ASF virus replication features in the presence of recombinant proteins CD2v, pX69R and pE248R

Cover Page


Cite item

Full Text

Abstract

Introduction. African swine fever (ASF), sever hemorrhagic disease of swine caused by a large DNA virus of the Asfaviridae family.

Since there are no effective and safe vaccines against ASF yet, it is urgent to study the functions of its proteins, which is applicable by analyzing the features of ASF virus replication in the presence of recombinant proteins in vitro.

Purpose. To study the effect of ASFV recombinant proteins CD2v, pE248R and pX69R on the speed and level of reproduction of ASF virus in vitro. Thus, obtain the necessary knowledge to develop approaches for creating a vaccine against ASF.

Materials and methods. ASFV isolate Krasnodar 07/17 and strain ASF/ARRIAH/CV-1 were used. Cloning of X69R, EP402R, and E248R genes was performed in the pJET1.2 / blunt vector and pCI-neo in E. coli JM-109 cells, according to the manufacturer's manual. Localization of recombinant proteins in CV-1 cell line carried out by direct immunofluorescence reaction (DIF) using polyclonal antibodies conjugated to FITC.

The ASF virus reproduction level was assessed by hemadsorption reaction and qPCR kit (Central Research Institute of Epidemiology).

Results. Recombinant plasmids pCI-neo / E248R, pCI-neo / EP402R and pCI-neo / X69R were constructed.

The localization and the specificity of the obtained recombinant proteins CD2v, pE248R and pX69R was confirmed. It was established that these recombinant proteins induce the level of ASF virus reproduction on days 3-5 of the experiment by ~ 1.2-1.5 lgHADU50/cm3 in comparison with the negative control.

Discussion. The data obtained demonstrate the important role of CD2v, pX69R and pE248R proteins in the reproduction of the virus, since they significantly affect its level. The exact function of pX69R protein was not determined, however, in the experiments its positive effect on ASF virus reproduction was established, manifested in an increase in its reproduction level.

Conclusion. This methodology allows us to study the nature of the effect of proteins with unknown function on ASF virus replication.

About the authors

A. Mazloum

«ARRIAH» Federal State Budgetary Institution «Federal Center for Animal Health»

Author for correspondence.
Email: ali.mazloum6@gmail.com
ORCID iD: 0000-0002-5982-8393

Mazloum Ali - Ph.D.

Vladimir region, Vladimir city, Yuryevets microdistrict, 600901.

Russian Federation

I. U. Zhukov

«ARRIAH» Federal State Budgetary Institution «Federal Center for Animal Health»

Email: fake@neicon.ru
ORCID iD: 0000-0002-3817-2129

Vladimir region, Vladimir city, Yuryevets microdistrict, 600901.

Russian Federation

E. B. Aronova

«ARRIAH» Federal State Budgetary Institution «Federal Center for Animal Health»

Email: fake@neicon.ru
ORCID iD: 0000-0002-2072-6701

Vladimir region, Vladimir city, Yuryevets microdistrict, 600901.

Russian Federation

A. S. Igolkin

«ARRIAH» Federal State Budgetary Institution «Federal Center for Animal Health»

Email: fake@neicon.ru
ORCID iD: 0000-0002-5438-8026

Vladimir region, Vladimir city, Yuryevets microdistrict, 600901.

Russian Federation

N. N. Vlasova

«ARRIAH» Federal State Budgetary Institution «Federal Center for Animal Health»

Email: fake@neicon.ru
ORCID iD: 0000-0001-8707-7710

Vladimir region, Vladimir city, Yuryevets microdistrict, 600901.

Russian Federation

References

  1. Garner G., Saville P., Fediavsky A., eds. African swine fever. Available at: http://lrd.spc.int/ext/Disease_Manual_Final/a120african_swine_fever.html
  2. Rowlands R.J., Michaud V., Heath L., Hutchings G., Oura C., Vosloo W., et al. African swine fever virus isolate, Georgia, 2007. Emerg. Infect. Dis. 2008; 14(12): 1870-4. Doi: https://doi.org/10.3201/eid1412.080591
  3. Gallardo C., Nieto R., Soler A., Pelayo V., Fernandez-Pinero J., Markowska-Daniel I., et al. Assessment of African swine fever diagnostic techniques as a response to the epidemic outbreaks in Eastern European Union Countries: How to improve surveillance and control programs. J. Clin. Microbiol. 2015; 53(8): 2555-65. Doi: https://doi.org/10.1128/JCM.00857-15
  4. EFSA Panel on Animal Health and Welfare (AHAW). African swine fever virus. EFSA J. 2015; 13(7): 4163-92. Doi: https://doi.org/10.2903/j.efsa.2015.4163
  5. World Organisation for Animal Health (OIE). African swine fever in Moldova. Immediate notification report. REF OIE: 21095. Available at: http://www.oie.int/wahis_2/temp/reports/en_imm_0000021095_20161004_170450.pdf
  6. OIE, 2017. World Animal Health Information Disease (WAHID). Paris, France: World Organisation of Animal Health.
  7. OIE, 2018. World Animal Health Information Disease (WAHID). Paris, France: World Organisation of Animal Health.
  8. World Organisation for Animal Health. African swine fever in Cote d’Ivoire. Immediate notification report. REF OIE: 15914. Available at: http://www.oie.int/wahis_2/temp/reports/en_imm_0000015914_20140828_131035.pdf
  9. Болгова М.В., Балышев В.М., Пономарев В.Н., Неверовская Н.С. Паспортизация изолята «Девис» вируса африканской чумы свиней. В кн.: Материалы международной научно-практической конференции «Актуальные вопросы контроля инфекционных болезней животных». Часть 1. Покров; 2014: 43-7.
  10. Galindo I., Hernaez B., Diaz-Gil G., Escribano J.M., Alonso C. A179L, a viral Bcl-2 homologue, targets the core Bcl-2 apoptotic machinery and its upstream BH3 activators with selective binding restrictions for Bid and Noxa. Virology. 2008; 375(2): 561-72. Doi: https://doi.org/10.1016/j.virol.2008.01.050
  11. Rodriguez J.M., Yanez R.J., Almazan F., Vinuela E., Rodriguez J.F. African swine fever virus encodes a CD2 homolog responsible for the adhesion of erythrocytes to infected cells. J. Virol. 1993; 67(9): 5312-20.
  12. Galindo I., Almazan F., Bustos M.J., Vinuela E., Carrascosa A.L. African swine fever virus EP153R open reading frame encodes a glycoprotein involved in the hemadsorption of infected cells. Virology. 2000; 266(2): 340-51. Doi: https://doi.org/10.1006/viro.1999.0080
  13. Rowlands R.J., Duarte M.M., Boinas F., Hutchings G., Dixon L.K. The CD2v protein enhances African swine fever virus replication in the tick vector, Ornithodoros erraticus. Virology. 2009; 393(2): 319¬28. Doi: https://doi.org/10.1016/j.virol.2009.07.040
  14. Rodriguez I., Nogal M.L., Redrejo-Rodriguez M., Bustos M.J., Salas M.L. The African swine fever virus virion membrane protein pE248R is required for virus infectivity and an early postentry event. J. Virol. 2009; 83(23): 12290-300. Doi: https://doi.org/10.1128/JVI.01333-09
  15. Xiang Z., Mobley H.L.T. Vaxign: the first web-based vaccine design program for reverse vaccinology and applications for vaccine development. J. Biomed. Biotechnol. 2010; 2010: 297505. Doi: https://doi.org/10.1155/2010/297505
  16. Xiang Z., He Y. Genome-wide prediction of vaccine targets for humanherpes simplex viruses using Vaxign reverse vaccinology. BMC Bioinformatics. 2013; 14(Suppl. 4): S2. Doi: https://doi.org/10.1186/1471-2105-14-S4-S2
  17. Мазлум А., Зиняков Н.Г., Иголкин А.С., Власова Н.Н. Клонирование генов, кодирующих трансмембранные белки и белки, ответственные за вирулентность вируса африканской чумы свиней. Ветеринария сегодня. 2018; (2): 3-7. Doi: https://doi.org/10.29326/2304-196X-2018-2-25-3-7
  18. Sambrook J., Russell D.W. The Condensed Protocols from Molecular Cloning: a Laboratory Manual. New York: Cold Spring Harbor; 2006.
  19. Burgess R.R. Elution of proteins from gels.MethodsEnzymol. 2009; 463: 565-72. Doi: https://doi.org/10.1016/S0076-6879(09)63032-9
  20. Мазлум А., Шарыпова Д.В., Гаврилова В.Л. и др. Методические рекомендации по выделению и титрованию вируса африканской чумы свиней в культуре клеток селезёнки свиней. Владимир; 2019.
  21. Мазлум А. и др. Методические рекомендации по оценке уровня репродукцию вируса африканской чумы свиней с использованием полимеразной цепной реакции в режиме реального времени. Владимир; 2019.
  22. Мазлум А., Жуков И.Ю., Першин А.С., Иголкин А.С., Власова Н.Н. Влияние рекомбинантного белка p30 на репродукцию вируса африканской чумы свиней in vitro. Ветеринария сегодня. 2018; (3): 3-7. Doi: https://doi.org/10.29326/2304-196X-2018-3-26-3-7
  23. Мазлум А., Власова Н.Н., Аронова Е.В., Иголкин А.С., Кривонос Р.А., Черных О.Ю. Определение корреляции показателя Ct и титра вируса африканской чумы свиней в биологических жидкостях. Ветеринария Кубани. 2018; 24(6): 4-7.
  24. Власова Н.Н., Жуков И.Ю., Мазлум А., Шарыпова Д.В., Першин А.С., Иголкин А.С. Штамм «АЧС/ВНИИЗЖ/ CV-1» вируса африканской чумы свиней, со сниженной вирулентностью для свиней, для вирусологических, диагностических, молекулярно-генетических и мониторинговых исследований. Патент РФ № 2675535; 2019.

Supplementary files

Supplementary Files
Action
1. JATS XML
Download

Copyright (c) 2019 Mazloum A., Zhukov I.U., Aronova E.B., Igolkin A.S., Vlasova N.N.

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.
СМИ зарегистрировано Федеральной службой по надзору в сфере связи, информационных технологий и массовых коммуникаций (Роскомнадзор).
Регистрационный номер и дата принятия решения о регистрации СМИ: серия ПИ № ФС77-77676 от 29.01.2020.


This website uses cookies

You consent to our cookies if you continue to use our website.

About Cookies