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Glycoprotein Micelles for Live Agent Collection and Stabilization

Investigators
Shelton, Daniel
Institutions
USDA - Agricultural Research Service
Start date
2008
End date
2009
Objective
  1. Select fimbrial type(s) for investigation.
  2. Produce antibodies against target fimbriae.
  3. Conduct immunoassay studies elucidating fimbrial expression as a function of environmental parameters.
  4. Screen fimbria against glycan microarray(s) to optimize affinity.
  5. Conduct attachment studies with micelles to establish optimal binding kinetics and survival.
More information
Approach: Type I fimbriae are relatively ubiquitous among Salmonella and E. coli. They were originally described based on the observation that they agglutinated erythrocytes, and that agglutination was inhibited by mannose. Subsequent research has indicated that there are several antigenically distinct sub-types, encoded by separate operons. A number of investigators have produced antibodies (including polyclonal and monoclonal) against various fimbriae. We will obtain antibodies from other researchers to the extent that they are made available. However, we also propose to produce our own antibodies. Fimbriae will be harvested from selected strains and polyclonal antibodies produced. Various Salmonella and pathogenic E. coli will be screened with the antibodies developed in objective 2 to identify strains expressing the target fimbriae. Strain(s) selected for objective 1 will be used as control strains. Isolates from our in-house culture collections of Salmonella and pathogenic E. coli will be screened for comparative purposes and to assess the prevalence of the target fimbriae. Selected strains will be assayed for fimbrial expression under a variety of growth and environmental conditions, including: growth phase, temperature, dissolved oxygen (aerobic vs. anaerobic), and media composition (e.g., C:N:P ratios). Selected Salmonella and/or pathogenic E. coli expressing the target fimbriae will be screened against available glycan arrays. Lipid micelles of various sizes will be prepared coated with the appropriate glycoprotein(s). Time-course studies will be conducted with different cell concentrations and cell/micelle ratios to quantify and optimize the kinetics of attachment. In addition, survival of captured cells will be assessed as a function of time and temperature.
Funding Source
Agricultural Research Service
Project number
1265-32420-004-06T
Accession number
414331
Categories
Escherichia coli
Bacterial Pathogens
Salmonella