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Loop-mediated isothermal amplification for visual detection of Vibrio parahaemolyticus using gold nanoparticles

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A Correction to this article was published on 28 December 2017

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Abstract

Loop-mediated isothermal amplification (LAMP) eradicates the need of thermocycler in DNA amplification. Signals are usually obtained via fluorometry or turbidimetry, but such methods need improvement in order to become more effortless and reliable. The authors describe a set of six specific primers targeting the species-specific tlh gene of Vibrio parahaemolyticus which were used in accelerated LAMP reaction. Gold nanoparticles (AuNPs) were functionalized with streptavidin (Avidin-AuNPs), and engineered to signal the LAMP reaction. Two of the loop primers for LAMP were biotinylated and then can produce a DNA that can cause clusterization of Avidin-AuNPs based on the formation of avidin-biotin complex. This leads to a color change of the solution from red to blue. Amplification is completed within 30 min and can be visually detected within 5 min. The detection limit of the method is found to be 8.6 cfu per reaction. This visual detection scheme does not require any fluorescent reagents and detection instruments. Conceivably, the method has a wide scope because such Avidin-AuNPs can be used as nanoprobes for a variety of other LAMP products. This rapid and universal strategy holds promise in point of care testing and food testing, particularly in resource-limited regions.

Six specific primers (two of them are biotinylated) were used to realize the accelerated Loop-Mediated Isothermal Amplification. Streptavidin modified gold nanoparticles (Avidin-AuNPs) cluster on the DNA products, leading to the apparent change of color from red to blue, which is readily identified even by unaided eye.

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Change history

  • 28 December 2017

    The published version of this article, unfortunately, contained error. Modifications have been made to the Abstract, Introduction, Results and discussion, and Acknowledgements section. The original article has been corrected.

References

  1. Jones J, Lydon K, Kinsey T, Friedman B, Curtis M, Schuster R, Bowers J (2017) Effects of ambient exposure, refrigeration, and icing on Vibrio vulnificus and Vibrio parahaemolyticus abundances in oysters. Int J Food Microbiol 253:54–58

    Article  CAS  Google Scholar 

  2. Xiang G, Pu X, Jiang D, Liu L, Liu C, Liu X (2013) Development of a real-time resistance measurement for vibrio parahaemolyticus detection by the lecithin-dependent hemolysin gene. PLoS One 8(8):e72342

    Article  CAS  Google Scholar 

  3. Martinez-Urtaza J, Lozano-Leon A, Viña-Feas A, de Novoa J, Garcia-Martin O (2006) Differences in the API 20E biochemical patterns of clinical and environmental Vibrio parahaemolyticus isolates. FEMS Microbiol Lett 255(1):75–81

    Article  CAS  Google Scholar 

  4. Zeng J, Wei H, Zhang L, Liu X, Zhang H, Cheng J, Ma D, Zhang X, Fu P, Liu L (2014) Rapid detection of Vibrio parahaemolyticus in raw oysters using immunomagnetic separation combined with loop-mediated isothermal amplification. Int J Food Microbiol 174:123–128

    Article  CAS  Google Scholar 

  5. Yi M, Ling L, Neogi SB, Fan Y, Tang D, Yamasaki S, Shi L, Ye L (2014) Real time loop-mediated isothermal amplification using a portable fluorescence scanner for rapid and simple detection of Vibrio parahaemolyticus. Food Control 41:91–95

    Article  CAS  Google Scholar 

  6. Duan N, Shen M, Wu S, Zhao C, Ma X, Wang Z (2017) Graphene oxide wrapped Fe3O4@ Au nanostructures as substrates for aptamer-based detection of Vibrio parahaemolyticus by surface-enhanced Raman spectroscopy. Microchim Acta 184(8):2653–2660

    Article  CAS  Google Scholar 

  7. Teng J, Ye Y, Yao L, Yan C, Cheng K, Xue F, Pan D, Li B, Chen W (2017) Rolling circle amplification based amperometric aptamer/immuno hybrid biosensor for ultrasensitive detection of Vibrio parahaemolyticus. Microchim Acta 184(9):3477–3485

    Article  CAS  Google Scholar 

  8. 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

    Article  CAS  Google Scholar 

  9. Mori Y, Notomi T (2009) Loop-mediated isothermal amplification (LAMP): a rapid, accurate, and cost-effective diagnostic method for infectious diseases. J Infect Chemother 15(2):62–69

    Article  CAS  Google Scholar 

  10. Kaneko H, Kawana T, Fukushima E, Suzutani T (2007) Tolerance of loop-mediated isothermal amplification to a culture medium and biological substances. J Biochem Biophys Methods 70(3):499–501

    Article  CAS  Google Scholar 

  11. 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

    Article  CAS  Google Scholar 

  12. Notomi T, Mori Y, Tomita N, Kanda H (2015) Loop-mediated isothermal amplification (LAMP): principle, features, and future prospects. J Microbiol 53(1):1–5

    Article  CAS  Google Scholar 

  13. Ayukawa Y, Hanyuda S, Fujita N, Komatsu K, Arie T (2017) Novel loop-mediated isothermal amplification (LAMP) assay with a universal QProbe can detect SNPs determining races in plant pathogenic fungi. Sci Rep 7(1):4253. https://doi.org/10.1038/s41598-017-04084-y

    Article  Google Scholar 

  14. Boehme CC, Nabeta P, Henostroza G, Raqib R, Rahim Z, Gerhardt M, Sanga E, Hoelscher M, Notomi T, Hase T (2007) Operational feasibility of using loop-mediated isothermal amplification for diagnosis of pulmonary tuberculosis in microscopy centers of developing countries. J Clin Microbiol 45(6):1936–1940

    Article  CAS  Google Scholar 

  15. Goto M, Honda E, Ogura A, Nomoto A, Hanaki KI (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

    Article  CAS  Google Scholar 

  16. Rafati A, Gill P (2015) Microfluidic method for rapid turbidimetric detection of the DNA of Mycobacterium tuberculosis using loop-mediated isothermal amplification in capillary tubes. Microchim Acta 182(3–4):523–530

    Article  CAS  Google Scholar 

  17. Tian B, Ma J, Zardán Gómez de la Torre T, Bálint A, Donolato M, Hansen MF, Svedlindh P, Strömberg M (2016) Rapid Newcastle disease virus detection based on loop-mediated isothermal amplification and Optomagnetic readout. Acs Sensors 1(10):1228–1234

    Article  CAS  Google Scholar 

  18. Martin A, Grant KB, Stressmann F, Ghigo J-M, Marchal D, Limoges B (2016) Ultimate single-copy DNA detection using real-time electrochemical LAMP. Acs Sensors 1(7):904–912

    Article  CAS  Google Scholar 

  19. Nagamine K, Hase T, Notomi T (2002) Accelerated reaction by loop-mediated isothermal amplification using loop primers. Mol Cell Probes 16(3):223–229

    Article  CAS  Google Scholar 

  20. Zhang H, Wang S, Chen Z, Ge P, Jia R, Xiao E, Zeng W (2017) A turn-on fluorescent nanoprobe for lead(II) based on the aggregation of weakly associated gold(I)-glutathione nanoparticles. Microchim Acta 184(10):1–7

    Google Scholar 

  21. Li M, Shi L, Xie T, Jing C, Xiu G, Long Y-T (2017) An ultrasensitive Plasmonic Nanosensor for aldehydes. ACS Sensors 2(2):263–267

    Article  CAS  Google Scholar 

  22. Yu R-J, Sun J-J, Song H, Tian J-Z, Li D-W, Long Y-T (2017) Real-time sensing of O-Phenylenediamine oxidation on gold nanoparticles. Sensors 17(3):530

    Article  Google Scholar 

  23. Han X, Liu Y, Yin Y (2014) Colorimetric stress memory sensor based on disassembly of gold nanoparticle chains. Nano Lett 14(5):2466–2470. https://doi.org/10.1021/nl500144k

    Article  CAS  Google Scholar 

  24. Shi H-y, Yang L, Zhou X-y, Bai J, Gao J, Jia H-x, Li Q-g (2017) A gold nanoparticle-based colorimetric strategy coupled to duplex-specific nuclease signal amplification for the determination of microRNA. Microchim Acta 184(2):525–531

    Article  CAS  Google Scholar 

  25. Nordstrom JL, Vickery MCL, Blackstone GM, Murray SL, DePaola A (2007) Development of a multiplex real-time PCR assay with an internal amplification control for the detection of Total and pathogenic Vibrio parahaemolyticus bacteria in oysters. Appl Environ Microbiol 73(18):5840–5847. https://doi.org/10.1128/aem.00460-07

    Article  CAS  Google Scholar 

  26. Haiss W, Thanh NT, Aveyard J, Fernig DG (2007) Determination of size and concentration of gold nanoparticles from UV− Vis spectra. Anal Chem 79(11):4215–4221

    Article  CAS  Google Scholar 

  27. Jazayeri MH, Amani H, Pourfatollah AA, Pazoki-Toroudi H, Sedighimoghaddam B (2016) Various methods of gold nanoparticles (GNPs) conjugation to antibodies. Sens Bio-Sens Res 9:17–22. https://doi.org/10.1016/j.sbsr.2016.04.002

    Article  Google Scholar 

  28. National food safety standard food microbiological examination: Vibrio parahaemolyticus (2013) China standard protocols, GB4789.7-2013. National health and family planning commission of the PRC, the People’s Republic of China

  29. Zhong Q, Tian J, Wang B, Wang L (2016) PMA based real-time fluorescent LAMP for detection of Vibrio parahaemolyticus in viable but nonculturable state. Food Control 63:230–238

    Article  CAS  Google Scholar 

  30. Wang R, Xiao X, Chen Y, Wu J, Qian W, Wang L, Liu Y, Ji F, Wu J (2017) A loop-mediated, isothermal amplification-based method for visual detection of Vibrio parahaemolyticus within only 1 h, from shrimp sampling to results. Anal Methods 9(11):1695–1701

    Article  CAS  Google Scholar 

  31. Yamazaki W, Ishibashi M, Kawahara R, Inoue K (2008) Development of a loop-mediated isothermal amplification assay for sensitive and rapid detection of Vibrio parahaemolyticus. BMC Microbiol C7–163 8(1):1–7. https://doi.org/10.1186/1471-2180-8-163

    Article  Google Scholar 

  32. Chen S, Wang F, Beaulieu JC, Stein RE, Ge B (2011) Rapid detection of viable salmonellae in produce by coupling propidium monoazide with loop-mediated isothermal amplification. Appl Environ Microbiol 77(12):4008–4016

    Article  CAS  Google Scholar 

  33. Ahmad F, Stedtfeld RD, Waseem H, Williams MR, Cupples AM, Tiedje JM, Hashsham SA (2017) Most probable number-loop mediated isothermal amplification (MPN-LAMP) for quantifying waterborne pathogens in< 25min. J Microbiol Methods 132:27–33

    Article  CAS  Google Scholar 

  34. Wang L, Shi L, Su J, Ye Y, Zhong Q (2013) Detection of Vibrio parahaemolyticus in food samples using in situ loop-mediated isothermal amplification method. Gene 515(2):421–425. https://doi.org/10.1016/j.gene.2012.12.039

    Article  CAS  Google Scholar 

  35. Gao H, Lei Z, Jia J, Wang S, Chen Y, Sun M, Liang C (2009) Application of loop-mediated isothermal amplification for detection of Yersinia enterocolitica in pork meat. J Microbiol Methods 77(2):198–201

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors acknowledge the financial support provided by the Yangfan project (14YF1408100) from Science and Technology Commission of Shanghai Municipality – PR China and the special research fund for the national non-profit institutes (East China Sea Fisheries Research Institute) (No. 2014 T05). E.K.F. thanks the World Academy of Sciences (TWAS) under the Grant No. 16-510 RG/CHE/AF/AC_G–FR3240293301 for its financial support.

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Authors and Affiliations

  1. Key Laboratory of East China Sea Fishery Resources Exploitation, Ministry of Agriculture, East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai, 200090, China

    Cong Kong, Yuan Wang, Guang-xin Yang & Feng Han

  2. Laboratory of Physical Chemistry, Université Felix Houphouet-Boigny, 22 BP 582, Abidjan 22, Côte d’Ivoire

    Essy Kouadio Fodjo

  3. Key Laboratory of Oceanic and Polar Fisheries, Ministry of Agriculture, East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai, 200090, China

    Xiao-sheng Shen

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  1. Cong Kong
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  2. Yuan Wang
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  3. Essy Kouadio Fodjo
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  4. Guang-xin Yang
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  5. Feng Han
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Correspondence to Cong Kong or Xiao-sheng Shen.

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Additional information

The original version of this article was revised: The published version of this article, unfortunately, contained error. Modifications have been made to the Abstract, Introduction, Results and discussion, and Acknowledgments section.

A correction to this article is available online at https://doi.org/10.1007/s00604-017-2617-1.

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Kong, C., Wang, Y., Fodjo, E.K. et al. Loop-mediated isothermal amplification for visual detection of Vibrio parahaemolyticus using gold nanoparticles. Microchim Acta 185, 35 (2018). https://doi.org/10.1007/s00604-017-2594-4

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