Serotypic identification of local Bluetongue virus isolates using Plaque Reduction Neutralization (PRN) and Reverse Transcriptase Polymerase Chain Reaction (RTPCR) Techniques

Abstract

Bluetongue virus (BTV) is a vector-borne disease of ruminants disseminated in the tropics and sub-tropics. It is also an important problem in the Middle East. The purpose of this study is serotypic identification of local Bluetongue viruses (BTV) isolated between 1998 and 2005. For this purpose, generic (g) and serotype specific (s) RT-PCR systems and the Plaque Reduction Neutralization Assay (PRNA) were used. Generic and serotype specific RT-PCR was performed on the segment 10 and segment 2 levels, respectively. Generic RT-PCR applications revealed specific DNA product (822 bp in length) in all 26 BTV field isolates. On the other hand, 19 BTV isolates were identified as serotype 9 and one isolate was found to be serotype 16, using the sRT-PCR technique. No DNA amplification was observed as a result of sRT-PCR for serotypes 2 and 4. Plaque reduction and neutralization assay (PRNA) was used for serologic identification of BTV isolates. Hyperimmune serums specific to three serotypes (BTV-4, BTV9 and BTV-16) produced in rabbits, were used in PRNA. The test showed that 19 of the isolates were BTV-9, and the remaining two isolates were identified as BTV-4 and BTV-16

Keywords

• Bluetongue virus
• serotyping identification
• PRNA
• RT-PCR

Yerel Mavidil virusu izolatlarının Plak Redüksiyon Nötralizasyon (PRN) ve Reverz TranskriptazPolimeraz Zincir Reaksiyonu (RT-PZR) teknikleri ile serotipik identifikasyonu

Abstract

Mavidil virusu dünyanın subtropik ve tropik bölgelerinde yaygın olan, ruminantların vektörle bulaşan hastalığıdır. Hastalık özellikle Ortadoğu için önem taşımaktadır. Bu çalısmada 1998-2005 yılları arasında izole edilen yerel BT viruslarının serotipik identifikasyonu amaçlanmıştır. Bu amaç için jenerik ve serotip spesifik RT-PZR sistemleri ile PRN testi kullanıldı. Jenerik RT-PZR ve spesifik RT-PZR sırasıyla segment 10 ve segment 2 düzeyinde gerçekleştirildi. Jenerik RT-PCR uygulamalarında 26 adet BTV izolatının tamamında spesifik DNA ürünü (822 bp. boyunda) elde edildi. Diğer taraftan sRT-PZR uygulaması sonunda ise izolatlardan 19 adedi serotip 9 ve 1 adedi serotip 16 olarak identifiye edildi. Serotip 2 ve 4 için uygulanan spesifik RT-PZR sonunda herhangi bir DNA ürünü elde edilemedi. Plak redüksiyon ve nötralizasyon testi BTV izolatlarının serotipik identifikasyonu için kullanıldı. Tavşanlarda üretilen BTV-4, BTV-9 ve BTV-16’ya spesifik hiperimmun serumlar PRN testinde kullanıldı. PRN testi sonunda 19 izolat BTV-9, 1’er izolat BTV-4 ve BTV-16 olarak identifiye edildi

Keywords

• Mavidil virusu
• serotipik identifikasyon
• PRNT
• RT-PZR

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