Veterinary World

     Open access and peer reviewed journal  

ISSN (Online): 2231-0916

ISSN (Print): 0972-8988


Home l Editorial board l Instructions for authors l Reviewer guideline l Open access policy l Archives l FAQ

Open Access

Copyright: The authors. This article is an open access article licensed under the terms of the Creative Commons Attribution License

( which permits unrestricted use, distribution and reproduction in any medium, provided the work is properly cited.

Research (Published online: 13-10-2014)

11. Detection of African swine fever virus from formalin fixed and non-fixed tissues by polymerase chain reaction - P. D. Luka, A. R. Jambol and B. Yakubu

Veterinary World, 7(10): 811-815



   doi: 10.14202/vetworld.2014.811-815



P. D. Luka: Applied Molecular Biology Department, National Veterinary Research Institute, Vom, Plateau State, Nigeria;

A. R. Jambol: Applied Molecular Biology Department, National Veterinary Research Institute, Vom, Plateau State, Nigeria;

B. Yakubu; Applied Molecular Biology Department, National Veterinary Research Institute, Vom, Plateau State, Nigeria;


Received: 17-06-2014, Revised: 08-09-2014, Accepted: 11-09-2014, Published online: 13-10-2014


Corresponding author: P. D. Luka, e-mail:

Aim: Formalin fixing and paraffin embedding of tissue samples is one of the techniques for preserving the structural integrity of cells for a very long time. However, extraction and analysis of genomic material from formalin fixed tissue (FFT) remains a challenge despite numerous attempts to develop a more effective method. The success of polymerase chain reaction (PCR) depends on the quality of DNA extract.

Materials and Methods: Here we assessed the conventional method of DNA extraction from FFT for African swine fever virus (ASFV) detection. The modified conventional method gave a higher quality DNA when compared with commercially available DNA extraction kits (QIAamp® DNA Mini Kit, DNeasy® Blood and Tissue Kit, and ZR Genomic DNATissue MiniPrep).

Results: An average A260/A280 DNA purity of 0.86-1.68 and 3.22-5.32 μg DNA/mg for formalin fixed and non-fixed tissues, respectively using a conventional method. In a reproducible and three times repeat PCR, the ASFV DNA expected product size of 278 bp was obtained from the DNA extract of the conventional method but not from the DNA extract of the commercial kits.

Conclusion: The present study has demonstrated that the conventional method extracts ASFV genome better than commercial kit. In summary, the commercial kit extraction appeared not suitable to purify ASFV DNA from FFT. We, therefore, recommend that the use of the conventional method be considered for African swine fever DNA extraction from FFT.

Keyword: African swine fever, detection, formalin fixed, non-formalin fixed, polymerase chain reaction.

1. Dixon, L.K., Escribano, J.M., Martins, C., Rock, D.L., Salas, M.L. and Wilinson, P.J. (2005). Asfarviridae. In: Fauquet CM, Mayo MA, Maniloff J, Desselberger U, Ball LA, editors. Virus Taxonomy, VIIIth Report of the ICTV. Elsevier/Academic Press, London (UK). p135-143.
2. Jori, F., Vial, L., Penrith, M.L., Pérez-Sánchez, R., Etter, E., Albina, E., Michaud V. and Roger, F. (2012) Review of the sylvatic cycle of African swine fever in sub-Saharan Africa and the Indian Ocean. Virus. Res., 173(1): 212-227.
3. Babalobi, O.O., Olugasa, B.O., Oluwayelu, D.O., Ijagbone, I.F., Ayoade, G.O. and Agbede, S.A. (2007) Analysis and evaluation of mortality losses of the 2001 African swine fever outbreak, Ibadan, Nigeria. Trop. Anim. Health Prod., 39(7): 533-542.
4. Gallardo, C., Anchuelo, R., Pelayo, V., Poudevigne, F., Leon, T., Nzoussi, J., Bishop, R., Pérez, C., Soler, A., Nieto, R., Martín, H. and Arias, M. (2011) African Swine fever virus p72 genotype Ix in domestic pigs, Congo, 2009. Emerg. Infect Dis., 17(8): 1556-1558.
PMid:21801650 PMCid:PMC3381578
5. Cubillos, C., Gómez-Sebastianb, S., Morenoa, N., Nunez, M.C., Mulumba-Mfumuc, L.K., Quembod, C.J., Heathe, L., Etter, E.M.C., Jori, F., Escribano, J.M. and Blanco, E. (2013) African swine fever virus serodiagnosis: A general review with a focus on the analyses of African serum samples. Virus Res., 173(1): 159-167.
6. Owolodun, O.A., Yakubu, B., Antiabong, J.F., Ogedengbe, M.E., Luka, P.D., John Audu, B., Ekong, P.S. and Shamaki, D. (2010) Spatio-Temporal dynamics of African swine fever outbreaks in Nigeria, 2002-2007. Transbound. Emerg. Dis., 57(5): 330-9.
7. Sánchez-Vizcaíno, M.J., Martínez-López, B., Martínez-Avilés, M., Martins, C., Boinas, F., Vial, L., Michaud, V., Jori, F., Etter, E., Albina, E. and Roger, F. (2009) Scientific Report Submitted to EFSA on African Swine Fever, p1-141.
8. Gallardo, C., Soler, A., Nieto, R., Carrascosa, A.L., De Miac, G.M., Bishop R.P., Martinse, C., Fasina, F.O., Couacy-Hymman, E., Heath, L., Pelayo, V., Martı’n, E., Simon, A., Martın, R., Okurut, A.R., Lekolol, I., Okoth, E. and Arias, M. (2012) Comparative evaluation of novel African swine fever virus (ASF) antibody detection techniques derived from specific ASF viral genotypes with the OIE internationally prescribed serological tests. Vet. Microbiol., 162(1): 32-43.
9. Wandeler, P., Hoeck, P.E. and Keller, L.F. (2007) Back to the future: Museum specimens in population genetics. Trends Ecol. Evol., 22(12): 634-642.
10. Von Ahlfen, S., Missel, A., Bendrat, K. and Schlumpberger, M. (2007) Determinants of quality from FFPE samples. PLoS. One., 2(12): e1261.
PMid:18060057 PMCid:PMC2092395
11. Kayser, K., Stute, H., Lübcke, J. and Wazinski U. (1988) Rapid microwave fixation - a comparative morphometric study. Histochem. J., 20(6-7): 347-352.
12. Perlmutter, M.A., Best, C.J., Gillespie, J.W., Gathright, Y., González, S., Velasco, A., Linehan, W.M., Emmert-Buck, M.R. and Chuaqui, R.F. (2004) Comparison of snap freezing versus ethanol fixation for gene expression profiling of tissue specimens. J. Mol. Diagn., 6(4): 371-377.
13. Gilbert, M.T., Haselkorn, T., Bunce, M., Sanchez, J.J., Lucas, S.B., Jewell, L.D., Van Marck, E. and Worobey, M. (2007) The isolation of nucleic acids from fixed, paraffin embedded tissues-which methods are useful when? PLoS One., 2(6): e537.
PMid:17579711 PMCid:PMC1888728
14. Gillio-Tos, A., De Marco, L., Fiano, V., Garcia-Bragado, F., Dikshit, R., Boffetta, P. and Merletti, F. (2007) Efficient DNA extraction from 25-year-old paraffin-embedded tissues: Study of 365 samples. Pathology., 39(3): 345-348.
15. Poinar, H.N. and Stankiewikz, A.B. (1999) Protein preservation and DNA retrieval from ancient tissues. Proc. Natl. Acad. Sci., 96(15): 8426-8431.
PMid:10411891 PMCid:PMC17532
16. Lin, J., Kenedy, S.H., Svarovsky, T., Rogers, J., Kemnitz, J., Xu, A. and Zondervan, K.T. (2009) High quality genomic DNA extraction from formalin-fixed and paraffin-embedded samples deparaffinized using mineral oil. Anal. Biochem., 395(2): 265-267.
PMid:19698695 PMCid:PMC2764035
17. Huang, W., Sheehy, T.M., Moore, L.E., Hsing, A.W. and Purdue, M.P. (2010) Simultaneous recovery of DNA and RNA from formalin-fixed paraffin-embedded tissue and application in epidemiologic studies. Cancer Epidemiol. Biomarkers. Prev., 19(4): 973-7.
PMid:20332269 PMCid:PMC2864144
18. Campos, P.F., Thomas, M. and Gilbert, P. (2012) DNA extraction from formalin-fixed material. Methods Mol. Biol., 840: 81-85.
19. Giammarioli, M., Gallardo, C., Oggiano, A., Iscaro, C., Nieto, R., Pellegrini, C., Dei Giudici, S., Arias, M. and De Mia, G.M. (2011) Genetic characterization of African swine fever viruses from recent and historical outbreaks in Sardinia (1978-2009). Virus Genes., 42(3): 377-387.
20. Koidl, C., Bozic, M., Hadzisejdic, I., Grahovac, M., Grahovac, B., Kranewitter, W., Marth, E. and Kessler, H.H. (2008) Comparison of molecular assays for detection and typing of human papillomavirus. Am. J. Obstet. Gynecol., 199(2): 144.
21. Klaassen, C.H., Prinsen, C.F., De Valk, H.A., Horrevorts, A.M., Jeunink, M.A. and Thunnissen, F.B. (2004) DNA microarray format for detection and subtyping of human papillomavirus. J. Clin. Microbiol., 42(5): 2152-2160.
PMid:15131183 PMCid:PMC404665
22. Atuhaire, D.K., Afayoa, M., Ochwo, S., Mwesigwa, S., Okuni, J.B., Olaho-Mukani, W. and Ojok, L. (2013) Molecular characterization and phylogenetic study of African swine fever isolates from recent outbreaks in Uganda (2010-2013). Virol. J., 10: 247.
PMid:23914918 PMCid:PMC3735491
23. Cao, W., Hashibe, M., Rao, J.Y., Morgenstern, H. and Zhang, Z.F. (2003) Comparison of methods for DNA extraction from paraffin-embedded tissues and buccal cells. Cancer Detect. Prev., 27(5): 397-404.
24. Huijsmans, C.J.J., Damen, J., Van Der Linden J.C., Savelkoul, P.H.M. and Hermans, M.H.A. (2010) Comparative analysis of four methods to extract DNAfrom paraffin-embedded tissues: Effect on downstream molecular applications. BMC Res. Notes., 3: 239.
PMid:20840759 PMCid:PMC2954845
25. Mirmomeni, H.M., Sujjadi Majd, S., Sisakhtnezhad, S. and Doranegard, F. (2010) Comparison of the three methods for the extraction of paraffin-embedded tissues. J. Biol. Sci., 10(3): 261-266.