motamedi M, Saghafinia M (2011) A review of the current isothermal amplification techniques: applications, advantages and disadvantages. J Glob Infect Dis 3(3):293–302. https://doi.org/10.4103/0974-777X.83538
Kesmen Z, Yetiman AE, Sahin F, Yetim H (2012) Detection of chicken and turkey meat in meat mixtures by using real-time PCR assays. J Food Sci 77(2):C167–C173. https://doi.org/10.1111/j.1750-3841.2011.02536.x
Keyvan E, Çil G, Çınar Kul B, Bilgen N, Şireli U (2017) Identification of meat species in different types of meat products by PCR. Ankara Üniversitesi Vet Fakültesi Derg 64:261–266. https://doi.org/10.1501/Vetfak_0000002809
Kim M-J, Kim H-Y (2019) A fast multiplex real-time PCR assay for simultaneous detection of pork, chicken, and beef in commercial processed meat products. LWT 114:108390. https://doi.org/10.1016/j.lwt.2019.108390
Kim M-J, Yoo I, Yang S-M, Suh S-M, Kim H-Y (2018) Development and validation of a multiplex PCR assay for simultaneous detection of chicken, turkey and duck in processed meat products. Int J Food Sci Technol 53(12):2673–2679. https://doi.org/10.1111/ijfs.13876
Kitpipit T, Sittichan K, Thanakiatkrai P (2014) Direct-multiplex PCR assay for meat species identification in food products. Food Chem 163:77–82. https://doi.org/10.1016/j.foodchem.2014.04.062
Koh B-R-D, Kim J-Y, Chang H-M, Park S-D, Kim Y-H (2011) Development of species-specific multiplex PCR assays of mitochondrial 12S rRNA and 16S rRNA for the identification of animal species. Korean J Vet Serv 34:417–428. https://doi.org/10.7853/kjvs.2011.34.4.417
Kumar A, Kumar RR, Sharma BD, Gokulakrishnan P, Mendiratta SK, Sharma D (2015) Identification of species origin of meat and meat products on the DNA basis: a review. Crit Rev Food Sci Nutr 55(10):1340–1351. https://doi.org/10.1080/10408398.2012.693978
Mane BG, Mendiratta SK, Tiwari AK (2009) Polymerase chain reaction assay for identification of chicken in meat and meat products. Food Chem 116(3):806–810. https://doi.org/10.1016/j.foodchem.2009.03.030
Martín I, García T, Fajardo V, López-Calleja I, Rojas M, Pavón MA, Hernández PE, González I, Martín R (2007) Technical note: detection of chicken, turkey, duck, and goose tissues in feedstuffs using species-specific polymerase chain reaction. J Anim Sci 85(2):452–458. https://doi.org/10.2527/jas.2006-350
Montowska M, Pospiech E (2010) Authenticity determination of meat and meat products on the protein and DNA basis. Food Rev Int 27(1):84–100. https://doi.org/10.1080/87559129.2010.518297
Mori Y, Nagamine K, Tomita N, Notomi T (2001) Detection of loop-mediated isothermal amplification reaction by turbidity derived from magnesium pyrophosphate formation. Biochem Biophys Res Commun 289(1):150–154. https://doi.org/10.1006/bbrc.2001.5921
Nie K, Zhao X, Ding X, Li XD, Zou SM, Guo JF, Wang DY, Gao RB, Li XY, Huang WJ, Shu YL, Ma XJ (2013) Visual detection of human infection with influenza A (H7N9) virus by subtype-specific reverse transcription loop-mediated isothermal amplification with hydroxynaphthol blue dye. Clin Microbiol Infect 19(8):e372–e375. https://doi.org/10.1111/1469-0691.12263
Niessen L, Bechtner J, Fodil S, Taniwaki MH, Vogel RF (2018) LAMP-based group specific detection of aflatoxin producers within Aspergillus section Flavi in food raw materials, spices, and dried fruit using neutral red for visible-light signal detection. Int J Food Microbiol 266:241–250. https://doi.org/10.1016/j.ijfoodmicro.2017.12.013
Notomi T, Okayama H, Masubuchi H, Yonekawa T, Watanabe K, Amino N, Hase T (2000) Loop-mediated isothermal amplification of DNA. Nucleic Acids Res 28(12):e63–e63. https://doi.org/10.1093/nar/28.12.e63
O’Mahony PJ (2013) Finding horse meat in beef products—a global problem. QJM an Int J Med 106(6):595–597. https://doi.org/10.1093/qjmed/hct087
Oliveira BB, Veigas B, Baptista PV (2021) Isothermal amplification of nucleic acids: the race for the next “gold standard.” Front Sensors 2:14. https://doi.org/10.3389/fsens.2021.752600
Omran GA, Tolba AO, El-Sharkawy EEE-D, Abdel-Aziz DM, Ahmed HY (2019) Species DNA-based identification for detection of processed meat adulteration: is there a role of human short tandem repeats (STRs)? Egypt J Forensic Sci 9(1):15. https://doi.org/10.1186/s41935-019-0121-y
Premanandh J (2013) Horse meat scandal – a wake-up call for regulatory authorities. Food Control 34(2):568–569. https://doi.org/10.1016/j.foodcont.2013.05.033
Rodríguez MA, García T, González I, Asensio L, Hernández PE, Martin R (2003) Qualitative PCR for the detection of chicken and pork adulteration in goose and mule duck foie gras. J Sci Food Agric 83(11):1176–1181. https://doi.org/10.1002/jsfa.1527
Schrader C, Schielke A, Ellerbroek L, Johne R (2012) PCR inhibitors – occurrence, properties and removal. J Appl Microbiol 113(5):1014–1026. https://doi.org/10.1111/j.1365-2672.2012.05384.x
Seetang-Nun Y, Jaroenram W, Sriurairatana S, Suebsing R, Kiatpathomchai W (2013) Visual detection of white spot syndrome virus using DNA-functionalized gold nanoparticles as probes combined with loop-mediated isothermal amplification. Mol Cell Probes 27(2):71–79. https://doi.org/10.1016/j.mcp.2012.11.005
Shears P (2010) Food fraud – a current issue but an old problem. Br Food J 112:198–213. https://doi.org/10.1108/00070701011018879
Sul S, Kim M-J, Kim H-Y (2019) Development of a direct loop-mediated isothermal amplification (LAMP) assay for rapid and simple on-site detection of chicken in processed meat products. Food Control 98:194–199. https://doi.org/10.1016/j.foodcont.2018.11.025
Tanabe S, Hase M, Yano T, Sato M, Fujimura T, Akiyama H (2007) A real-time quantitative PCR detection method for pork, chicken, beef, mutton, and horseflesh in foods. Biosci Biotechnol Biochem 71(12):3131–3135. https://doi.org/10.1271/bbb.70683
Tanner NA, Zhang Y, Evans TC (2015) Visual detection of isothermal nucleic acid amplification using pH-sensitive dyes. Biotechniques 58(2):59–68. https://doi.org/10.2144/000114253
Thangsunan P, Temisak S, Morris P, Rios-Solis L, Suree N (2021) Combination of loop-mediated isothermal amplification and AuNP-oligoprobe colourimetric assay for pork authentication in processed meat products. Food Anal Methods 14(3):568–580. https://doi.org/10.1007/s12161-020-01901-5
Tomita N, Mori Y, Kanda H, Notomi T (2008) Loop-mediated isothermal amplification (LAMP) of gene sequences and simple visual detection of products. Nat Protoc 3(5):877–882. https://doi.org/10.1038/nprot.2008.57
Unajak S, Meesawat P, Anyamaneeratch K, Duangtipanuwareepong SK, Choowongkomon K (2010) Identification of species (meat and blood samples) using nested-PCR analysis of mitochondrial DNA. African J Biotechnol 10(29):5670–5676. https://doi.org/10.5897/AJB10.2432
Wang F, Wu X, Xu D, Chen L, Ji L (2020) Identification of chicken-derived ingredients as adulterants using loop-mediated isothermal amplification. J Food Prot 83(7):1175–1180. https://doi.org/10.4315/JFP-19-542
Wang J, Wan Y, Chen G, Liang H, Ding S, Shang K, Li M, Dong J, Du F, Cui X, Tang Z (2019) Colorimetric detection of horse meat based on loop-mediated isothermal amplification (LAMP). Food Anal Methods 12(11):2535–2541. https://doi.org/10.1007/s12161-019-01590-9
Wood T (1956) Some applications of paper chromatography to the examination of meat extract. J Sci Food Agric 7(3):196–200. https://doi.org/10.1002/jsfa.2740070305
Xiong J, Huang B, Xu J, Huang W (2020) A closed-tube loop-mediated isothermal amplification assay for the visual detection of Staphylococcus aureus. Appl Biochem Biotechnol 191(1):201–211. https://doi.org/10.1007/s12010-020-03278-x
Yang B-Y, Liu X-L, Wei Y-M, Wang J-Q, He X-Q, Jin Y, Wang Z-J (2014) Rapid and sensitive detection of human astrovirus in water samples by loop-mediated isothermal amplification with hydroxynaphthol blue dye. BMC Microbiol 14(1):38. https://doi.org/10.1186/1471-2180-14-38
Acknowledgements
The authors would like to thank Prof. Paul A. Millner from School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, UK, and Prof. Richard L. Deming from California State University, Fullerton, USA, for helpful discussions and English language editing.
Funding
This research was supported by the Program Management Unit for Human Resources & Institutional Development, Research and Innovation, Office of National Higher Education Science Research and Innovation Policy Council, Thailand (NXPO) (Grant Number B16F640001). This work was also partially supported by the Center of Excellence in Materials Science and Technology, the Materials Science Research Center (MSRC), Faculty of Science, Chiang Mai University, Thailand.
Author information
Authors and Affiliations
Contributions
Pattanapong Thangsunan, conceptualisation, writing — original draft, writing — review and editing, methodology, and formal analysis. Sasithon Temisak, writing — review and editing, and methodology. Thanapak Jaimalai, methodology. Leonardo Rios-Solis, writing — review and editing. Nuttee Suree, funding acquisition and writing — review and editing.
Corresponding authors
Ethics declarations
Ethics Approval
This article does not contain any studies with human participants or animals performed by any of the authors.
Consent to Participate
Not applicable.
Conflict of Interest
Pattanapong Thangsunan declares that he has no conflict of interest. Sasithon Temisak declares that she has no conflict of interest. Thanapak Jaimalai declares that she has no conflict of interest. Leonardo Rios-Solis declares that he has no conflict of interest. Nuttee Suree declares that he has no conflict of interest.
Additional information
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Thangsunan, P., Temisak, S., Jaimalai, T. et al. Sensitive Detection of Chicken Meat in Commercial Processed Food Products Based on One-Step Colourimetric Loop-Mediated Isothermal Amplification. Food Anal. Methods 15, 1341–1355 (2022). https://doi.org/10.1007/s12161-021-02210-1
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12161-021-02210-1