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I Wayan T. Wibawan
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Agriculture, Biological Sciences & Forestry Biochemistry, Genetics & Molecular Biology Veterinary

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Development of a nested PCR for detection of Bovine herpesvirus-1 (BHV-1) in bovine nasal secretion and semen Saepulloh, Muharam; Adjid, R.M Abdul; T. Wibawan, I Wayan; ., Darminto
Indonesian Journal of Animal and Veterinary Sciences Vol 13, No 2 (2008)
Publisher : Indonesian Animal Sciences Society

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (283.13 KB) | DOI: 10.14334/jitv.v13i2.609

Abstract

A nested polymerase chain reaction (nPCR) assay for detection of Bovine herpesvirus-1 (BHV-1) in bovine semen and nasal secretions was successfully developed. The nested Polymerase Chain Reaction was based on external and internal primers from the viral gD glycoprotein gene. This nPCR assay was 1000-fold more sensitive than using PCR external primer. The nested PCR has a detection limit as low as 5 ag/ml pure BHV-1 DNA and 100,75 TCID50/500 mL BHV-1 infected cells. On the other hand,  PCR using external primer had detection limit of about 5 fg/ml pure BHV-1 DNA. Specificity studies showed that nPCR could only detect BHV-1, whereas BHV-4, PRV, PI-3 and BRSV can not be detected. In addition, nPCR was also capable in detecting BHV-1 in nasal secretion samples from animal without clinical signs. A total of 405 samples consisted of 381 nasal secretion and 24 fresh semen samples have been tested with the nPCR. The result revealed that from 381 nasal secretion samples tested, 14 samples showed to be positive (3.68%), consisting of 13 out of 294 (4.42%) nasal secretion samples collected from Pangalengan West Java, and 1 out of 87 (1.54%) samples collected from Bogor. Furthermore, 2 out of 11 (18.18%) extended semen samples collected from Bogor and 2 out of 13 (15.38%) fresh semen samples collected from Pasuruan also showed to be positive. In addition, the nPCR was faster and easier to perform than the standard viral isolation test. It is concluded that, the nPCR can be used as test of choice for routine diagnosis of BHV-1. Key Words: Nested PCR, BHV-1, Semen, Glycoprotein D Gene, TCID50
Streptococcus agalactiae isolates FROM SUBCLINICAL MASTITIS CATTLE : I. In vitto phenotypic expression of isolates Iwan H. Utama; Ninuk S. Rejeki; I Made Sukada; I Wayan T. Wibawan
Media Veteriner Vol. 4 No. 1 (1997): Media Veteriner
Publisher : Media Veteriner

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (207.134 KB)

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Thirty six isolates Streptococcus agalactiae from subclinical mastitis cattle in Bogor were esanlincd their phenotypic espressions such as hemolytic pattern. supernatant turbidity inliquid medium, and colony morphology in semisolid medium (agar semi solid). Nine, fifteen and ttvelve isolates showed their a, 0, and y hemolytic patterns respectively. Fourteen isolates showed turbid supernatant, 18 isolates with less turbid supernatant, and the rest gave clear supernatant in fluid medium. In agar semi solid, 15 isolates showed mostly thick diffuse colonies in combination with thin diffuse and compact colonies, 17 isolates with mostly thin diffuse colonies in combination with thick diffuse and compact colonies. The rest isolates showed compact without diffuse colonies. There was a relation between growth pattern in fluid medium and colony morphology in agar semi solid, and the varia
Pemurnian Ekstraseluler Hyaluronidase Streptococcus agalactiae (Streptokokus Grup B ) (Extracellular Hyaluronidase Purification of Streptococcus agalactiae (Group B of Streptococus) Wendry Setiyadi Putranto; Sri Budiarti; Maggy T. Suhartono; I Wayan T. Wibawan; Zainatul Hayati
Jurnal Ilmu Ternak Vol 6, No 1 (2006)
Publisher : Fakultas Peternakan, Universitas Padjadjaran

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.24198/jit.v6i1.2260

Abstract

Hyaluronidase (EC 4.2.2.1) merupakan enzim ekstraseluler yang dihasilkan Streptococcus agalactiae (Streptokokus Grup B). Penelitian ini bertujuan untuk mengetahui aktivitas spesifik dari hyaluronidase S. agalactiae dan berat molekul proteinnya. Pemurnian enzim dengan sentrifugasi, pengendapan amonium sulfat 45% dan kromatographi kolom dengan Sephadex G-100 dan Sodiumdodecyl sulfate polyacrylainide gel electrophoresis (SDS PAGE). Enzim hyaluronidase dari S. Agalactiae memiliki aktivitas spesifik sebesar 0,0012 U/mg (ekstrak kasar), 0,0125 U/mg (pengendapan dengan 45% ammonium sulfate) and 0,032 U/mg (Gel Filtration). Berat molekul protein hyaluronidase adalah 102 - 106 kD.Kata kunci : hyaluronidase, aktivitas spesifik, pemurnian
Development of a nested PCR for detection of Bovine herpesvirus-1 (BHV-1) in bovine nasal secretion and semen Muharam Saepulloh; R.M Abdul Adjid; I Wayan T. Wibawan; Darminto .
Jurnal Ilmu Ternak dan Veteriner Vol 13, No 2 (2008): JUNE 2008
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (283.13 KB) | DOI: 10.14334/jitv.v13i2.609

Abstract

A nested polymerase chain reaction (nPCR) assay for detection of Bovine herpesvirus-1 (BHV-1) in bovine semen and nasal secretions was successfully developed. The nested Polymerase Chain Reaction was based on external and internal primers from the viral gD glycoprotein gene. This nPCR assay was 1000-fold more sensitive than using PCR external primer. The nested PCR has a detection limit as low as 5 ag/ml pure BHV-1 DNA and 100,75 TCID50/500 mL BHV-1 infected cells. On the other hand,  PCR using external primer had detection limit of about 5 fg/ml pure BHV-1 DNA. Specificity studies showed that nPCR could only detect BHV-1, whereas BHV-4, PRV, PI-3 and BRSV can not be detected. In addition, nPCR was also capable in detecting BHV-1 in nasal secretion samples from animal without clinical signs. A total of 405 samples consisted of 381 nasal secretion and 24 fresh semen samples have been tested with the nPCR. The result revealed that from 381 nasal secretion samples tested, 14 samples showed to be positive (3.68%), consisting of 13 out of 294 (4.42%) nasal secretion samples collected from Pangalengan West Java, and 1 out of 87 (1.54%) samples collected from Bogor. Furthermore, 2 out of 11 (18.18%) extended semen samples collected from Bogor and 2 out of 13 (15.38%) fresh semen samples collected from Pasuruan also showed to be positive. In addition, the nPCR was faster and easier to perform than the standard viral isolation test. It is concluded that, the nPCR can be used as test of choice for routine diagnosis of BHV-1. Key Words: Nested PCR, BHV-1, Semen, Glycoprotein D Gene, TCID50
Co-Authors Darminto . Darminto . I Made Sukada Iwan H. Utama MAGGY T. SUHARTONO Muharam Saepulloh Muharam Saepulloh Ninuk S. Rejeki R.M Abdul Adjid Sri Budiarti Poerwanto Wendry Setiyadi Putranto Zainatul Hayati