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Genetic Analysis of GBS Adherence to Fibronectin

Investigators
Tamura, Glen
Institutions
Children's Hospital and Regional Medical Center - Seattle
Start date
1998
End date
2003
Objective
  1. To study the role of GBS adherence to fibronectin in the pathogenesis of neonatal pneumonia and sepsis.
  2. To clone the transposon insertion sites of the additional two mutants, and to create targeted mutations in genes of interest as well as complement mutants to confirm the role of the transposon insertion in the phenotype.
  3. To clone the structural gene for the fibronectin adhesin.
    If necessary, create additional fibronectin adherence mutants.
More information

GBS adherence to epithelial cells may play an important role in pathogenesis of GBS pneumonia. I propose that GBS adherence to epithelial cells is mediated, at least in part, by GBS adherence to fibronectin.

I have isolated four transposon mutants with single insertions which show decreased adherence to fibronectin; 3/4 of these show decreased adherence to epithelial cells. One mutant has been tested in a neonatal rat model of pulmonary colonization, and shows a significantly decreased ability to persist in the lungs of neonatal rats at 24 hours.

I have cloned the transposon insertion sites from two of these mutants. One mutant contains a mutation in a gene which shows significant homology to ABC transporters, while the other contains a mutation immediately upstream of a gene with homology to multiple surface receptors. Disruption of the ABC transport gene by plasmid insertional mutagenesis demonstrates that this gene is required for maximal fibronectin adherence.

The role of these genes will be further studied in the mouse model of neonatal pulmonary colonization. These studies will establish a role for fibronectin adherence in the pathogenesis of GBS disease. The role of these putative adhesin genes will then be tested after hyperexpression of the adhesin gene by blockade of binding of GBS to fibronectin with both the cloned adhesin and antisera to the adhesin. These studies will allow for identification of the structural gene(s) for fibronectin adhesin(s).

Funding Source
Nat'l. Inst. of Allergy and Infectious Diseases
Project number
5R29AI041484-03
Categories
Prevention and Control
Bacterial Pathogens
Sanitation and Quality Standards