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Diagnosis and Molecular Marker Analysis of Bali’s Rabies Virus Isolates

Prof. Dr. Drh. I Gusti Ngurah Kade Mahardika, I GUSTI NGURAH KADE MAHARDIKA (2014) Diagnosis and Molecular Marker Analysis of Bali’s Rabies Virus Isolates. Jurnal Veteriner, 15. ISSN 1411 - 8327

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Abstract

The direct fluorescent antibody test (dFAT) was recommended by both World Health Organization (WHO) and Office International des Epizooties (OIE) as a standard diagnostic technique for rabies. Since the outbreak of rabies in Bali, it was ascertain the importance to develop a reverse transcriptase-polymerase chain reaction (RT-PCR) technique with specific primers as an alternative diagnostic method. The aim of this study was to develop a RT-PCR technique for rabies diagnosis in animals and find out the molecular marker of Bali’s rabies virus (BRV) isolates based on the sequence of nucleoprotein (N) gene. Brain samples were obtained during 2009 from 14 suspected rabid dogs and one cattle, where rabies viruses were isolated. The dFAT was used to detect the presence of rabies viral antigen. Ribonucleic acid (RNA) of rabies viruses was extracted with TRIzol reagent. Fragment of N gene was amplified using one-step RTPCR method with specifically-designed primer pairs and sequenced using ABI automatic sequencer. Multiple alignment of nucleotide and deduced amino acid sequences were analyzed using ClustalW of MEGA 4.0 program. This study found that twelve out of fifteen animal brain samples confirmed as rabies by dFAT. Similarly, a single band of 1215 bp PCR product for rabies virus was also detected in twelve out of twelve (100%) dFAT rabies positive samples. It is therefore evident that alternative diagnostic of rabies in animals can be established using RT-PCR technique. The results showed that the RT-PCR has a very high agreement with dFAT. Polymorphic sites of N gene of twelve BRV isolates were identified at the position 186, 501, 801, 840, 1068 and 1153. Bali’s rabies virus isolates have conserved amino acid (isoleucine) alterations at position 308 (open reading frame). Isoleucine distinguished between all Bali’s isolates and the all of isolates from other area of Indonesia and other part of the world. This finding significantly different as compared to other rabies virus isolates from other part of Indonesia or the world documented on the GenBank. Accordingly it is proposed that it can be used as molecular marker and believed to be the first study of molecular marker of rabies virus in Indonesia. Keywords : rabies virus, diagnosis, molecular marker, nucleoprotein

Item Type: Article
Uncontrolled Keywords: -
Subjects: L Education > L Education (General)
Divisions: Faculty of Law, Arts and Social Sciences > School of Education
Depositing User: Mr. Repository Admin
Date Deposited: 07 Jun 2016 21:58
Last Modified: 21 Jun 2016 05:58
URI: http://erepo.unud.ac.id/id/eprint/156

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