2013 Annual Report
1a.Objectives (from AD-416):
To conduct computational evolutionary sequence analysis in the context of contemporary and historical SIV for determination of phylogenetic relationships, lineages, and reassortment events.
1b.Approach (from AD-416):
The USDA SIV Surveillance System was initiated in 2009 and roughly 150 SIV isolates have entered into the system. Currently, 3 gene segments (HA, NA, and M) are being sequenced routinely by participating NAHLN laboratories and the sequences submitted to the GenBank database. There has not been a systematic approach for analyzing and reporting summarized results of the sequencing efforts on a single gene or whole virus genome level. This is a gap in providing a useful output from the surveillance system for determination of significant virus evolution and identification of viruses of interest. Additionally, NVSL will conduct whole virus sequencing on approximately 400 viruses currently in the SIV repository using the Illumina-based approach developed at St. Jude Children’s Research Hospital and adapted at NADC. NADC will be instrumental in transferring this technique to NVSL for use at the ISU Sequencing and Synthesis core facility. Preliminary data generated from the Illumina sequencing run will be initially assessed and assembled at NADC with subsequent transfer of the bioinformatics methods developed by the NADC to APHIS-VS personnel. ARS will then perform a computational evolutionary biology analysis where the fully assembled sequences are analyzed in the context of contemporary and historical SIV for determination of phylogenetic relationships, lineages, and reassortment. To conduct large scale sequences analyses and subsequent quarterly analyses, a dedicated temporary scientist must be hired who has skills in sequence analysis as well as an understanding of influenza biology and ecology in the swine host. The results will be shared with APHIS-VS NVSL and NSU staff for reporting purposes as well as appropriate GenBank sequence deposition and peer-reviewed publication. The temporary scientist will interact and liaise with NVSL staff to transfer knowledge and skills as appropriate and with NADC scientific staff for associated in vitro and in vivo study of select SIV isolates.
Using data derived from the USDA swine influenza A (IAV) surveillance system, we constructed a baseline characterization of swine IAV genetic diversity in the United States. Phylogenetic analyses have revealed changes in the population dynamics among multiple clades of A/H1N1, A/H3N2, and A/H1N2 co-circulating in U.S. swine populations. The genetic differences are considerable, and we suggested a re-evaluation of phylogenetic nomenclature based upon objective genetic and statistical parameters. In addition, these data have provided criteria for selecting contemporary isolates for further antigenic characterization. In doing so, this work aided agricultural production and pandemic preparedness by allowing for the identification and a benchmark from which to measure success of intervention strategies. These interventions include timely vaccine and diagnostic updates for use in swine, as well as factors to prevent infection and transmission, such as changes in production practices or farm management.
Interaction and training of appropriate staff has been achieved via two mechanisms. The first was a joint National Institutes of Health-USDA workshop: this workshop covered introductory and advanced phylogenetic techniques, and allowed for practical experience and training with the appropriate analysis of sequence data. The second mechanism has been via meetings, consultation, and e-mails with staff in the National Veterinary Services Laboratories (NVSL). Outreach to the swine health community is ongoing, with conference presentations at the Alan Leman Swine Conference and American Association of Veterinary Laboratory Diagnosticians in the final quarter of 2013. A notable achievement is the collaboration with the Influenza Research Database, which will soon allow for rapid "typing" of sequence data via an online and user-friendly interface.
The sequencing effort of the USDA has resulted in an H1 dataset of nearly 750 unique isolates, and an H3 dataset of nearly 350 unique isolates (including publically available data). These data represent a considerable challenge to quality check, and analyze. In collaboration with NVSL staff we have begun to characterize the genetic diversity, search for reassortment events, and describe patterns in the spatial and temporal transmission of swine IAV in the U.S. Preliminary analyses were presented in May 2013, and manuscripts and sequence publication are in progress.