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Iron Uptake by Escherichia coli Cultured with Antibodies from Cows Immunized with High-Affinity Ferric Receptors^*

Department of Animal Sciences, The Ohio State University, Ohio Agricultural Research and Development Center, Wooster 44691

Corresponding author: J. S. Hogan; e-mail: hogan.4{at}osu.edu.

The synergistic effects of immunoglobulin G (IgG) from cows vaccinated with ferric citrate receptor (FecA) and IgG from cows vaccinated with ferric enterobactin receptor (FepA) were measured in an in vitro iron uptake assay. Serum was isolated and pooled within treatment from five cows each vaccinated with FepA or FecA or not vaccinated. Immunoglobulin G was isolated by ammonium sulfate precipitation and protein G affinity chromatography. Six Escherichia coli isolates from bovine intramammary infections were cultured in an iron-depleted medium to induce high-affinity iron acquisition systems and, in iron-depleted conditions, to specifically induce the expression of FecA. The bacterial cells were mixed with either 3 or 6 mg/mL of purified IgG and ⁵⁵Fe. The radioactivity of ⁵⁵Fe taken up by the bacterial cells was measured by a liquid scintillation counter after 5-, 10-, and 15-min incubations at 37°C. The combination of anti-FecA IgG and anti-FepA IgG reduced ⁵⁵Fe uptake compared with either anti-FecA or anti-FepA alone. Iron uptake was reduced more by anti-FecA IgG than by anti-FepA IgG when the ferric citrate system was induced. Reduction of iron uptake did not differ between anti-FepA alone and anti-FecA alone when citrate was absent from the medium.

Key Words: Escherichia coli • FecA • FepA • immunoglobulin G

Abbreviation key: FecA = ferric citrate receptor, FepA = ferric enterobactin receptor, IROMP = iron-regulated outer membrane proteins