Ornithobacterium rhinotracheale North American field isolates express a hemolysin-like protein.

Ornithobacterium rhinotracheale is a gram-negative bacterium responsible for the sporadic outbreaks of airsacculitis in poultry, accounting for millions of dollars in losses to the poultry industry annually. Although the organism was originally classified as non-beta-hemolytic, recent North American field isolates of O. rhinotracheale obtained from pneumonic lungs and air sacs indicated hemolytic activity on blood agar plates upon extended incubation for 48 hr at room temperature in air after initial incubation at 37 C for 48 hr under 7.5% CO2. This report characterizes the beta-hemolytic activity of O. rhinotracheale isolates by using in vitro kinetic hemolysis assays with sheep red blood cells, western blotting with leukotoxin-specific monoclonal antibodies, and isobaric tagging and relative and absolute quantitative (iTRAQ) analysis of O. rhinotracheale outer membrane protein digest preparations. The kinetic analyses of the hemolytic activity with red blood cells indicated that the protein is a pore former. iTRAQ analysis with membrane preparations revealed four peptides with homology to Mannheimia haemolytica leukotoxin and two peptides with homology to Actinobacillus actinoacetemcomitans leukotoxin. This is the first report that North American field isolates of O. rhinotracheale may express a hemolysin-like activity.

Iron acquisition by Ornithobacterium rhinotracheale.

Ornithobacterium rhinotracheale (ORT) is an emerging respiratory pathogen of poultry in North America that is causing millions of dollars in economic losses to the poultry industry. Ornithobacterium rhinotracheale is associated with airsacculitis, pleuritis, pneumonia, and consolidation of lungs. Little is known about the molecular mechanisms of infection. In this study, the mechanism of iron acquisition by O. rhinotracheale was explored. O. rhinotracheale strains grown under iron deprivation in media containing 200 microM 2,2'-dipyridyl did not secrete siderophores as measured by the chrome azurol S (CAS) agar and CAS solution assays. Filter disks impregnated with various protein-bound iron compounds and inorganic iron salts of Fe(III) and Fe(II) placed on iron-restricted agar inoculated with a lawn of O. rhinotracheale supported growth from sheep and porcine hemoglobins, ovotransferrin, Fe(III), and Fe(II), but they did not support growth from bovine transferrin, bovine apo-transferrin, bovine lactoferrin, and hemin. However, both bovine hemoglobin and transferrin supported growth of O. rhinotracheale serotype C. Four immunoreactive proteins involved in iron acquisition were identified in an O. rhinotracheale membrane extract by using mass spectrometry. Furthermore, O. rhinotracheale field strains showed differential sensitivity to 2,2'-dipyridyl. Of the 72 field strains tested, 22 strains were resistant to the iron chelator at concentrations of 50 microM and 100 microM, suggesting this attribute may be related to disease-producing potential of these strains. This is the first report on the identification of the iron acquisition mechanism of O. rhinotracheale.