Advertisement

Archives of Virology

, Volume 163, Issue 6, pp 1455–1461 | Cite as

Development of a reverse transcription recombinase-aided amplification assay for the detection of coxsackievirus A10 and coxsackievirus A6 RNA

  • Teng-fei Yan
  • Xin-na Li
  • Le Wang
  • Chen Chen
  • Su-xia Duan
  • Ju-ju Qi
  • Li-Xin Li
  • Xue-jun Ma
Original Article

Abstract

Hand, foot and mouth disease (HFMD) is a serious public health problem, and coxsackievirus A6 (CVA6) and coxsackievirus A10 (CVA10) are two of the major causative pathogens, in addition to enterovirus 71 (EV71) and coxsackievirus A16 (CVA16). A simple and rapid reverse transcription recombinase-aided amplification assay (RT-RAA) was developed for the detection of CVA10 and CVA6 in this study. The analytical sensitivity for detection of CVA10 and CVA6 at 95% probability by probit regression analysis was 35 copies per reaction and 38 copies per reaction, respectively, with 100% specificity. Compared with commercial RT-qPCR assays, when testing 455 fecal specimens, the kappa value of the RT-RAA assay for CVA10 and CVA6 was 0.920 (p < 0.001) and 0.952 (p < 0.001), respectively. Moreover, four samples that were positive for CVA10 and five that were positive for CVA6 by RT-RAA but negative by RT-qPCR were further determined to be true positives. These results demonstrate that the proposed RT-RAA assays are very valuable tools for the detection of CVA10 and CVA6 and have potential for use in resource-limited settings.

Notes

Funding

This work was supported by grants from the National Key Research and Development Plan of China (2016TFC1202700, 2016YFC1200903 and 2017YFC1200503)and the China Mega-Project for Infectious Disease (2017ZX10302301-004 and 2017ZX10104001).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

All operations in the experiments were approved by the Institutional Review Boards of the National Institute for Viral Disease Control and Prevention, Center for Disease Control and Prevention of China. Caregivers of the children with HFMD were informed, and informed consent documents were signed.

References

  1. 1.
    Solomon T, Lewthwaite P, Perera D, Cardosa MJ, Mcminn P, Ooi MH (2010) Virology, epidemiology, pathogenesis, and control of enterovirus 71. Lancet Infectious Dis 10(11):778–790CrossRefGoogle Scholar
  2. 2.
    Yip CCY, Lau SKP, Woo PCY, Yuen KY (2013) Human enterovirus 71 epidemics: what’s next? Emerging Health Threats J 6(6):19780CrossRefGoogle Scholar
  3. 3.
    Lo SH, Huang YC, Huang CG, Tsao KC, Li WC, Hsieh YC, Chiu CH, Lin TY (2011) Clinical and epidemiologic features of Coxsackievirus A6 infection in children in northern Taiwan between 2004 and 2009. J Microbiol, Immunol, Infect 44(4):252CrossRefGoogle Scholar
  4. 4.
    Tryfonos C, Richter J, Koptides D, Yiangou M, Christodoulou CG (2011) Molecular typing and epidemiology of enteroviruses in Cyprus, 2003–2007. J Med Microbiol 60(10):1433–1440CrossRefPubMedGoogle Scholar
  5. 5.
    Gopalkrishna V, Patil PR, Patil GP, Chitambar SD (2012) Circulation of multiple enterovirus serotypes causing hand, foot and mouth disease in India. J Med Microbiol 61(Pt 3):420CrossRefPubMedGoogle Scholar
  6. 6.
    Yang F, Ren L, Xiong Z, Li J, Xiao Y, Zhao R, He Y, Bu G, Zhou S, Wang J (2009) Enterovirus 71 Outbreak in the People’s Republic of China in 2008. J Clin Microbiol 47(7):2351CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Oberste MS, Maher K, Kilpatrick DR, Pallansch MA (1999) Molecular evolution of the human enteroviruses: correlation of serotype with VP1 sequence and application to picornavirus classification. J Virol 73(3):1941–1948PubMedPubMedCentralGoogle Scholar
  8. 8.
    Perera D, Shimizu H, Yoshida H, Tu PV, Ishiko H, Mcminn PC, Cardosa MJ (2010) A comparison of the VP1, VP2, and VP4 regions for molecular typing of human enteroviruses. J Med Virol 82(4):649–657CrossRefPubMedGoogle Scholar
  9. 9.
    Zhu RN, Qian Y, Deng J, Xing JF, Zhao LQ, Wang F, Liao B, Ren XX, Li Y (2007) Zhang Q (2007) [Study on the association of hand, foot and mouth disease and enterovirus 71/CA16 among children in Beijing. Chin J Epidemiol 28(10):1004–1008Google Scholar
  10. 10.
    Bible JM, Pantelidis P, Chan PK, Tong CY (2007) Genetic evolution of enterovirus 71: epidemiological and pathological implications. Rev Med Virol 17(6):371–379CrossRefPubMedGoogle Scholar
  11. 11.
    Yan W, Yeo A, Phoon MC, Tan EL, Poh CL, Quak SH, Chow VTK (2010) The largest outbreak of hand; foot and mouth disease in Singapore in 2008: the role of enterovirus 71 and coxsackievirus A strains. Int J Infect Dis 14(12):e1076CrossRefGoogle Scholar
  12. 12.
    Mirand A, Henquell C, Archimbaud C, Ughetto S, Antona D, Bailly JL, Peigue-Lafeuille H (2012) Outbreak of hand, foot and mouth disease/herpangina associated with coxsackievirus A6 and A10 infections in 2010, France: a large citywide, prospective observational study. Clin Microbiol Infect 18(5):E110–E118CrossRefPubMedGoogle Scholar
  13. 13.
    Blomqvist S, Klemola P, Kaijalainen S, Paananen A, Simonen ML, Vuorinen T, Roivainen M (2010) Co-circulation of coxsackieviruses A6 and A10 in hand, foot and mouth disease outbreak in Finland. J Clin Virol 48(1):49–54CrossRefPubMedGoogle Scholar
  14. 14.
    Javier LDMD, Pablo HBMD, Violeta ZNMD, María ABPD, Fernando G-CPD, Antonio SMD, Gobernado M, Isabel FBMD (2011) Onychomadesis outbreak in valencia, spain associated with hand, foot, and mouth disease caused by enteroviruses. Pediatr Dermatol 28(1):1–5CrossRefGoogle Scholar
  15. 15.
    Xu M, Su L, Cao L, Zhong H, Dong N, Xu J (2013) Enterovirus genotypes causing hand foot and mouth disease in Shanghai, China: a molecular epidemiological analysis. BMC Infect Dis 13(1):489CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Chonmaitree T, Ford C, Sanders C, Lucia HL (1988) Comparison of cell cultures for rapid isolation of enteroviruses. J Clin Microbiol 26(12):2576PubMedPubMedCentralGoogle Scholar
  17. 17.
    Lim KA, Benyeshmelnick M (1960) Typing of viruses by combinations of antiserum pools. Application to typing of enteroviruses (Coxsackie and ECHO). J Immunol 84(3):309–317PubMedGoogle Scholar
  18. 18.
    Yolken RH, Torsch VM (1981) Enzyme-linked immunosorbent assay for detection and identification of coxsackieviruses A. Infect Immun 31(2):742–750PubMedPubMedCentralGoogle Scholar
  19. 19.
    Hu XM, Zhang Y, Xu BL, Yang MJ, Wang M, Zhang C, Li J, Bai RY, Zhou XM, Xu WB (2011) Development of a GeXP based multiplex RT-PCR assay for simultaneous differentiation of nine human hand food mouth disease pathogens. Bing Du Xue Bao 27(27):331–336PubMedGoogle Scholar
  20. 20.
    Fuschino ME, Lamson DM, Rush K, Carbone LS, Taff ML, Hua Z, Landi K, George KS (2012) Detection of coxsackievirus A10 in multiple tissues of a fatal infant sepsis case. J Clin Virol Off Publ Pan Am Soc Clin Virol 53(3):259–261CrossRefGoogle Scholar
  21. 21.
    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. Nucleic Acids Res 28(12):E63CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Nagamine K, Hase T, Notomi T (2002) Accelerated reaction by loop-mediated isothermal amplification using loop primers. Mol Cell Probes 16(3):223CrossRefPubMedGoogle Scholar
  23. 23.
    Zhang X, Guo L, Ma R, Cong L, Wu Z, Wei Y, Xue S, Zheng W, Tang S (2017) Rapid detection of Salmonella with recombinase aided amplification. J Microbiol Methods 139:202–204CrossRefPubMedGoogle Scholar
  24. 24.
    Bei L, Cheng HR, Yan QF, Huang ZJ, Shen GF, Zhang ZF, Yinv LI, Deng ZX, Lin M (2010) Recombinase-Aid Amplification:a Novel Technology of in vitro Rapid Nucleic Acid Amplification. Scientia SinicaGoogle Scholar
  25. 25.
    Chen C, Li X-n, Li G-x, Zhao L, Duan S-x, Yan T-f, Feng Z-s, Ma X-j Use of a Rapid Reverse-transcription Recombinase Aided Amplification Assay for Respiratory Syncytial Virus Detection. Diagnostic Microbiology and Infectious Disease.  https://doi.org/10.1016/j.diagmicrobio.2017.10.005
  26. 26.
    Nie K, Zhang Y, Luo L, Yang MJ, Hu XM, Wang M, Zhu SL, Han F, Xu WB, Ma XJ (2011) Visual detection of human enterovirus 71 subgenotype C4 and Coxsackievirus A16 by reverse transcription loop-mediated isothermal amplification with the hydroxynaphthol blue dye. J Virol Methods 175(2):283–286CrossRefPubMedGoogle Scholar
  27. 27.
    Nie K, Qi SX, Zhang Y, Luo L, Xie Y, Yang MJ, Zhang Y, Li J, Shen H, Li Q (2012) Evaluation of a direct reverse transcription loop-mediated isothermal amplification method without RNA extraction for the detection of human enterovirus 71 subgenotype C4 in nasopharyngeal swab specimens. PLoS One 7(12):e52486CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Guan H, Wang J, Wang C, Yang M, Liu L, Yang G, Ma X (2015) Etiology of multiple non-EV71 and non-CVA16 enteroviruses Associated with Hand, Foot and Mouth Disease in Jinan, China, 2009-June 2013. PLoS One 10(11):e0142733CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Tian H, Zhang Y, Sun Q, Zhu S, Li X, Pan Z, Xu W, Xu B (2014) Prevalence of Multiple Enteroviruses Associated with Hand, Foot, and Mouth Disease in Shijiazhuang City, Hebei Province, China: Outbreaks of Coxsackieviruses A10 and B3. PLoS One 9(1):e84233CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Oberste M, Maher K, Williams A, Dybdahl-Sissoko N, Brown B, Gookin M, Penaranda S, Mishrik N, Uddin M, Pallansch M (2006) Species-specific RT-PCR amplification of human enteroviruses: a tool for rapid species identification of uncharacterized enteroviruses. J Gen Virol 87(Pt 1):119CrossRefPubMedGoogle Scholar
  31. 31.
    Nasri D, Bouslama L, Pillet S, Bourlet T, Aouni M, Pozzetto B (2007) Basic rationale, current methods and future directions for molecular typing of human enterovirus. Expert Rev Mol Diagn 7(4):419–434CrossRefPubMedGoogle Scholar
  32. 32.
    Wang K, Wu Y, Yin D, Tang S, Hu G, He Y (2016) Development and evaluation of a rapid recombinase polymerase amplification assay for detection of coxsackievirus A6. Archives of Virology 1–4Google Scholar
  33. 33.
    Malek L, Sooknanan R, Compton J (2008) Nucleic acid sequence-based amplification (NASBA™). Humana PressGoogle Scholar
  34. 34.
    Brentano ST, Mcdonough SH (2000) Isothermal amplification of RNA by transcription-mediated amplification (TMA). Springer, BerlinCrossRefGoogle Scholar
  35. 35.
    Lizardi PM, Huang X, Zhu Z, Brayward P, Thomas DC, Ward DC (1998) Mutation detection and single-molecule counting using isothermal rolling-circle amplification. Nat Genet 19(3):225–232CrossRefPubMedGoogle Scholar
  36. 36.
    Ding X, Nie K, Shi L, Zhang Y, Guan L, Zhang D, Qi S, Ma X (2014) Improved detection limit in rapid detection of human enterovirus 71 and coxsackievirus A16 by a novel reverse transcription-isothermal multiple-self-matching-initiated amplification assay. J Clin Microbiol 52(6):1862–1870CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Wang J, Li R, Hu L, Sun X, Wang J, Li J (2015) Development of a quantitative fluorescence single primer isothermal amplification-based method for the detection of Salmonella. Int J Food Microbiol 219:22CrossRefPubMedGoogle Scholar
  38. 38.
    Xie XP, Huang LZ, Liu RW, Sheng-Fu HE, Chen HY, Chen JC, Tang L, Hospital GP (2014) Establishment of multiple fluorescent RT-PCR for simultaneously detecting Coxsackievirus A6,10 and 16 and its clinical application. Chin J Clin Lab Sci 10:721–725Google Scholar

Copyright information

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

Authors and Affiliations

  1. 1.Myasthenia Gravis Research InstituteThe First Hospital of ShijiazhuangShijiazhuangChina
  2. 2.Key Laboratory for Medical Virology, National Health and Family Planning Commission, National Institute for Viral Disease Control and PreventionChinese Center for Disease Control and PreventionBeijingChina
  3. 3.Pediatric Research InstituteChildren’s Hospital of Hebei ProvinceShijiazhuangChina

Personalised recommendations