Project Number: 6612-32000-064-13
Project Type: Reimbursable

Start Date: Oct 21, 2011
End Date: Mar 31, 2012

Objective:
1. To characterize Newcastle disease virus (NDV) and Avian Influeza virus isolates from East Europe. Waterfowl samples from Ukraine and Russia will be characterized using sequencing and phylogenetic analyses. Selected viral isolates will be used as a template to clone the F and HN genes into a eukaryotic expression vector (plasmid) and used as vaccines. 2. To complete three different overlapping training projects that will be provided to two students from ARRIAH in the 4 week visit to Southeast Poultry Research Laboratory.

Approach:
The approach for objective one of this work will be to determine the nucleotide sequences of Newcastle disease virus (NDV) isolates isolated in Ukraine between 2001 and 2010. Reverse transcription polymerase chain reaction (PCR), sequencing, and sequence based phylogenetic analysis will be conducted. Once the phylogenetic analysis is accomplished, a representative isolate will be selected and used as the source of RNA for the cloning of the F and HN genes. The genes will be expressed in bacteria and used as DNA vaccines. The approach for objective two includes four teaching modules with the following objectives: 1) training session to provide the methodology to sequence the non-coding region of influenza viruses. This will include hands RNA ligation techniques, real-time (RT)-PCR amplification, purification of sequencing product, and sequence the sequence; 2) An indepth review of the most important web based influenza bioinformatic sites will be explored to allow the students to understand how they can integrate these tools into their existing research for avian influenza; 3) Systematically evaluation of commonly used primers from the U.S. and Russia to evaluate how sequence variation can affect test results for both sensitivity and specificity. This includes the use of plasmids and primers specifically made to evaluate natural variation found in nature; and 4) Through a series of lectures and interactive exercises, a review of Biosafety and biosecurity performed at the Southeast Poultry Research Laboratory will be reviewed. The lectures will be overviews from the committee chairs that are assigned jurisdiction at the laboratory and include the Biosecurity Advisory Committee, the Institutional Biosafety committee, and the Institutional Animal Care and Use Committee.