Sel’skokhozyaistvennaya Biologiya [Agricultural Biology], 2018, Vol. 53, № 2, p. 430-437.
(how to cite this article)

doi: 10.15389/agrobiology.2018.2.430eng

UDC 636.4:619:578:612.017.1:57.083

Acknowledgements:
The authors thank V.P. Bolobolova N.G. Sai, Goloskok P.V. (Belarus State Veterinary Center, Minsk, Belarus) and V.M. Lyska (FRC for Virology and Microbiology, Russia) for help in the research trials.

Supported financially by a subsidy from Ministry of Education and Science of the Russian Federation, State Contract № 14.601.21.0016, unique agreement identifier: RFMEFI60117X0016

 

VALIDATION OF A TEST SYSTEM FOR AFRICAN SWINE FEVER
SERODIAGNOSIS USING IMMUNOBLOTTING

O.A. Dubrovskaya, A.D. Sereda, A.S. Kazakova, A.R. Imatdinov,
O.M. Strizhakova, A.P. Vasil’ev, I.V. Nogina, M.E. Vlasov,
V.M. Balyshev, D.V. Kolbasov

Federal Research Center for Virology and Microbiology, Federal Agency of Scientific Organizations, 1, ul. Akademika Bakuleva, pos. Vol’ginskii, Petushinskii Region, Vladimir Province, 601125 Russia, e-mail sereda-56@mail.ru (✉ corresponding author), olgadubrovskaya@list.ru, almazlcf@yandex.ru, annakazakova85@yandex.ru, omstr@ya.ru, apvas@list.ru, irinanogina1911@gmail.com, vlasovmikhail1993@yandex.ru, balyshevvm@rambler.ru, kolbsovdenis@gmail.com

ORCID:
Dubrovskaya O.A. orcid.org/0000-0002-3168-7947
Nogina I.V. orcid.org/0000-0002-1787-5546
Sereda A.D. orcid.org/0000-0001-8300-5234
Vlasov M.E. orcid.org/0000-002-8324-3256
Kazakova A.S. orcid.org/0000-0002-0126-9023
Balyshev V.M. orcid.org/0000-0001-8224-9333
Imatdinov A.R. orcid.org/0000-0003-2889-6112
Kolbasov D.V. orcid.org/0000-0002-4935-0891
Strizhakova O.M. orcid.org/0000-0003-0023-0028

Received November 22, 2017

 

Because of the lack of a vaccine, African swine fever (ASF, caused by African swine fever virus (ASFV) of Asfivirus genus, Asfarviridae family) control strategy is based on making a rapid and early diagnosis and taking strict veterinary and sanitary measures. In the Eastern Europe countries where the infection has currently spread, highly virulent isolates are usually detected (J.M. Sanchez-Vizcaino et al., 2013). In the laboratory diagnosis, polymerase chain reaction (PCR) and direct immunofluorescence method are predominantly used. However, since 2012, researchers have observed some alteration in biological and genetic properties of a number of ASFV isolates. Therefore, serological methods may become prevalent in the laboratory diagnosis as it was during an ASF epizooty in the Iberian Peninsula in 1960-1990. We have earlier reported the development of a test system for the disease immunoblotting serodiagnosis (Rec p30-IB) based on a recombinant structural ASFV protein p30 (A.S. Kazakova et al., 2014). In this paper, the Rec p30-IB test system validation is shown. The diagnostic sensitivity of the Rec p30-IB was of 99.3 %, and the specificity was 100 %. Antibodies against p30 were detected in blood serum and organ samples taken from domestic pigs or wild boars irrespective of the seroimmunotypic membership and the virulence levels of the ASFV strains. In the blood serum samples collected from domestic pigs infected with heterologous viruses, no false-positive results were seen. In the serum of domestic pigs which were survived after intramuscular injection of attenuated strains LK-111, KK-262/С, MK-200, FK-135, PSA-1-NH and SCA 2015 VNIIVViM at 103 to 104 HAU50/CPE50, antibodies to p30 were detected on day 7 to 10. For organ samples from domestic pigs that had died from ASF 5 to 10 days post intramuscular infection with highly virulent strains Lisbon-57, Mozambique-78 or Stavropol 01/08 at a dose of 103 HAE50, the antibodies to p30 were detected in 30 % of the animals. The validation results indicate that the Immunoblotting Test System for African Swine Fever Serodiagnosis (Rec p30-IB) can be used for laboratory practice and monitoring of blood sera and organ samples collected from ASFV-infected domestic pigs or wild boars.

Keywords: African swine fever, protein p30 ASFV, serodiagnosis, immunoblotting, validation.

 

Full article (Rus)

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REFERENCES

  1. Dixon L.K., Costa J.V., Escribano J.M., Rock D.L., Vinuela E., Wilkinson P.J. Family Asfarviridae. M.H.V.V. Regenmortel (ed.). London Academic Press, 2000: 159-165.
  2. Sereda A.D., Balyshev V.M. Voprosy virusologii, 2011, 4: 38-42.
  3. Malogolovkin A., Burmakina G., Titov I., Sereda A., Gogin A., Baryshnikova E., Kolbasov D. Comparative analysis of African swine fever virus genotypes and serogroups. Emerg. Infect. Dis., 2015, 21(2): 312-315 CrossRef
  4. Gogin A., Gerasimov V., Malogolovkin A., Kolbasov D. African swine fever in the North Caucasus region and the Russian Federation in years 2007-2012. Virus Res., 2013, 173: 198-203 CrossRef
  5. Abrahantes J.C., Gogin A., Richardson J., Gervelmeyer A. Epidemiological analyses on African swine fever in the Baltic countries and Poland. EFSA Journal, 2017, 15(3): 4732 CrossRef
  6. Sanchez-Vizcaino J.M., Mur L., Martinez-Lopez B. African swine fever (ASF): five years around Europe. Vet. Microbiol., 2013, 165: 45-50 CrossRef
  7. Gallardo C., Fernández-Pinero J., Pelayo V., Gazaev I., Markowska-Daniel I., Pridotkas G., Nieto R, Fernández-Pacheco P., Bokhan S., Nevolko O., Drozhzhe Z., Pérez C., Soler A., Kolvasov D., Arias M. Genetic variation among African swine fever genotype II viruses, Eastern and Central Europe. Emerg. Infect. Dis., 2014, 20(9): 1544-1547 CrossRef
  8. Vlasova N.N., Varentsova A.A., Shevchenko I.V., Zhukov I.Yu., Remyga S.G., Gavrilo-
    va V.L., Puzankova O.S., Shevtsov A.A, Zinyakova N.G., Gruzdev K.N. Comparative analysis of clinical and biological characteristics of African swine fever virus isolates from 2013 year Russian Federation. British Microbiology Research Journal. 2015, 5(3): 203-215 CrossRef
  9. Goller K.V., Malogolovkin A.S., Katorkin S., Kolbasov D., Titov I., Hoper D., Beer M., Keil G.M., Portugal R., Blome S. Tandem repeat insertion in African swine fever virus. Russia, 2012. Emerg. Infect. Dis., 2015, 21(4): 731-732 CrossRef
  10. Fr?czyk M., Wo?niakowski G., Kowalczyk A., Bocian ?., Kozak E., Niemczuk K., Pejsak Z. Evolution of African swine fever virus genes related to evasion of host immune response. Vet. Microbiol., 2016, 25(193): 133-44 CrossRef
  11. Reis A.L., Parkhouse R.M.E., Penedos A.R., Martins C., Leitão A. Systematic analysis of longitudinal serological responses of pigs infected experimentally with African swine fever virus. J. Gen. Virol., 2007, 88(9): 2426-2434 CrossRef
  12. Arias M., Escribano J.M., Sánchez-Vizcaíno J.M. Persistence of African swine fever antibody reactivity on ELISA and immunoblotting assays. Vet. Rec., 1993, 133(8): 189-190.
  13. Pastor M.J., Laviada M.D., Sanchez-Vizcaino J.M., Escribano J.M. Detection of African swine fever virus antibodies by immunoblotting assay. Can. J. Vet. Res., 1989, 53(1): 105-107.
  14. World Organization for Animal Health (OIE), 2012. African swine fever. Chapter 2.8.1. In: Ma-nual of diagnostic tests and vaccines for terrestrial animals (mammals, birds and bees). Available http://www.oie.int/fileadmin/Home/eng/Health_standards/tahm/2.08.01_ASF.pdf. No date.
  15. Escribano J.M., Pastor M.J., Sánchez-Vizcaino J.M. Antibodies to bovine serum albumin in swine sera: implications for false-positive reactions in the serodiagnosis of African swine fever. Am. J. Vet. Res., 1989, 50: 1118-1122.
  16. Alcaraz C., Rodriguez F., Oviedo J.M., Eiras A., De Diego M., Alonso C., Escribano J.M. Highly specific confirmatory western blot test of African swine fever virus antibody detection using the recombinant virus protein p54. J. Virol. Methods, 1995, 52(1-2): 111-119 CrossRef
  17. Kollnberger S.D., Gutierrez-Castaneda B., Foster-Cuevas M., Corteyn A., Parkhouse R.M. Identification of the principal serological immunodeterminants of African swine fever virus by screening a virus cDNA library with antibody. J. Gen. Virol., 2002, 83(6): 1331-1342 CrossRef
  18. Gallardo C., Reis A.L., Kalema-Zikusoka G., Malta J., Soler A., Blanco E., Parkhouse R.M., Leitão A. Recombinant antigen targets for serodiagnosis of African swine fever. Clin. Vaccine Immunol., 2009, 16(7): 1012-1020 CrossRef
  19. Gallardo C., Dufton M.M., Macharia J.M., Arias M., Taracha E.A., Soler A., Okoth E., Martin E., Kasiti J., Bishop R.P. Enhanced discrimination of African swine fever virus isolates through nucleotide sequencing of the p54, p72, and pB602L (CVR) genes. Virus Genes, 2009, 38: 85-95 CrossRef
  20. Oviedo J.M., Rodríguez F., Gómez-Puertas P., Brun A., Gómez N., Alonso C., Escribano J.M. High level expression of the major antigenic African swine fever virus proteins p54 and p30 in baculovirus and their potential use as diagnostic reagents. J. Virol. Methods, 1997, 64(1): 27-35 CrossRef
  21. Pérez-Filgueira D.M., González-Camacho F., Gallardo C., Resino-Talaván P., Blanco E., Gómez-Casado E., Alonso C., Escribano J.M. Optimization and validation of recombinant serological tests for African swine fever diagnosis based on detection of the p30 protein produced in Trichoplusia ni larvae. J. Clin. Microbiol., 2006, 44 (9): 3114-3121 CrossRef
  22. Kazakova A.S., Imatdinov I.R., Dubrovskaya O.A., Imatdinov A. R., Sidlik M.V., Balyshev V.M., Krasochko P.A., Sereda A.D. Recombinant protein p30 for serological diagnosis of African swine fever by immunoblotting assay. Transbound. Emerg. Dis., 2017, 64: 1479-1492 CrossRef
  23. Sereda A.D., Dubrovskaya O.A., Imatdinov A.R., Strizhakova O.M., Vasil’ev A.P., Sindryako-va I.P., Lunitsin A.V. Laboratory diagnostics of chronic and asymptomatic forms of African swine fever. Agricultural Biology, 2016, 51(4): 459-466 CrossRef
  24. Prudnikova E.Yu. Adaptatsiya virusa afrikanskoi chumy svinei, vydelennogo na territorii Rossiiskoi Federatsii, k perevivaemym kul'turam kletok i izuchenie ego biologicheskikh svoistv. Kandidatskaya dissertatsiya [Adaptation of the African swine fever virus Russian isolate to transplantable cell cultures and the study of its biological properties. PhD Thesis]. Pokrov, 2013. Available http://search.rsl.ru/ru/record/01005543218. No date (in Russ.).
  25. Balyshev V.M., Kalantaenko Yu.F., Bolgova M.V., Prudnikova E.Yu. Doklady RASKHN, 2011, 5: 52-53 (in Russ.).
  26. Strizhakova O.M., Lyska V.M., Malogolovkin A.S., Novikova M.B., Sidlik M.V., Nogina I.V., Shkaev A.E., Balashova E.A., Kurinnov V.V., Vasil’ev A.P.Validation of an ELISA kit for detection of antibodies against ASF virus in blood or spleen of domestic pigs and wild boars. Aricultural Biology, 2016, 51(6): 845-852 CrossRef
  27. Cubillos C., Gómez-Sebastian S., Moreno N., Nuñez M.C., Mulumba-Mfumu L.K., Quembode C.J., Heath L., Etter E.M.C., Jori F., Escribano J.M., Blanco E. African swine fever virus serodiagnosis: a general review with a focus on the analyses of African serum samples. Virus Res., 2013, 173: 159-167 CrossRef

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