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Identification and Characterization of Marekâs Disease Virus Serotype 1 Using Molecular Approaches Hartawan, Risza; Dharmayanti, NLPI
Indonesian Bulletin of Animal and Veterinary Sciences Vol 25, No 1 (2015)
Publisher : Indonesian Animal Sciences Society

Marekâs disease is an important disease in the commercial poultry farm and causes significant economical loss. The disease is characterized by syndrome of paralysis and neoplastic formation in various organs and tissues in the host. The etiological agent is Marekâs disease virus serotype 1 (MDV-1). Eventhough the outbreaks in the field are well controlled by vaccination, several cases in the vaccinated flocks indicating virus evolution into more pathogenic strains. Therefore, monitoring of the disease circumstance in the field is indispensable for guiding better policies in disease controlling program. This paper describes several molecular methods that have been developed for identification and characterization of MDV-1. The identification and characterization of newly found virus strain in the field can be done by in vivo challenge test which is a conventional method especially to determine pathogenecity. However, this method requires several stages with time consuming procedures. The development of alternative methods for identification and characterization of MDV-1 viruses has been conducted mainly using molecular biology approach. Several molecular methods give satisfying result and have been implemented in both laboratory and field condition. Key words: Marekâs disease virus serotype 1, identification, characterization, molecular

Characterisation of the H5 and N1 genes of an Indonesian highly pathogenic Avian Influenza virus isolate by sequencing of multiple clone approach Risza Hartawan; Karl Robinson; Timothy Mahony; Joanne Meers
Jurnal Ilmu Ternak dan Veteriner Vol 15, No 3 (2010): SEPTEMBER 2010
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Hemagglutinin and neuraminidase are the main antigenic determinants of highly pathogenic avian influenza (HPAI) virus. The features of these surface glycoproteins have been intensively studied at the molecular level. The objective of this research was to characterise the genes encoding these glycoproteins by sequencing of multiple clones. The H5 and N1 genes of isolate A/duck/Tangerang/Bbalitvet-ACIAR-TE11/2007 were each amplified in one or two fragments using reverse transcriptase-PCR (RT-PCR), and subsequently cloned into pGEM-T Easy TA cloning system. The sequencing result demonstrated high homology between respective clones but with several variations that were identified as single nucleotide polymorphisms (SNPs). A total of 1,707 base pair and 1,350 base pair of H5 and N1 genes respectively were successfully assembled from multiple clones containing the genes of interest. The features of both H5 and N1 genes from this isolate resemble the typical characteristics of Indonesian strains of H5N1 virus from sub-clade 2.1.3. Key Words: Avian Influenza, Characterization, Gene Cloning, Hemagglutinin, Neuraminidase

In vitro expression of native H5 and N1 genes of avian influenza virus by using Green Fluorescent Protein as reporter Risza Hartawan; K. Robinson; T. Mahony; J. Meer
Jurnal Ilmu Ternak dan Veteriner Vol 16, No 3 (2011): SEPTEMBER 2011
Publisher : Indonesian Center for Animal Research and Development (ICARD)

The hemagglutinin and neuraminidase are important immunogen of avian influenza virus that are suitable for recombinant experimentation. However, both genes have been experienced rapid mutation resulting in diverse variety of genotypes. Hence, gene expression in recombinant systems will be difficult to predict. The objective of the study was to examine expression level of H5 and N1 genes from a field isolate by cloning the genes into expression vector pEGFP-C1. Two clones respresenting fulllength of H5 and N1 gene in plasmid pEGFP-C1 were transfected into chicken embryo fibroblasts (CEF), rabbit kidney (RK13) and African green monkey kidney (VERO) cells using Lipofectamine ‘Plus’ reagent. The experiment showed level of gene expression in the VERO cell was higher than in the RK13 and CEF cells. Observations using fluorescent microscopy and Western blotting revealed that the N1 gene was expressed better in all cells compared to the H5 gene. Key Words: H5N1 Virus, Hemagglutinin, Neuraminidase, Gene Expression, Green Fluorescent Protein

Development of Recombinant Vaccine Using Herpesvirus of Turkey (Hvt) as Vector for Several Viral Diseases in Poultry Industry Risza Hartawan
WARTAZOA, Indonesian Bulletin of Animal and Veterinary Sciences Vol 21, No 1 (2011): MARCH 2011
Publisher : Indonesian Center for Animal Research and Development

Herpesvirus of turkey (HVT) has been utilised as live vaccine against Marek’s disease in poultry industry world-wide for many years. However, the potency of HVT is not limited on the Marek’s disease only. Along with rapid development of recombinant technique, the potency of HVT can be broaden for other diseases. As naturally apathogenic virus, HVT is a suitable candidate as vector vaccine to express important antigens of viral pathogens. Several researches have been dedicated to design HVT recombinant vaccine by inserting gene of important virus, such as Marek’s disease virus (MDV), immuno bursal disease virus (IBDV), Newcastle disease virus (NDV) and Avian Influenza virus (AIV). Therefore, the future recombinant of HVT has been expected to be better in performance along with the improvement of recombinant technique. Key words: Herpesvirus of turkey, live vector vaccine, viral pathogens

Identification and Characterization of Marek’s Disease Virus Serotype 1 Using Molecular Approaches Risza Hartawan; NLPI Dharmayanti
WARTAZOA, Indonesian Bulletin of Animal and Veterinary Sciences Vol 25, No 1 (2015): MARCH 2015
Publisher : Indonesian Center for Animal Research and Development

Marek’s disease is an important disease in the commercial poultry farm and causes significant economical loss. The disease is characterized by syndrome of paralysis and neoplastic formation in various organs and tissues in the host. The etiological agent is Marek’s disease virus serotype 1 (MDV-1). Eventhough the outbreaks in the field are well controlled by vaccination, several cases in the vaccinated flocks indicating virus evolution into more pathogenic strains. Therefore, monitoring of the disease circumstance in the field is indispensable for guiding better policies in disease controlling program. This paper describes several molecular methods that have been developed for identification and characterization of MDV-1. The identification and characterization of newly found virus strain in the field can be done by in vivo challenge test which is a conventional method especially to determine pathogenecity. However, this method requires several stages with time consuming procedures. The development of alternative methods for identification and characterization of MDV-1 viruses has been conducted mainly using molecular biology approach. Several molecular methods give satisfying result and have been implemented in both laboratory and field condition. Key words: Marek’s disease virus serotype 1, identification, characterization, molecular

Pengembangan Sejumlah Primer untuk Reverse Transcriptase Polymerase Chain Reaction Guna Melacak Virus Flu Burung di Indonesia (DEVELOPMENt OF PRIMERS FOR REVERSE TRANSCRIPTASE POLYMERASE CHAIN REACTION TO DETECT AVIAN INFLUENZA VIRUS IN INDONESIA) Ni Luh Putu Indi Dharmayanti; Risza Hartawan; Dyah Ayu Hewajuli
Jurnal Veteriner Vol 17 No 2 (2016)
Publisher : Faculty of Veterinary Medicine, Udayana University and Published in collaboration with the Indonesia Veterinarian Association

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Until recently, two clades of of avian influenza viruses (AIVs) designated as 2.3.2 and 2.2.3 havebeen circulating in Indonesia. Mutations of AIV genes have cretaed many more variants of the virus. It istherefore important to evaluate the appropriate methods used for the detection and diagnosis of AI virusin the field. Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) have been used as a standardmethod for detection of AIV in many laboratories in Indonesia. The success of RT-PCR for detection ofAIV virus is dependent on the nucleotide sequences of primer that match with the circulating of AIVs. Theaims of this study was to develop RT-PCR by designing primers for H5 subtype specific to the circulatingAIVs in the field. The primers were designed using Primer Design software, and optimization andvalidation of the primer were conducted using AIVs that have been characterized in the previous study.The primers were then used RT-PCR using AIV isolates from field samples and their sensitivity andspecificity were then determined. The results showed that the H5 primers designed in this study, H5-IDand H5-NLP, was able to detect the AIVs in field samples better than the H5-specific primers have beenused previously. In conclusion, H5 primers designed based on recent viruses in the field showed betterresults in the detection of AI virus as compared to the previous primers. As AIV-H5N1 subtype in the fieldwill continue to change and evolve, the use of primers designed in this study is recommended for diagnosisof H5 AIV.

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