Improvement of Enzyme-Linked Immunoelectrotransfer Blot Assay by Quantitative Approach for Foot-and-Mouth Disease diagnosis

Year 2022, Volume: 11 Issue: 2, 162 - 170, 30.12.2022

Can Çokçalışkan , Ünal Parlak , Beyhan Sareyyüpoğlu , Abdülnaci Bulut , Ayca Babak , Fuat Özyörük

https://doi.org/10.31196/huvfd.1113565

Abstract

Foot-and-mouth disease (FMD) is a highly contagious animal disease that causes devastating economic losses. The trade of live animals and derived products is only possible if the exporting country is free from disease, according to the World Organization for Animal Health (WOAH) code for FMD. One of the most important ways to prove disease-free is to measure the levels of non-structural protein antibodies (NPS) of FMD virus in the target population sampled. For detection of the disease status of a herd, mass screening and assays such as Enzyme-linked immunosorbent assay (ELISA) and Enzyme-linked immunoelectrotransfer blot assay (EITB) were developed and described in the WOAH diagnostic manual. In this study, recombinant FMDV NS proteins were produced and tested with sera panels collected from uninfected and naturally infected animals using a quantitative Western blot assay as an improved EITB, which enables numerical documentation and statistical analysis. NSP band intensities were used to determine the cut-off values, differentiating infected from non-infected animals and revealing variable sensitivity among the different NSPs. The quantitative EITB results also showed a correlation with the NSP-ELISA results.

Keywords

Enzyme-Linked Immunoelectrotransfer Blot Assay (EITB), Foot-and-Mouth Disease Virus (FMDV), Non-Structural Proteins (NSP), NSP-ELISA, Quantitative Western blot (QCWB).

Supporting Institution

Tarımsal Araştırmalar ve Politikalar Genel Müdürlüğü

Project Number

TAGEM/HS/01/02/07/118

Thanks

We would like to thank Graham Belsham from the Technical University of Denmark and Giulia Pezzoni from the IZSLER of Italy for their contributions.

Şap Hastalığı Teşhisi için Enzim Immunotransfer Blot Testinin kantitatif bir yaklaşım ile iyileştirilmesi

Abstract

Şap hastalığı büyük ekonomik kayıplara neden olan çok bulaşıcı bir hayvan hastalığıdır. Dünya Hayvan Sağlığı Örgütü Şap Hastalığı koduna göre canlı hayvanların ve bunların ürünlerinin ticareti, ancak, ihraç eden ülkenin hastalıktan ari olması şartıyla mümkündür. Hastalıktan ariliğin gösterilmesi için en önemli yöntemlerden biri örneklenen hedef popülasyonun Şap virüsü yapısal olmayan proteinlerine (YOP) karşı oluşan antikor seviyelerinin ölçümü temeline dayanır. Bir sürünün hastalık durumunun belirlenmesinde yaygın tarama ve Enzim bağlı Immunosorbent test (ELISA) ve Enzim bağlı Immunotransfer testi (EITB) geliştirilmiş ve WOAH (Dünya Hayvan Sağlığı Örgütü) teşhis el kitabında tanımlanmıştır. Bu çalışmada Şap virüsünün yapısal olmayan proteinleri rekombinant olarak üretilmiş, enfekte olmayan, aşılı ve doğal olarak enfekte olan hayvanlardan toplanan serumlardan oluşan panellerle, sayısal belgelemeye ve istatistik analize olanak veren, iyileştirilmiş, kantitatif bir Western blot kullanılarak test edilmiştir. YOP bant yoğunlukları, hasta ve enfekte hayvanların birbirinden ayırt edecek eşik değerlerin ve farklı YOP’lerin bu teşhiste farklı hassasiyet gösterdiğinin belirlenmesinde kullanılmıştır. Kantitatif EITB sonuçları aynı zamanda YOP-ELISA sonuçları ile korelasyon göstermiştir.

Keywords

Enzim-Bağlı İmmunoelekrotransfer Blot Testi (EITB), Şap Virüsü, Yapısal Olmayan Proteinler (YOP), YOP-ELISA, Kantitatif Western blot.

Project Number

TAGEM/HS/01/02/07/118

References

• Anonymous, 2011: Foot and mouth disease, Chapter 8.5. In O. T. A. H. S. Commission (ed.), Terrestrial Animal Health Code, 20th ed., OIE World Organisation for Animal Health, Paris, France.
• Bergmann IE, Astudillo V, Malirat V, Neitzert E, 1998: Serodiagnostic strategy for estimation of foot-and-mouth disease viral activity through highly sensitive immunoassays using bioengineered nonstructural proteins. Vet Q 20: 6-9.
• Bergmann IE, de Mello PA, Neitzert E, Beck E, Gomes I, 1993: Diagnosis of persistent aphthovirus infection and its differentiation from vaccination response in cattle by use of enzyme-linked immunoelectrotransfer blot analysis with bioengineered nonstructural viral antigens. Am J Vet Res 54: 825-831.
• Bergmann IE, Malirat V, Neitzert E, 2005: Non-capsid proteins to identify foot-and-mouth disease viral circulation in cattle irrespective of vaccination. Biologicals 33: 235-239.
• Bergmann IE, Malirat V, Neitzert E, Beck E, Panizzutti N, Sánchez C, Falczuk A, 2000: Improvement of a serodiagnostic strategy for foot-and-mouth disease virus surveillance in cattle under systematic vaccination: a combined system of an indirect ELISA-3ABC with an enzyme-linked immunoelectrotransfer blot assay Arch Virol 145: 473–489.
• Bergmann IE, Neitzert E, Malirat V, Ortiz S, Colling A, Sanchez C, Correa Melo E, 2003: Rapid serological profiling by enzyme-linked immunosorbent assay and its use as an epidemiological indicator of foot-and-mouth disease viral activity. Arch Virol 148: 891-901.
• Bronsvoort BM, Toft N, Bergmann IE, Sørensen KJ, Anderson J, Malirat V, Tanya VN, Morgan KL, 2006: Evaluation of three 3ABC ELISAs for foot-and-mouth disease non-structural antibodies using latent class analysis. BMC Vet Res 2: 30.
• Clavijo A, Hole K, Mingyi L, Collignon B, 2006: Simultaneous detection of antibodies to foot-and-mouth disease non-structural proteins 3ABC, 3D, 3A and 3B by a multiplexed Luminex assay to differentiate infected from vaccinated cattle. Vaccine 24: 1693–1704.
• Clavijo A, Wright P, Kitching P, 2004: Developments in diagnostic techniques for differentiating infection from vaccination in foot-and-mouth disease. Vet J 167: 9-22.
• Davies G, 2002: Foot and mouth disease. Res Vet Sci 73: 195-199.
• Dekker A, Gijsen E, 1998: The possible use of native foot-and-mouth disease non-structural protein 3A in a serological screening test. Vet Q; 20: 27-28.
• Espinoza AM, Maradei E, Mattion N, Cadenazzi G, Maddonni G, Robiolo B, La Torre J, Bellinzoni R, Smitsaart E, 2004: Foot-and-mouth disease polyvalent oil vaccines inoculated repeatedly in cattle do not induce detectable antibodies to non-structural proteins when evaluated by various assays. Vaccine 23: 69-77.
• Fu Y, Cao I, Sun P, Bao H, Bai X, Li P, Li D, Lu Z, Liu Z, 2011: Development of a dot immunoblot method for differentiation of animals infected with foot-and-mouth disease virus from vaccinated animals using non-structural proteins expressed prokaryotically. J Virol Methods 171: 234-240.
• Greiner M, Pfeiffer D, Smith RD, 2000: Principles and practical application of the receiver-operating characteristic analysis for diagnostic tests. Prev Vet Med 45: 23-41.
• Kitching RP, 2002: Identification of foot and mouth disease virus carrier and subclinically infected animals and differentiation from vaccinated animals. Rev Sci Tech 21: 531-538.
• Kweon CH, Ko YJ, Kim WI, Lee SY, Nah JJ, Lee KN, Sohn HJ, Choi KS, Hyun BH, Kang SW, Joo YS, Lubroth J, 2003: Development of a foot-and-mouth disease NSP ELISA and its comparison with differential diagnostic methods. Vaccine 21: 1409-1014.
• Lee F, Ming-Hwa J, Der-Wei Y, 2006: Presence of antibodies to non-structural proteins of foot-and-mouth disease virus in repeatedly vaccinated cattle. Vet Microbiol 115: 14–20.
• Lewis SA, Morgan DO, Grubman MJ, 1991: Expression, Processing, and Assembly of Foot-and-Mouth Disease Virus Capsid Structures in Heterologous Systems: Induction of a Neutralizing Antibody Response in Guinea Pigs. J Virol 65: 6572-6580.
• Lubroth J, Grubman MJ, Burrage TG, Newman JF, Brown F, 1996: Absence of protein 2C from clarified foot-and-mouth disease virus vaccines provides the basis for distinguishing convalescent from vaccinated animals. Vaccine 14: 419-427.
• Mackay DK, 1998: Differentiating infection from vaccination in foot-and-mouth disease. Vet Q 20: 2-5.
• Orsel K, Bouma A, 2009: The effect of foot-and-mouth disease (FMD) vaccination on virus transmission and the significance for the field. Can Vet J 50: 1059-1063.
• Paton DJ, De Clercq K, Greiner M, Dekker A, Brocchi E, Bergmann I, Sammin D, Gubbins S, Parida S, 2006: Application of non-structural protein antibody tests in substantiating freedom from foot-and-mouth disease virus infection after emergency vaccination of cattle. Vaccine 24: 6503-6512.
• Perry BD, Rich KM, 2007: Poverty impacts of foot-and-mouth disease and the poverty reduction implications of its control. Vet Rec 160: 238-241.
• Shen F, Chen PD, Walfield AM, Ye J, House J, Brown F, Wang CY, 1999: Differentiation of convalescent animals from those vaccinated against foot-and- mouth disease by a peptide ELISA. Vaccine 17: 3039-3049.