Abstract
Varicella-zoster virus (VZV) infects more than 90% of the population worldwide and has a high incidence of postherpetic neuralgia in elderly patients, seriously affecting their quality of life. Combined with clustered regularly interspaced short palindromic repeats (CRISPR) system, we develop a quantum dot nanobeads (QDNBs) labeled lateral flow assay for VZV detection. Our assay allows the identification of more than 5 copies of VZV genomic DNA in each reaction. The entire process, from sample preparation to obtaining the results, takes less than an hour. In 86 clinical vesicles samples, the test shows 100% concordance with quantitative real-time PCR for VZV detection. Notably, when vesicles are present in specific areas, such as the genitals, our method outperforms clinical diagnosis. Compared to traditional detection methods, only a minute amount of blister fluid is required for accurate detection. Therefore, we anticipate that our method could be translated to clinical applications for specific and rapid VZV detection.
Key Points
• CRISPR/Cas12a and quantum dot nanobead-based lateral flow assay achieved 5 copies per reaction for VZV detection
• Specific identification of VZV in atypical skin lesions
• Results read by the naked eye within one hour
Graphical Abstract
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 analyzed during this study are included in this published article (and its supplementary information files).
References
Bhaya D, Davison M, Barrangou R (2011) CRISPR-Cas systems in bacteria and archaea: versatile small RNAs for adaptive defense and regulation. Annu Rev Genet 45:273–297. https://doi.org/10.1146/annurev-genet-110410-132430
Bienes KM, Mao L, Selekon B, Gonofio E, Nakoune E, Wong G, Berthet N (2022) Rapid detection of the varicella-zoster virus using a recombinase-aided amplification-lateral flow system. Diagnostics (Basel, Switzerland) 12(12). https://doi.org/10.3390/diagnostics12122957
Binkhamis K, Al-Siyabi T, Heinstein C, Hatchette TF, LeBlanc JJ (2014) Molecular detection of varicella zoster virus while keeping an eye on the budget. J Virol Methods 202:24–27. https://doi.org/10.1016/j.jviromet.2014.02.009
Broughton JP, Deng X, Yu G, Fasching CL, Servellita V, Singh J, Miao X, Streithorst JA, Granados A, Sotomayor-Gonzalez A, Zorn K, Gopez A, Hsu E, Gu W, Miller S, Pan CY, Guevara H, Wadford DA, Chen JS, Chiu CY (2020) CRISPR-Cas12-based detection of SARS-CoV-2. Nat Biotechnol 38(7):870–874. https://doi.org/10.1038/s41587-020-0513-4
Chen JS, Ma E, Harrington LB, Da Costa M, Tian X, Palefsky JM, Doudna JA (2018) CRISPR-Cas12a target binding unleashes indiscriminate single-stranded DNase activity. Science 360(6387):436–439. https://doi.org/10.1126/science.aar6245
Chen X, Zhou Q, Li S, Yan H, Chang B, Wang Y, Dong S (2021) Rapid and visual detection of SARS-CoV-2 using multiplex reverse transcription loop-mediated isothermal amplification linked with gold nanoparticle-based lateral flow biosensor. Front Cell Infect Microbiol 11:581239. https://doi.org/10.3389/fcimb.2021.581239
Chen X, Zhou Q, Tan Y, Wang R, Wu X, Liu J, Liu R, Wang S, Dong S (2022) Nanoparticle-based lateral flow biosensor integrated with loop-mediated isothermal amplification for rapid and visual identification of Chlamydia trachomatis for point-of-care use. Front Microbiol 13:914620. https://doi.org/10.3389/fmicb.2022.914620
Chen Y, Shi Y, Chen Y, Yang Z, Wu H, Zhou Z, Li J, Ping J, He L, Shen H, Chen Z, Wu J, Yu Y, Zhang Y, Chen H (2020) Contamination-free visual detection of SARS-CoV-2 with CRISPR/Cas12a: A promising method in the point-of-care detection. Biosens Bioelectron 169:112642. https://doi.org/10.1016/j.bios.2020.112642
Deng H, Liu Q, Wang X, Huang R, Liu H, Lin Q, Zhou X, Xing D (2017) Quantum dots-labeled strip biosensor for rapid and sensitive detection of microRNA based on target-recycled nonenzymatic amplification strategy. Biosens Bioelectron 87:931–940. https://doi.org/10.1016/j.bios.2016.09.043
Forbes HJ, Thomas SL, Smeeth L, Clayton T, Farmer R, Bhaskaran K, Langan SM (2016) A systematic review and meta-analysis of risk factors for postherpetic neuralgia. Pain 157(1):30–54. https://doi.org/10.1097/j.pain.0000000000000307
Gershon AA, Breuer J, Cohen JI, Cohrs RJ, Gershon MD, Gilden D, Grose C, Hambleton S, Kennedy PG, Oxman MN, Seward JF, Yamanishi K (2015) Varicella zoster virus infection. Nat Rev Dis Primers 1:15016. https://doi.org/10.1038/nrdp.2015.16
Gross GE, Eisert L, Doerr HW, Fickenscher H, Knuf M, Maier P, Maschke M, Müller R, Pleyer U, Schäfer M, Sunderkötter C, Werner RN, Wutzler P, Nast A (2020) S2k guidelines for the diagnosis and treatment of herpes zoster and postherpetic neuralgia. J Dtsch Dermatol Ges 18(1):55–78. https://doi.org/10.1111/ddg.14013
Huang X, Aguilar ZP, Xu H, Lai W, Xiong Y (2016) Membrane-based lateral flow immunochromatographic strip with nanoparticles as reporters for detection: a review. Biosens Bioelectron 75:166–180. https://doi.org/10.1016/j.bios.2015.08.032
Ivanov AV, Safenkova IV, Zherdev AV, Dzantiev BB (2020) Nucleic acid lateral flow assay with recombinase polymerase amplification: solutions for highly sensitive detection of RNA virus. Talanta 210:120616. https://doi.org/10.1016/j.talanta.2019.120616
Jiang N, Ahmed R, Damayantharan M, Ünal B, Butt H, Yetisen AK (2019) Lateral and vertical flow assays for point-of-care diagnostics. Adv Healthc Mater 8(14):e1900244. https://doi.org/10.1002/adhm.201900244
Karakkat BB, Hockemeyer K, Franchett M, Olson M, Mullenberg C, Koch PL (2018) Detection of root-infecting fungi on cool-season turfgrasses using loop-mediated isothermal amplification and recombinase polymerase amplification. J Microbiol Methods 151:90–98. https://doi.org/10.1016/j.mimet.2018.06.011
Kobayashi T, Yagami A, Suzuki K, Ihira M, Yoshikawa T, Matsunaga K (2013) Clinical utility of loop-mediated isothermal amplification assay for the diagnosis of common alpha herpesvirus skin infections. J Dermatol 40(12):1033–1037. https://doi.org/10.1111/1346-8138.12325
Laing KJ, Ouwendijk WJD, Koelle DM, Verjans G (2018) Immunobiology of varicella-zoster virus infection. J Infect Dis 218(suppl_2):S68–S74. https://doi.org/10.1093/infdis/jiy403
Li SY, Cheng QX, Liu JK, Nie XQ, Zhao GP, Wang J (2018a) CRISPR-Cas12a has both cis- and trans-cleavage activities on single-stranded DNA. Cell Res 28(4):491–493. https://doi.org/10.1038/s41422-018-0022-x
Li SY, Cheng QX, Wang JM, Li XY, Zhang ZL, Gao S, Cao RB, Zhao GP, Wang J (2018b) CRISPR-Cas12a-assisted nucleic acid detection. Cell Discov 4:20. https://doi.org/10.1038/s41421-018-0028-z
Lv H, Wang J, Zhang J, Chen Y, Yin L, Jin D, Gu D, Zhao H, Xu Y, Wang J (2021) Definition of CRISPR Cas12a trans-cleavage units to facilitate CRISPR diagnostics. Front Microbiol 12:766464. https://doi.org/10.3389/fmicb.2021.766464
Min SW, Kim YS, Nahm FS, Yoo DH, Choi E, Lee PB, Choo H, Park ZY, Yang CS (2016) The positive duration of varicella zoster immunoglobulin M antibody test in herpes zoster. Medicine 95(33):e4616. https://doi.org/10.1097/md.0000000000004616
Mukama O, Yuan T, He Z, Li Z, Habimana JD, Hussain M, Li W, Yi Z, Liang Q, Zeng L (2020) A high fidelity CRISPR/Cas12a based lateral flow biosensor for the detection of HPV16 and HPV18. Sens Actuators B Chem 316:128119. https://doi.org/10.1016/j.snb.2020.128119
Oliver SL, Xing Y, Chen DH, Roh SH, Pintilie GD, Bushnell DA, Sommer MH, Yang E, Carfi A, Chiu W, Arvin AM (2021) The N-terminus of varicella-zoster virus glycoprotein B has a functional role in fusion. PLoS Pathog 17(1):e1008961. https://doi.org/10.1371/journal.ppat.1008961
Patchsung M, Jantarug K, Pattama A, Aphicho K, Suraritdechachai S, Meesawat P, Sappakhaw K, Leelahakorn N, Ruenkam T, Wongsatit T, Athipanyasilp N, Eiamthong B, Lakkanasirorat B, Phoodokmai T, Niljianskul N, Pakotiprapha D, Chanarat S, Homchan A, Tinikul R et al (2020) Clinical validation of a Cas13-based assay for the detection of SARS-CoV-2 RNA. Nat Biomed Eng 4(12):1140–1149. https://doi.org/10.1038/s41551-020-00603-x
Resch-Genger U, Grabolle M, Cavaliere-Jaricot S, Nitschke R, Nann T (2008) Quantum dots versus organic dyes as fluorescent labels. Nat Methods 5(9):763–775. https://doi.org/10.1038/nmeth.1248
Schmader K (2018) Herpes Zoster. Ann Intern Med 169(3):Itc19–itc31. https://doi.org/10.7326/aitc201808070
Sohrabi H, Majidi MR, Fakhraei M, Jahanban-Esfahlan A, Hejazi M, Oroojalian F, Baradaran B, Tohidast M, Guardia M, Mokhtarzadeh A (2022) Lateral flow assays (LFA) for detection of pathogenic bacteria: a small point-of-care platform for diagnosis of human infectious diseases. Talanta 243:123330. https://doi.org/10.1016/j.talanta.2022.123330
Strich JR, Chertow DS (2019) CRISPR-Cas biology and its application to infectious diseases. J Clin Microbiol 57(4). https://doi.org/10.1128/jcm.01307-18
Wang M, Zhang R, Li J (2020a) CRISPR/Cas systems redefine nucleic acid detection: principles and methods. Biosens Bioelectron 165:112430. https://doi.org/10.1016/j.bios.2020.112430
Wang X, Ji P, Fan H, Dang L, Wan W, Liu S, Li Y, Yu W, Li X, Ma X, Ma X, Zhao Q, Huang X, Liao M (2020b) CRISPR/Cas12a technology combined with immunochromatographic strips for portable detection of African swine fever virus. Commun Biol 3(1):62. https://doi.org/10.1038/s42003-020-0796-5
Warren-Gash C, Forbes H, Maple P, Quinlivan M, Breuer J (2018) Viral load and antibody boosting following herpes zoster diagnosis. J Clin Virol 103:12–15. https://doi.org/10.1016/j.jcv.2018.03.010
Yan W, Fan L, Li J, Wang Y, Han H, Tan F, Zhang P (2020) Bimodal size distribution immuno-quantum dots for fluorescent western blotting assay with high sensitivity and extended dynamic range. Mikrochim Acta 187(11):598. https://doi.org/10.1007/s00604-020-04578-z
Zhang P, Lu H, Chen J, Han H, Ma W (2014) Simple and sensitive detection of HBsAg by using a quantum dots nanobeads based dot-blot immunoassay. Theranostics 4(3):307–315. https://doi.org/10.7150/thno.8007
Zhang Q, Li J, Li Y, Tan G, Sun M, Shan Y, Zhang Y, Wang X, Song K, Shi R, Huang L, Liu F, Yi Y, Wu X (2022) SARS-CoV-2 detection using quantum dot fluorescence immunochromatography combined with isothermal amplification and CRISPR/Cas13a. Biosens Bioelectron 202:113978. https://doi.org/10.1016/j.bios.2022.113978
Zhao J, Ao C, Wan Z, Dzakah EE, Liang Y, Lin H, Wang H, Tang S (2021) A point-of-care rapid HIV-1 test using an isothermal recombinase-aided amplification and CRISPR Cas12a-mediated detection. Virus Res 303:198505. https://doi.org/10.1016/j.virusres.2021.198505
Zheng C, Wang K, Zheng W, Cheng Y, Li T, Cao B, Jin Q, Cui D (2021) Rapid developments in lateral flow immunoassay for nucleic acid detection. The Analyst 146(5):1514–1528. https://doi.org/10.1039/d0an02150d
Zhou B, Ye Q, Li F, Xiang X, Shang Y, Wang C, Shao Y, Xue L, Zhang J, Wang J, Ding Y, Chen M, Wu Q (2022) CRISPR/Cas12a based fluorescence-enhanced lateral flow biosensor for detection of Staphylococcus aureus. Sens Actuators B Chem 351. https://doi.org/10.1016/j.snb.2021.130906
Funding
This study was supported by grants from Program of the Science and Technology Commission of Shanghai Municipality “Science and Technology Innovation Action Plan” Medical Innovation Research Special Project (grant/award number: 22Y11905700), Program of Shanghai Municipal Health Commission (grant/award number: 2020YJZX0108), and Anhui Provincial Education Department Scientific Research Project (Grant/Award Number: YJS20210323).
Author information
Authors and Affiliations
Contributions
XZ: investigation, methodology, data curation, and writing—original draft. QF: project administration and data curation. YW: project administration and data curation. LL: investigation and data curation. WJ: investigation and sample collection. MC: data curation. HX: data curation. PZ: supervision and conceptualization. FT: supervision and writing—review and editing. All authors read and approved the manuscript.
Corresponding authors
Ethics declarations
Ethics approval
This study was performed in line with the principles of the Declaration of Helsinki. Approval was granted by the Clinical Trial Ethics Committee of Shanghai Dermatology Hospital (no. 201811).
Conflict of interest
The authors declare no competing interests.
Additional information
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary materials
Rights and permissions
Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.
About this article
Cite this article
Zhong, X., Fu, Q., Wang, Y. et al. CRISPR-based quantum dot nanobead lateral flow assay for facile detection of varicella-zoster virus. Appl Microbiol Biotechnol 107, 3319–3328 (2023). https://doi.org/10.1007/s00253-023-12509-0
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00253-023-12509-0