Volume 17 | Number 3 | Fall 2005 | Safety Issue

GTRI Receives USDA Grant to Develop Interferometric Biosensor as a Tool for the Detection of Avian Influenza

David Gottfried, senior research scientist, is spearheading a study to determine the feasibility of using GTRI’s interferometric biosensor for AI detection. Photo by Gary W. Meek

Researchers with the Georgia Tech Research Institute (GTRI) recently received a grant from the United States Department of Agriculture (USDA) to develop biosensor technology for avian influenza detection.

In 2004, GTRI was part of a proposal submission to the USDA’s Cooperative State Research Education and Extension Service, National Research Initiative, Cooperative Agricultural Project in Animal and Plant Security (USDA-CSREES-NRI-CAP). This project, a three-year, $5-million program titled “Prevention and Control of Avian Influenza in the U.S.,” organized by the University of Maryland and including multiple states and research centers, was funded in January 2005. The overall objective of the program is the prevention and control of avian influenza in the United States through research, education, and outreach. The research objectives for the sensor development component of the diagnostics subgroup include:

a. Development and optimization of assays using currently available polyclonal and monoclonal antibodies coupled to the GTRI waveguide sensor. In the first year of the project, assays will be developed for subtype H7 influenza to demonstrate proof of the concept and evaluate the sensitivity.

b. Testing of the assay protocol in the lab using split samples (oropharyngeal swabs) collected from experimentally infected chickens for comparison to other diagnostic methods.

c. Testing of the prototype in field applications. Methods for simple and rapid sample clean up will be evaluated as needed.

The team at GTRI, headed by Senior Research Scientist David Gottfried, working on biosensor development and assay optimization is collaborating with Dr. David Suarez (USDA Southeast Poultry Research Laboratory) for obtaining assay reagents, standard virus samples, split oropharyngeal swab samples, and testing. Additional funding for this program has been provided by the Georgia Research Alliance Innovation Fund.

Gottfried explains that early identification of avian influenza (AI) infection in poultry is critical for aiding in the control of outbreaks. Currently, three methods of direct identification of infected flocks can be used: virus isolation, real-time reverse-transcriptase polymerase chain reaction (RT-PCR), and commercially available antigen capture tests. Virus isolation is a sensitive technique, but typically requires five to seven days for testing. RT-PCR is becoming more commonly available in veterinary diagnostic laboratories, but requires expensive equipment and appropriate laboratory facilities. While the time to get test results can be in as little as three hours, routine surveillance samples are more often processed in 24 hours. The antigen capture immunoassays, such as the Directigen or Binax tests, can provide rapid test results, but suffer from low sensitivity and high cost.

“There still remains a need for a rapid and sensitive penside diagnostic test to identify flocks and birds infected with avian influenza. For this test to be practical, it must be cost-effective and not require expensive instrumentation. The availability of a rapid, sensitive, and economical diagnostic test for on-site analysis would greatly aid in the control of avian influenza at the outset and during an outbreak,” says Gottfried.

“A field-deployable biosensor will provide a low-cost device for assessment of avian influenza infection on the farm in poultry flocks and at live bird markets. The use of on-site assays will decrease the response time to respond to outbreaks. As an inexpensive presumptive test, this will allow for voluntary monitoring programs and flock containment. In addition, limited funds for testing can be allocated to confirmatory assays when required,” adds Gottfried.

Gottfried and his research team have researched, designed, developed, and tested a prototype of a cost-effective biosensor for the direct measurement of antibody-antigen complex formation as well as other biological interactions. Their optical interferometric biosensor has been used for detection of bacteria such as Salmonella typhimurium, Campylobacter jejuni, and Bacillus anthracis using polyclonal and monoclonal antibodies. Detection of concentrations as low as 103 cfu/ml can be achieved within 30-60 minutes, and no antibody labeling, reporter antibody, extended incubation, or washing steps are required. Because signal transduction is based on the direct binding of antigen to antibody, the effects of species that might interfere with nucleic acid or signal amplification (required by other methods) are minimized.

Initial work under the recently funded grant has begun. To date, explains Gottfried, assays were performed with two strains of the H7 influenza subtype and a control H8 subtype to demonstrate proof of the concept. Three antibodies (two monoclonal and one polyclonal) were evaluated for their sensitivity. Although much additional work remains, he says, the GTRI biosensor was more sensitive than the Binax Now Flu A assay by 2-3 orders of magnitude, and provided discrimination of the H antigen subtypes with a 60-minute assay.

“With continued development, this biosensor technology should be able to rival the virus isolation and PCR technologies in sensitivity with a more rapid and lower cost assay. The availability of this type of diagnostic test for on-site analysis will be helpful in the control of avian influenza,” says Gottfried.