Deteksi Molekuler Gen Fusion (F) dan Analisis Perbandingan Beberapa Enzim Restriksi sebagai Penentu Patotipe Virus Newcastle Disease

Medania Purwaningrum; Verawati Verawati; Aris Haryanto

doi:10.22146/jntt.44935

Deteksi Molekuler Gen Fusion (F) dan Analisis Perbandingan Beberapa Enzim Restriksi sebagai Penentu Patotipe Virus Newcastle Disease

https://doi.org/10.22146/jntt.44935

Medania Purwaningrum^(1*), Verawati Verawati⁽²⁾, Aris Haryanto⁽³⁾

(1) Departemen Biokimia dan Biologi Molekuler, Fakultas Kedokteran Hewan, Universitas Gadjah Mada
(2) Laboratorium Bioteknologi, Balai Besar Veteriner Wates, Yogyakarta
(3) Departemen Biokimia dan Biologi Molekuler, Fakultas Kedokteran Hewan, Universitas Gadjah Mada
(*) Corresponding Author

Abstract

Newcastle disease (ND) is a contagious viral disease caused by Avian Paramyxovirus Serotype-1 (APMV-1). This viral infection is responsible for devastating outbreak by attacking nerve, respiration, and also digestive system. This disease often followed with decreasing of eggs production and also responsible for economic losses in the poultry industries around the globe. The main goal was to differentiate virulent or avirulent strain of ND virus from F gene, which is the virulent marker of ND virus, by Reverse Transcription Polymerase Chain Reaction (RT-PCR) and Restrion Enzyme Analysis using BamH1, Hin 1l, and Apa 1. Ten ND virus samples came from Animal Disease Investigation Center (ADIC) Wates virus collection, collected from field case in 2012-2013. Newcastle disease virus was collected by extraction from the samples. The RNA product of extraction were used as a template for amplification in RT-PCR. The target of RT-PCR amplification was F gene. The results indicated positive reaction due to existing of DNA fragment band in size of 767 bp. RT-PCR and Restrion Enzyme Analysis can be used as tool to determine the pathotype of ND virus showed different restriction visualized by gel agarose electrophoresis.

Keywords

Newcastle Disease Virus, F gene, RT-PCR, Restriction Enzyme, Pathotype

Full Text:

PDF

References

Adi, A. A. A. M., Astawa, N. M., Putra, K. S. A., Matsumoto, Y. (2008). Deteksi Virus Penyakit Tetelo Isolat Lapangan dengan Metode Nested Reverse Transcriptase Polymerase Chain Reaction. Jurnal Veteriner. 9, 3:1-7.

Aldous, E. W., Mynn, J. K., Irvine, R.M., Alexander, D.J., Brown, I.H. (2010). A Molecular Epidemiological Investigation of Avian Paramyxovirus Type 1 Viruses Isolated from Game Birds of the Order Galliformes. Avian Pathology 39, 6, 519-524.

Dortmans, J. C. F. M., Koch, G., Rottier, P. J. M., Peeters, B. P. H. (2009). Two Genetically Closely Related Pigeon Paramyxovirus Type 1 (PPMV-1) Variants with Identical Velogenic Fusion Protein Cleavage Sites but with Strongly Contrasting Virulence. J Gen Virol. 90, 2746-2750.

Dortmans, J. C. F. M., Rottier, P. J. M., Koch, G., Peeters, B. P. H. (2011). Passaging of a Newcastle Disease Virus Pigeon Variant in Chickens Results in Selection of Viruses with Mutations in the Polymerase Complex Enhancing Virus Replication and Virulence dalam Virulence Determinants of Newcastle Disease Virus, hal : 77-96.

Fatchiyah, Estri Laras, Sri Widyarti dan Sri Rahayu., (2011). Biologi Molekuler, Prinsip Dasar Analitis. Penerbit Erlangga. 46 – 49.

Lee, E-K., Jeon, W-J., Kwon, J-H., Yang, C-B., Choi, K-S. 2008. Molecular Epidemiological Investigation of Newcastle Disease Virus from Domestic Ducks in Korea. Veterinary Microbiology 134, 241–248.

Kim, L. M., Suarez, D. L., Afonso, C. L. 2008. Detection of A Broad Range of Class I and II Newcastle Disease Viruses Using A Multiplex Real-Time Reverse Transcription Polymerase Chain Reaction Assay. J Vet Diagn Invest. 20,414–425.

Mase, M. dan Kanehira, K. 2012. Differentiation of APMV-1 by PCR and REA. The Journal of Veterinary Medical Science.1-10. Miller, P. J., Decanini, E. L., Alfonso, C. L. 2010. Newcastle disease: Evolution of Genotypes and the Related Diagnostic Challenges. Infect. Gen. Evol. 10: 26-35.

Murphy, F. A., Gibbs, E. P. J., Horzinek, M. C., Studdert, M. J. 2008. Paramyxoviridae dalam Veterinary Virology. 3th edition. USA. 416-420.

Ngili, Y., 2010. Biokimia Dasar. Rekayasa Sains, Bandung. Office International des Epizooties (OIE). 2012. Newcastle Disease. Manual Standards for Diagnostic Test and Vaccine, 576-589.

Purwaningrum, M., Verawati, Aris H. 2018. In Proceeding of International Symposium in Veterinary Science. Pathotyping of Newcastle Disease Local Isolate Virus in Poultry by RTPCR RFLP. pp.57-62. Universitas Gadjah Mada, Yogyakarta.

Purwaningrum, M., 2014. Patotyping of Newcastle Disease Local Isolate Virus in Poultry By Reverse Transcription Polymerase Chain Reaction and Restriction Endonuclease Analysis Method (RT-PCR and REA). Theses. Faculty of Veterinary Medicine, Universitas Gadjah Mada, Indonesia.

Qin, Z. M., Sun, L., Ma, B. C., Cui, Z., Zhu,Y., Kitamura, Y., Liu, W. 2008. F gene Recombination between Genotype II and VII Newcastle Disease Virus. Virus Research. 131, 299-303.

Sianita, N., Hasan, Z., Kusriningrum R. 2011. Respon Antibodi dan Protektivitas pada Ayam Pasca Vaksinasi Menggunakan Vaksin Nd Aktif Lv12. Veterinaria Medika. 4, 2.

Tan, SW., Ideris, A., Omara, A. R., Yusoff , K., Hair-Bejoa, M. 2009. Detection and Differentiation of Velogenic and Lentogenic Newcastle Disease Viruses using SYBR Green I Real-Time PCR with Nucleocapsid Gene-specific Primers. Journal of Virological Methods 160, 149–156.

DOI: https://doi.org/10.22146/jntt.44935