Determination of RNA genome in the low titer zoonotic RNA virus samples

Abstract

There are numerous RNA virus infections in mammals as emerging zoonosis originating from wildlife. The total viral diversity is unknown. The total number of mammalian viruses is estimated to be more than 320000. Many of these viruses have genomic RNA and are important as zoonotic agents. In many RNA virus infections, the virus load in the serum can be used as a marker for severity. For diagnostic purposes dealing with the low virus titer in serum and biotechnological applications, it is mandatory and highly important, to have a native, innovative, and sensitive RNA isolation kit. For this aim, we used an RNA isolation kit with RNA carrier, produced by Research Institute Molecular Biological System Transfer (MBST, Tehran, Iran) and the Avian infectious bronchitis virus vaccine of Nobilis, 2500 Doses, 103 ID50 (Intervet, Netherlands) as virus RNA probe. The RNA was extracted from diluted solutions (up to 10-9 ). Subsequently, cDNA was synthesized. The cDNA was then amplified using a specific primer pair derived from the RNA genome of the Avian infectious bronchitis virus. Our results showed that it was possible to detect RNA viruses in the prepared samples with virus titers of up to 10-9 or 0.00001 × ID50. These results were confirmed by the Iranian National Reference Laboratory, Diagnosis and Applied Studies Center, and Veterinary Organization using their evaluation matrix Avian influenza virus sample. In conclusion, this kit is suitable for samples with low RNA virus titers.

Keywords

• rt-pcr
• carrier rna
• avian infectious bronchitis virus
• avian influenza virus
• rna virus