Skip to main content
SpringerLink
Log in

Rapid detection of bovine herpesvirus 1 in bovine semen by loop-mediated isothermal amplification (LAMP) assay

  • Original Article
  • Published:
Archives of Virology Aims and scope Submit manuscript

Abstract

Bovine herpesvirus 1 (BoHV-1) is the most common viral pathogen found in bovine semen, causing numerous reproductive disorders leading to economic losses to the cattle industry. For rapid detection of BoHV-1 in bovine semen, in this study, we applied a loop-mediated isothermal amplification (LAMP) assay. The assay could be completed within 90 min, including total DNA isolation, target amplification, and visual interpretation of positive or negative results with the naked eye. The assay detected as little as 10 fg of BoHV-1 DNA per reaction. The analytical sensitivity of the assay was 0.2 TCID50 BoHV-1 per reaction, which was 100 times more sensitive than conventional PCR and comparable to TaqMan real-time PCR. The applicability of the assay was assessed by analysing 118 semen samples collected from breeding bulls. On comparison with TaqMan real-time PCR, the LAMP assay had a diagnostic sensitivity of 97 %, specificity of 100 %, and accuracy of 99.2 % for detection of BoHV-1 in bovine semen. The LAMP assay developed in this study is a rapid, sensitive, and cost-effective alternative for detection of BoHV-1 in bovine semen.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  1. Nandi S, Kumar M, Manohar M, Chauhan RS (2009) Bovine herpes virus infections in cattle. Anim Health Res Rev 10:85–98

    Article  PubMed  CAS  Google Scholar 

  2. OIE (2010) Infectious bovine rhinotracheitis/infectious pustular vulvovaginitis. OIE Terrestrial Manual, Chapter 2.4.13

  3. Muylkens B, Thiry J, Kirten P, Schynts F, Thiry E (2007) Bovine herpesvirus 1 infection and infectious bovine rhinotracheitis. Vet Res 38:181–209

    Article  PubMed  CAS  Google Scholar 

  4. Gibbs EP, Rweyemamu MM (1977) Bovine herpesviruses, part 1. Vet Bull 47:317–343

    Google Scholar 

  5. Miller JM, Whetstone CA, Van der Maaten MJ (1991) Abortifacient property of bovine herpesvirus type 1isolates that represent three subtypes determined by restriction endonuclease analysis of viral DNA. Am J Vet Res 52:458–461

    PubMed  CAS  Google Scholar 

  6. Turin L, Russo S, Poli G (1999) BoHV-1: new molecular approaches to control a common and widespread infection. Mol Med 5:261–284

    PubMed Central  PubMed  CAS  Google Scholar 

  7. Jones C, Newby TJ, Holt T, Doster A, Stone MJ, Webster CJ, Jackwood MW (2000) Analysis of latency in cattle after inoculation with a temperature sensitive mutant of bovine herpesvirus 1 (RLB106). Vaccine 18:3185–3195

    Article  PubMed  CAS  Google Scholar 

  8. Van der Oirschot JT, Straver PJ, van Lieshout JAH, Quak J, Westenbrink F, van Exsel ACA (1993) A subclinical infection of bulls with bovine herpesvirus type 1 at an artificial insemination centre. Vet Rec 132:32–35

    Article  PubMed  Google Scholar 

  9. Afshar A, Eaglesome MD (1990) Viruses associated with bovine semen. Vet Bull 60:93–109

    Google Scholar 

  10. Wyler R, Engels M, Schwyzer M (1989) Infectious bovine rhinotracheitis/vulvovaginitis (BoHV-1). In: Witman G (ed) Herpesvirus diseases of cattle, horses, and pigs. Kluwer Academic Publishers, Boston, pp 1–72

    Chapter  Google Scholar 

  11. Van der Oirschot JT (1995) Bovine herpes virus 1 in semen of bulls and the risk of transmission: a brief review. Vet Quart 17:29–33

    Article  Google Scholar 

  12. Eaglesome MD, Garcia MM (1997) Disease risks to animal health from artificial insemination with bovine semen. Rev Sci Tech Off Int Epiz 16:215–225

    CAS  Google Scholar 

  13. Monke DR, Howard TH (1992) IBR research update and commentary. In: Proceedings 14th technical conference on artificial insemination and reproduction, vol 14. National Association of Animal Breeders, Milwaukee, pp 26–32

  14. Dennet DP, Barasa JO, Johnson RH (1976) Infectious bovine rhinotracheitis virus: studies on the venereal carrier state in range cattle. Res Vet Sci 20:77–83

    Google Scholar 

  15. Schultz RD, Adams LS, Letchworth G, Sheffy BE, Manning T, Bean B (1982) A method to test large numbers of bovine semen samples for viral contamination and results of a study using this method. Theriogenology 17:115–123

    Article  PubMed  CAS  Google Scholar 

  16. Weiblen R, Kreutz LC, Canabarro TF, Schuch LF, Rebelatto M (1992) Isolation of bovine herpesvirus 1 from preputial swabs and semen of bulls with balanoposthitis. J Vet Diagn Invest 4:341–343

    Article  PubMed  CAS  Google Scholar 

  17. Brunner D, Engels M, Schwyzer M, Wyle R (1988) A comparison of three techniques for detecting bovine herpesvirus type 1 (BoHV-1) in naturally and experimentally contaminated bovine semen. Reprod Domest Anim 23:1–9

    Article  CAS  Google Scholar 

  18. Xia JQ, Lofstedt RM, Yason CV, Kibenge FS (1995) The use of a polymerase chain reaction assay for the detection of bovine herpesvirus 1 in the semen of experimentally infected bulls by dot blot hybridization, polymerase chain reaction and virus isolation. Res Vet Sci 59:183–185

    Article  PubMed  CAS  Google Scholar 

  19. Kataria RS, Tiwari AK, Gupta PK, Mehrotra ML, Rai A, Bandyopadhyay SK (1997) Detection of bovine herpesvirus 1 (BHV-1) genomic sequences in bovine semen inoculated with BHV-1 by polymerase chain reaction. Acta Virol 41:311–315

    PubMed  CAS  Google Scholar 

  20. van Engelenburg FA, Maes RK, van Oirschot JT, Rijsewijk FA (1993) Development of a rapid and sensitive polymerase chain reaction assay for detection of bovine herpesvirus type 1 in bovine semen. J Clin Microbiol 31:3129–3135

    PubMed Central  PubMed  Google Scholar 

  21. Oliveira MT, Campos FS, Dias MM, Velho FA, Freneau GE, Brito WM, Rijsewijk FA, Franco AC, Roehe PM (2011) Detection of bovine herpesvirus 1 and 5 in semen from Brazilian bulls. Theriogenology 75:1139–1145

    Article  PubMed  CAS  Google Scholar 

  22. Rocha MA, Barbosa EF, Guimaraes SE, Neto D, Gouveia AM (1998) A high sensitivity nested PCR assay for BHV-1 detection in semen of naturally infected bulls. Vet Microbiol 63:1–11

    Article  PubMed  CAS  Google Scholar 

  23. Diallo IS, Corney BG, Rodwell BJ (2011) Detection and differentiation of bovine herpesvirus 1 and 5 using a multiplex real time polymerase chain reaction. J Virol Methods 175:46–52

    Article  PubMed  CAS  Google Scholar 

  24. Wang J, O’keefe J, Orr D, Loth L, Banks M, Wakeley P, West D, Card R, Ibata G, Van Maanen K, Thoren P, Isaksson M, Kerkhofs P (2007) Validation of a real-time PCR assay for the detection of bovine herpesvirus 1 in bovine semen. J Virol Methods 144:103–108

    Article  PubMed  CAS  Google Scholar 

  25. 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

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  26. Fu S, Qu G, Guo S, Ma L, Zhang N, Zhang S, Gao S, Shen Z (2011) Applications of loop-mediated isothermal DNA amplification. Appl Microbiol Biot 163:845–850

    CAS  Google Scholar 

  27. Mehrotra ML, Rajya BS, Kumar S (1976) IBR keratoconjunctivitis in calves. Indian J Vet Pathol 1:70–73

    Google Scholar 

  28. Gupta PK, Rai A (1993) Restriction enzyme analysis of bovine herpesvirus-1 DNA from an Indian isolate. Biochem Mol Biol Int 29:889–898

    PubMed  CAS  Google Scholar 

  29. Wang J, O’Keefe J, Orr D, Loth L, Banks M, Wakeley P, West D, Card R, Ibata G, Van Maanen K, Thoren P, Isaksson M, Kerkhofs P (2008) An international inter-laboratory ring trial to evaluate a real-time PCR assay for the detection of bovine herpesvirus 1 in extended bovine semen. Vet Microbiol 126:11–19

    Article  PubMed  CAS  Google Scholar 

  30. Mori Y, Nagamine K, Tomita N, Notomi T (2001) Detection of loop-mediated isothermal amplification reaction by turbidity derived from magnesium pyrophosphate formation. Biochem Bioph Res Co 289:150–154

    Article  CAS  Google Scholar 

  31. Nagamine KT, Hase T, Notomi T (2002) Accelerated reaction by loop-mediated isothermal amplification using loop primers. Mol Cell Probe 16:223–229

    Article  CAS  Google Scholar 

  32. 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 46:167–172

    Article  PubMed  CAS  Google Scholar 

  33. Parida M, Horioke K, Ishida H, Dash PK, Saxena P, Jana AM, Islam MA, Inoue S, Hosaka N, Morita K (2005) Rapid detection and differentiation of dengue virus serotypes by a real time reverse transcription loop-mediated isothermal amplification assay. J Clin Microbiol 43:2895–2903

    Article  PubMed Central  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This study was supported by a project of the National Fund for Basic, Strategic and Frontier Application Research in Agriculture (NFBSFARA), Indian Council of Agricultural Research (ICAR), Government of India. We sincerely thank ICAR and Indian Veterinary Research Institute (IVRI) for providing necessary facilities to carry out this work.

Author information

Author notes

  1. Sachin S. Pawar

    Present address: Project Directorate on foot-and-mouth disease, Mukteswar, 263138, India

Authors and Affiliations

  1. Division of Veterinary Biotechnology, Indian Veterinary Research Institute, Izatnagar, 243122, India

    Sachin S. Pawar, Chetan D. Meshram, Arvind A. Sonwane, B. P. Mishra & Praveen K. Gupta

  2. School of Animal Biotechnology, Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana, 141001, India

    Niraj K. Singh

  3. Center for Wildlife, Indian Veterinary Research Institute, Izatnagar, 243122, India

    Mohini Saini

  4. Disease Investigation Section, Animal Husbandry Department, Pune, 411007, India

    Sunil S. Rautmare

  5. Department of Microbiology, K.N.P. College of Veterinary Science, Shirwal, 412801, India

    Dushyant M. Muglikar

Authors
  1. Sachin S. Pawar
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Chetan D. Meshram
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Niraj K. Singh
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Arvind A. Sonwane
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Mohini Saini
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Sunil S. Rautmare
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Dushyant M. Muglikar
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. B. P. Mishra
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Praveen K. Gupta
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Praveen K. Gupta.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Pawar, S.S., Meshram, C.D., Singh, N.K. et al. Rapid detection of bovine herpesvirus 1 in bovine semen by loop-mediated isothermal amplification (LAMP) assay. Arch Virol 159, 641–648 (2014). https://doi.org/10.1007/s00705-013-1869-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00705-013-1869-2

Keywords

Search

Navigation