Abstract
Enzootic Bovine Leucosis (EBL) is one of viral diseases in cattle caused by bovine leukemia virus (BLV), from Retroviridae. The virus can be detected using severals methods such as Polymerase Chain Reaction (PCR), while antibody can be detected using Agar Gel Immunodifussion (AGID). The aim of this experiment was to study the effectivity of PCR and AGID methods to detect enzootic bovine leukosis virus in Indonesia. Samples of peripheral blood leukocyte (PBL) were collected from cattles those with and without showing clinical signs. A total of 307 blood and serum samples were tested against BLV using PCR and AGID tests, while 21 semen samples which were from similar animals for blood collection were collected only for PCR test. The results indicated that twelve cattles have positive results with PCR test in PBL, but from those cattles only seven were positive with AGID. On the other hand, the PCR did not detect EBL in 21 bovine semen samples tested, although one sample gave positive result with PCR in PBL. This results indicated that PCR method from blood samples was more sensitive than that AGID method. The PCR detection was also more sensitive for PBL than that for semen samples.
Highlights
Enzootic Bovine Leucosis (EBL) is one of viral diseases in cattle caused by bovine leukemia virus (BLV), from Retroviridae
The virus can be detected using severals methods such as Polymerase Chain Reaction (PCR), while antibody can be detected using Agar Gel Immunodifussion (AGID). The aim of this experiment was to study the effectivity of PCR and AGID methods to detect enzootic bovine leukosis virus in Indonesia
A total of 307 blood and serum samples were tested against BLV using PCR and AGID tests, while 21 semen samples which were from similar animals for blood collection were collected only for PCR test
Summary
Sebanyak 328 sampel yang terdiri dari 166 darah sapi (Limousin, Simmental, Angus, FH dan PO) asal peternakan sapi di Bogor; 30 sampel darah sapi dan 21 sampel semen sapi (Limousin, Simental, Angus, FH dan Ongole) asal peternakan sapi di Lembang; dan 111 sampel darah sapi Brahman asal peternakan sapi di Palembang. Buffy coat dipisahkan dengan cara sentrifugasi pada kecepatan 1.500 rpm selama 15 menit dan digunakan untuk ekstraksi DNA, sedangkan serum digunakan untuk uji serologik dengan menggunakan AGID. Perlakuan terhadap sampel semen untuk analisis PCR dilakukan berdasarkan Mohammadabadi et al (2011) sebagai berikut: (i) untuk menghindari faktor inhibitor yang terdapat pada semen maka semen tersebut dicampur dengan DEPC-dH2O dengan perbandingan sama banyak, (ii) kemudian campuran semen-DEPC-dH2O tersebut di beku cairkan sebanyak 3 kali dan (iii) akhirnya 300 μl digunakan untuk ekstraksi RNA. Uji serologik dilakukan dengan menggunakan kit AGID yang berasal dari Synbiotics Corporation (Prancis). Agar yang telah diisi dengan antigen, antiserum, dan serum lapangan ini kemudian dimasukkan ke dalam kotak plastik yang diberi alas kertas saring yang dibasahi dengan air, untuk menjaga agar keadaannya selalu lembab dan disimpan pada suhu kamar, serta diamati setiap hari sampai 3 hari. Reaksi serologik dinyatakan positif apabila dalam waktu 24-72 jam terbentuk garis presipitasi antara lubang yang berisi antigen dan lubang yang berisi serum lapangan, dan garis presipitasi ini bersambung dengan garis presipitasi yang terbentuk antara lubang yang berisi antigen dan lubang yang berisi antiserum
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