Loop-mediated isothermal amplification (LAMP) allows rapid amplification of nucleic acids under isothermal conditions. In this report, a 20-min LAMP amplification of the DPOL gene of infectious spleen and kidney necrosis virus (ISKNV) using a biotin-labeled primer was combined with lateral flow dipstick (LFD) chromatography for rapid and simple visual detection of ISKNV-specific amplicons. The LFD process involves a 5-min specific hybridization with an FITC-labeled DNA probe to confirm the presence of complement ISKNV amplicons that were biotinated in LAMP. The resulting DNA duplexes, consisting of labeled probes and amplicons, migrate along the LFD strip by chromatography for 5 min and are trapped at the test line and visualized by biotin labeling. The detection limit of ISKNV by LAMP-LFD was 10 copies. The results show that the LAMP-LFD method has the advantages of better sensitivity and speed and less dependence on equipment than the standard PCR for specifically detecting low levels of ISKNV DNA, and this can be useful in the field as a routine diagnostic tool.
This is a preview of subscription content, log in to check access.
Buy single article
Instant access to the full article PDF.
Price includes VAT for USA
Subscribe to journal
Immediate online access to all issues from 2019. Subscription will auto renew annually.
This is the net price. Taxes to be calculated in checkout.
Chinchar G, Essbauer S, He JG, Hyatt A, MIyazaki T (2005) Family iridoviridae. In: Fauquet CM, Mayo MA, Maniloff J, Desselberger U, Ball LA (eds) Virus taxonomy classification and nomenclature of viruses eight report of the international committee on the taxonomy of viruses. Academic Press, San Diego, pp 145–161
Jeong JB, Kim HY, Jun LJ, Lyu JH, Park NG, Kim JK, Jeong HD (2008) Outbreaks and risks of infectious spleen and kidney necrosis virus disease in freshwater ornamental fishes. Dis Aquat Organ 78:209–215
He JG, Zeng K, Weng SP, Chan SM (2002) Experimental transmission, pathogenicity and physical–chemical properties of infectious spleen and kidney necrosis virus (ISKNV). Aquaculture 204:11–24
Wang YQ, Lu L, Weng SP, Huang JN, Chan SM, He JG (2007) Molecular epidemiology and phylogenetic analysis of a marine fish infectious spleen and kidney necrosis virus-like (ISKNV-like) virus. Arch Virol 152:763–773
He JG, Deng M, Weng SP, Li Z, Zhou SY, Long QX, Wang XZ, Chan SM (2001) Complete genome analysis of the mandarin fish infectious spleen and kidney necrosis iridovirus. Virology 291:126–139
Deng M, He JG, Weng SP, Zeng K, Zeng Z, Long QX (2000) Infectious spleen and kidney necrosis virus (ISKNV) from Siniperca chuatsi: development of a PCR detection method and the new evidence of iridovirus. Chin J Virol 16:365–369
Lao HH, Ye X, Zou WM, Bai JJ, Tan AP (2009) Detection of infectious spleen and kidney necrosis virus (ISKNV) of mandarin fish (Siniperca chuatsi) by nested PCR. South China Fisheries Science 5:69–72
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:e63
Kiatpathomchai W, Jaroenram W, Arunrut N, Jitrapakdee S, Flegel TW (2008) Shrimp Taura syndrome virus detection by reverse transcription loop-mediated isothermal amplification combined with a lateral flow dipstick. J Virol Methods 153:214–217
Jaroenram W, Kiatpathomchai W, Flegel TW (2009) Rapid and sensitive detection of white spot syndrome virus by loop-mediated isothermal amplification combined with a lateral flow dipstick. Mol Cell Probes 23:65–70
Puthawibool T, Senapin S, Kiatpathomchai W, Flege TW (2009) Detection of shrimp infectious myonecrosis virus by reverse transcription loop-mediated isothermal amplification combined with a lateral flow dipstick. J Virol Methods 156:27–31
Wang XW, Ao JQ, Li QJ, Chen XH (2005) Quantitative detection of a marine fish iridovirus isolated from large yellow croaker, Pseudosciaena crocea, using a molecular beacon. J Virol Methods 133:76–81
Brice SL, Stockert SS, Jester JD, Huff JC, Bunker JD, Weston WL (1992) Detection of herpes simplex virus DNA in the peripheral blood during acute recurrent herpes labialis. J Am Acad Dermatol 26:594–598
Miranda MB, Handermann M, Darai G (2005) DNA polymerase gene locus of Cercopithecine herpesvirus 1 is a suitable target for specific and rapid identification of viral infection by PCR technology. Virus Genes 30:307–322
Tomoya K, Ram S, Masahiro S, Toshiaki I (2004) Detection of white spot syndrome virus in shrimp by loop-mediated isothermal amplification. J Virol Methods 115:59–65
Caipang CM, Haraguchi I, Ohira T, Hirono I, Aoki T (2004) Rapid detection of a fish iridovirus using loop-mediated isothermal amplification (LAMP). J Virol Methods 121:155–161
Gunimaladevi I, Kono T, Lapatra SE, Sakai M (2005) A loop mediated isothermal amplification (LAMP) method for detection of infectious hematopoietic necrosis virus (IHNV) in rainbow trout (Oncorhynchus mykiss). Arch Virol 150:899–909
Nagamine K, Hase T, Notomi T (2002) Accelerated reaction by loop-mediated isothermal amplication using loop primers. Mol Cell Probes 16:223–229
Endo S, Komori T, Ricci G, Sano A, Yokoyama K, Ohori A, Kamei K, Franco M, Miyaji M, Nishimura K (2004) Detection of gp43 of Paracoccidioides brasiliensis by the loop-mediated isothermal amplification (LAMP) method. FEMS Microbiol Lett 234:93–97
Blomström AL, Hakhverdyan M, Reid SM, Dukes JP, King DP, Belák S, Berg M (2008) A one-step reverse transcriptase loop-mediated isothermal amplification assay for simple and rapid detection of swine vesicular disease virus. J Virol Methods 147:188–193
Pillai D, Bonami JR, Widada JS (2006) Rapid detection of Macrobrachium rosenbergii nodavirus (MrNV) and extra small virus (XSV), the pathogenic agents of white tail disease of Macrobrachium rosenbergii (De Man), by loop-mediated isothermal amplification. J Fish Dis 29:275–283
Teng PH, Chen CL, Sung PF, Lee FC, Ou BR, Lee PY (2007) Specific detection of reverse transcription-loop-mediated isothermal amplification amplicons for Taura syndrome virus by colorimetric dot–blot hybridization. J Virol Methods 146:317–326
We thank Prof. Junjie Bai (Pearl River Fishery Research Institute, Chinese Academy of Fishery Science, Guangzhou city, China) for kindly providing ISKNV and Dr. Dengfeng Li (Ningbo University, Ningbo city, China) for kindly providing WSSV. Ms. Wenchao Ding was a student registered in Ningbo University for the degree of MSc. The project was supported by the Program of Science and Technology Department of Zhejiang Province (No. 2008C22049; No. 2008R40G2070059) and the Program for Changjiang Scholars and Innovative Research Team in University (IRT0734) and the KC Wong Magna Fund in Ningbo University.
About this article
Cite this article
Ding, W.C., Chen, J., Shi, Y.H. et al. Rapid and sensitive detection of infectious spleen and kidney necrosis virus by loop-mediated isothermal amplification combined with a lateral flow dipstick. Arch Virol 155, 385–389 (2010). https://doi.org/10.1007/s00705-010-0593-4
- White Spot Syndrome Virus
- Lamp Assay
- Lamp Reaction
- Lamp Primer
- Taura Syndrome Virus