Microchimica Acta

, 185:212 | Cite as

Detection of nucleic acids and elimination of carryover contamination by using loop-mediated isothermal amplification and antarctic thermal sensitive uracil-DNA-glycosylase in a lateral flow biosensor: application to the detection of Streptococcus pneumoniae

  • Yi Wang
  • Yan Wang
  • Dongxun Li
  • Jianguo Xu
  • Changyun Ye
Original Paper

Abstract

The authors report on a loop-mediated isothermal amplification (LAMP) scheme that uses antarctic thermally sensitive uracil-DNA-glycosylase (AUDG) for simultaneous detection of nucleic acids and elimination of carryover contamination. It was applied in a lateral flow assay (LFA) format. The assay has attractive features in that it does not require the use of labeled primers or probes, and can eliminate false-positive results generated by unwanted hybridization between two labeled primers or between a labeled primer and probe. LAMP amplification and AUDG digestion are conducted in a single pot, and the application of a closed-tube reaction prevents false-positives due to carryover contamination. The method was applied to the detection of the human pathogen Streptococcus pneumoniaein in pure cultures and spiked blood samples. This LFA can detect S. pneumoniae in pure cultures with a 25 fg.μL−1 detection limit and in spiked blood samples with a 470 cfu.mL−1 detection limit. Conceivably, this assay can be applied to the detection of various other targets if the specific LAMP primers are available.

Graphical abstract

Keywords

Diagnostic technique Nucleic acid amplification Limit of detection Nanoparticles False-positive result 

Notes

Acknowledgements

We acknowledge the financial supports of the grants (Mega Project of Research on the Prevention and Control of HIV/AIDS, Viral Hepatitis Infectious Diseases 2013ZX10004-101 to Changyun Ye) from the Ministry of Science and Technology, People’s Republic of China, and grant (2015SKLID507 to Changyun Ye) from State Key Laboratory of Infectious Disease Prevention and Control, China CDC.

Compliance with ethical standards

The author(s) declare that they have no competing interests.

Supplementary material

604_2018_2723_MOESM1_ESM.doc (5.1 mb)
ESM 1 (DOC 5184 kb)

References

  1. 1.
    Wang Y, Wang Y, Ma A-J, Li D-X, Luo L-J, Liu D-X, Jin D, Liu K, Ye C-Y (2015) Rapid and sensitive isothermal detection of nucleic-acid sequence by multiple cross displacement amplification. Sci Rep 5:11902CrossRefGoogle Scholar
  2. 2.
    Yan L, Zhou J, Zheng Y, Gamson AS, Roembke BT, Nakayama S, Sintim HO (2014) Isothermal amplified detection of DNA and RNA. Mol BioSyst 10(5):970–1003.  https://doi.org/10.1039/c3mb70304e CrossRefGoogle Scholar
  3. 3.
    Wang X, Liu W, Yin B, Sang Y, Liu Z, Dai Y, Duan X, Zhang G, Ding S, Tao Z (2017) An isothermal strand displacement amplification strategy for nucleic acids using junction forming probes and colorimetric detection. Microchim Acta 184(6):1603–1610.  https://doi.org/10.1007/s00604-017-2158-7 CrossRefGoogle Scholar
  4. 4.
    Zhao Y, Chen F, Li Q, Wang L, Fan C (2015) Isothermal Amplification of Nucleic Acids. Chem Rev 115(22):12491–12545.  https://doi.org/10.1021/acs.chemrev.5b00428 CrossRefGoogle Scholar
  5. 5.
    Zhou L, Wang J, Chen Z, Li J, Wang T, Zhang Z, Xie G (2017) A universal electrochemical biosensor for the highly sensitive determination of microRNAs based on isothermal target recycling amplification and a DNA signal transducer triggered reaction. Microchim Acta 184(5):1305–1313.  https://doi.org/10.1007/s00604-017-2129-z CrossRefGoogle Scholar
  6. 6.
    Xu M, He Y, Gao Z, Chen G, Tang D (2015) Isothermal cycling and cascade signal amplification strategy for ultrasensitive colorimetric detection of nucleic acids. Microchim Acta 182(1):449–454.  https://doi.org/10.1007/s00604-014-1385-4 CrossRefGoogle Scholar
  7. 7.
    Tanner NA, Evans TC (2014) Loop-mediated isothermal amplification for detection of nucleic acids. Curr Protoc Mol Biol 105:15.14. 1–15.14. 14Google Scholar
  8. 8.
    Zhang X, Lowe SB, Gooding JJ (2014) Brief review of monitoring methods for loop-mediated isothermal amplification (LAMP). Biosens Bioelectron 61:491–499.  https://doi.org/10.1016/j.bios.2014.05.039 CrossRefGoogle Scholar
  9. 9.
    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):523–530.  https://doi.org/10.1007/s00604-014-1354-y CrossRefGoogle Scholar
  10. 10.
    Wang Y, Li H, Wang Y, Zhang L, Xu J, Ye C (2017) Loop-mediated isothermal amplification label-based gold nanoparticles lateral flow biosensor for detection of enterococcus faecalis and staphylococcus aureus. Front Microbiol 8:192Google Scholar
  11. 11.
    Yongkiettrakul S, Jaroenram W, Arunrut N, Chareanchim W, Pannengpetch S, Suebsing R, Kiatpathomchai W, Pornthanakasem W, Yuthavong Y, Kongkasuriyachai D (2014) Application of loop-mediated isothermal amplification assay combined with lateral flow dipstick for detection of Plasmodium falciparum and Plasmodium vivax. Parasitol Int 63(6):777–784CrossRefGoogle Scholar
  12. 12.
    Plaon S, Longyant S, Sithigorngul P, Chaivisuthangkura P (2015) Rapid and sensitive detection of Vibrio alginolyticus by loop-mediated isothermal amplification combined with a lateral flow dipstick targeted to the rpoX gene. J Aquat Anim Health 27(3):156–163CrossRefGoogle Scholar
  13. 13.
    Mori Y, Kanda H, Notomi T (2013) Loop-mediated isothermal amplification (LAMP): recent progress in research and development. J Infect Chemother 19(3):404–411CrossRefGoogle Scholar
  14. 14.
    Santiago-Felipe S, Tortajada-Genaro LA, Puchades R, Maquieira Á (2016) Parallel solid-phase isothermal amplification and detection of multiple DNA targets in microliter-sized wells of a digital versatile disc. Microchim Acta 183(3):1195–1202.  https://doi.org/10.1007/s00604-016-1745-3 CrossRefGoogle Scholar
  15. 15.
    Nurul Najian AB, Engku Nur Syafirah EAR, Ismail N, Mohamed M, Yean CY (2016) Development of multiplex loop mediated isothermal amplification (m-LAMP) label-based gold nanoparticles lateral flow dipstick biosensor for detection of pathogenic Leptospira. Anal Chim Acta 903:142–148.  https://doi.org/10.1016/j.aca.2015.11.015 CrossRefGoogle Scholar
  16. 16.
    Salo P, Ortqvist A, Leinonen M (1995) Diagnosis of bacteremic pneumococcal pneumonia by amplification of pneumolysin gene fragment in serum. J Infect Dis 171(2):479–482CrossRefGoogle Scholar
  17. 17.
    Wang Y, Wang Y, Xu J, Ye C (2016) Development of multiple cross displacement amplification label-based gold nanoparticles lateral flow biosensor for detection of shigella spp. Front Microbiol 7:1834.  https://doi.org/10.3389/fmicb.2016.01834 Google Scholar
  18. 18.
    Le Rouzic E (2006) Contamination-pipetting: relative efficiency of filter tips compared to Microman® positive displacement pipette. Nat Methods 3(6).  https://doi.org/10.1038/nmeth887
  19. 19.
    Barhate RS, Ramakrishna S (2007) Nanofibrous filtering media: filtration problems and solutions from tiny materials. J Membr Sci 296(1):1–8CrossRefGoogle Scholar
  20. 20.
    Wang Y, Wang Y, Zhang L, Li M, Luo L, Liu D, Li H, Cao X, Hu S, Jin D, Xu J, Ye C (2016) Endonuclease restriction-mediated real-time polymerase chain reaction: a novel technique for rapid, sensitive and quantitative detection of nucleic-acid sequence. Front Microbiol 7:1104.  https://doi.org/10.3389/fmicb.2016.01104 Google Scholar
  21. 21.
    Zhao H, Dong J, Zhou F, Li B (2015) G-quadruplex − based homogenous fluorescence platform for ultrasensitive DNA detection through isothermal cycling and cascade signal amplification. Microchim Acta 182(15):2495–2502.  https://doi.org/10.1007/s00604-015-1608-3 CrossRefGoogle Scholar
  22. 22.
    Hagiwara E, Baba T, Shinohara T, Kitamura H, Sekine A, Komatsu S, Ogura T (2017) Ten-year trends and clinical relevance of the antimicrobial resistance genotype in respiratory isolates of streptococcus pneumoniae. Chemotherapy 62(4):256–261.  https://doi.org/10.1159/000470828 CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Austria, part of Springer Nature 2018

Authors and Affiliations

  • Yi Wang
    • 1
  • Yan Wang
    • 1
  • Dongxun Li
    • 2
  • Jianguo Xu
    • 1
  • Changyun Ye
    • 1
  1. 1.State Key Laboratory of Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, National Institute for Communicable Disease Control and PreventionChinese Center for Disease Control and PreventionBeijingPeople’s Republic of China
  2. 2.Changping District Center for Disease Control and PreventionBeijingPeople’s Republic of China

Personalised recommendations