Recipient Organization
TessArae LLC
46090 Lake Center Plaza, Suite 304
Potomac Falls,VA 20165
Performing Department
(N/A)
Non Technical Summary
National swine health statistics indicate a growing death rate due to respiratory disease in both the nursery and grower/finished phases in swine (Swine 2006, APHIS, USDA). In 2006, veterinary diagnostic testing revealed that Porcine Reproductive and Respiratory Syndrome was the most prevalent of the diagnosed diseases in breeding herd and nursery pigs (Swine 2006, APHIS, USDA). Additional diseases that contribute to the swine morbidity and mortality rate include Porcine Circovirus 2 associated diseases, swine influenza, foot and mouth disease, classic swine fever and swine vesicular disease. The proposed TessArray RPM assay is a simultaneously differential diagnosis platform for these and other targeted pathogens of the assay. The single test result can establish cause of infectious disease in individual animals as well as outbreaks of infectious disease in local herds or across swine production communities. No existing test other than RPM can enable rapid differential diagnosis, determine presence of multiple infectious agents co-infecting individual animals, or support epidemiological tracking in epidemic outbreaks to minimize effective response times.
Animal Health Component
100%
Research Effort Categories
Basic
5%
Applied
55%
Developmental
40%
Goals / Objectives
We will design and develop a highly multiplexed gene sequencing based assay for the detection and identification of viral and bacterial pathogens causing agriculturally and economically significant disease of swine. This approach, using a high-density gene sequencing microarray offers significant advantages over conventional single-target, biomarker-based (e.g. antibody or PCR) testing. In addition to maximum sensitivity for detection of one or more multiple pathogens that may be present in a single specimen, the assay result represents one or more specimen-specific gene sequences, rather than a measured signal for a presence/absence report of only one targeted pathogen. The reported pathogen gene sequences enable forensic and epidemiological tracking of the same detected pathogens from one specimen to another, which no biomarker-based assay can support. Furthermore, the microarray assay is capable of detection and identification, through gene sequences, of emergent pathogen strains and variants that are at least 80% sequence-similar. Biomarkers are so constrained to perfect sequence matching of assay probes and intended target pathogens that natural variants often result in false negative or false positive assay results. TessArae will collaborate with the Kansas State University College of Veterinary Medicine for the selection of simultaneously targeted pathogens, to include at least 15 bacterial genera and at least 18 categories of virus.
Project Methods
TessArae and collaborators at KSU-CVM will determine the multiple viral and bacterial pathogens of swine to be simultaneously targeted by the resequencing pathogen microarray (RPM). TessArae then undertakes a systematic survey of all natural variants of these pathogens that have been characterized by one or more partial or complete gene or genome sequences. Representative target gene sequences are selected from two or three different target genes per target pathogen to be used as detector tiles on the RPM. Part of the selection of these detector tiles is consideration of favorable flanking sequences from their respective genomes to support multiplexed PCR-like amplification in the RPM assay protocol. The target gene sequences that can be interrogated on the smallest RPM configuration total about 47 Kbp. The approximately 35 different bacterial and viral pathogens would thus be represented by at least two different genes each up to an average sequence length of about 670bp per target gene. TessArae will receive purified nucleic acid preparations from KSU archive specimens for testing the first batch of arrays (under USDA permit 105229). Once initial validations and quality assurance of the arrays and assay protocols are completed, the final Phase I SBIR report will be submitted, and remaining assay kits will be sent to KSU for further testing in anticipation of later Phase II activities.