Virulensi Virus Newcastle Disease Isolat Lapang Berdasarkan Analisis Bioinformatika Gen Protein Hemaglutinin - Neuraminidase

ABSTRACT Newcastle disease virus (NDV) is very contagious disease agent, and causes many outbreaks trough out the country of Indonesia. This study was conducted to determine the virulence of Bali field isolate of NDV based on haemaglutinin-neuraminidase (HN) protein genes. Four field isolates from different locations were propagated at fertilized chicken eggs of 9 days old. Allantoic fluid was harvested and NDV was confirmed using standard haemaglutination (HA) and haemaglutination inhibition test (HI) methods. The fragment of HN protein gene was amplified using RT-PCR. The product was sequenced using Big-Dye termination method. All four isolates grew well with the titer of 25-29 HA unit and could be confirmed using HI test. The HN genes, however, exhibited variations at its 3-D structure and hydrophobicity between the virus that previously circulated in Indonesia and vaccine virus. It is concluded that all four Bali’s isolates under this study are virulent VND of the genotype VII. Further testing is needed to justify the best formula of NDV vaccine to be used trough out Indonesia. Key words : Newcastle disease virus, genotype VII, HA, HI, RT-PCR, vaccine ABSTRAK Virus Newcastle disease (VND) merupakan agen penyakit yang sangat kontagius dan setiap tahun mewabah pada beberapa peternakan unggas di Indonesia. Tujuan penelitian ini adalah untuk menentukan virulensi VND isolat lapangan secara molekuler berdasarkan gen penyandi protein hemaglutinin-neuraminidase. Empat isolat VND yang berasal dari lokasi berbeda diperbanyak pada telur ayam bertunas (TAB) umur 9 hari melalui ruang allantois. Cairan alantois dipanen dan dilanjutkan dengan uji hemaglutinasi (HA) baku. Konfirmasi dilakukan dengan uji hambatan hemaglutinasi (HI) baku. Fragmen gen protein HN diamplifikasi dengan RT-PCR. Hasil RT-PCR divisualisasikan pada gel agarosa 1% dan disekuensing dengan Big-Dye termination method. Hasil uji HA keempat isolat VND memiliki titer kisaran 25-29 dan dapat dikonfirmasi dalam uji HI.Analisis sekuen asam amino protein HN menunjukkan variasi dalam struktur tiga dimensi dan hidrofobisitas antar VND yang pernah bersirkulasi di Indonesia dan virus vaksin. Dari hasil penelitian dapat disimpulkan bahwa 4 isolat VND adalah virulen dengan genotipe VII. Uji lanjutan perlu dilakukan untuk mendapatkan formulasi vaksin terbaik yang dapat digunakan di seluruh