Submitted to: Annual North American Cystic Fibrosis Conference
Publication Type: Abstract Only
Publication Acceptance Date: 7/12/2000
Publication Date: N/A
Technical Abstract: Antimicrobial peptides isolated from respiratory secretions are active in vitro against Gram-negative and Gram-positive bacteria, fungi, and Pseudomonas aeruginosa associated with chronic respiratory inflammation in cystic fibrosis (CF) patients. However, their activities against organisms in vivo have rarely, if ever, been demonstrated. In this study, we were interested to see if endogenous AP would attach to bacteria deposited directly into the lung. Sheep BAL fluid contains 0.83 +/ 0.33 mM AP to which Mannheimia haemolytica is susceptible (MIC 0.08 mM). To show this, 8 adult sheep (mean age 3.3 yrs.) were lightly sedated with 10 mg xylazine. A bronchoscope was inserted into the nasopharynx, and 2% lidocaine hydrochloride was administered at the larynx. The anterior portion of the right cranial lobe was saturated with 25 ml of M. haemolytica (10.2 log10 CFU/ml). Sheep were euthanized at 0, 5, 10 and 20 minutes. At necropsy, pieces of tissue were then taken for quantitative bacteriological culture, histopathology, and immunoelectron microscopy. Organisms were clearly seen by histopathology and electron microscopy to be attached to the alveolar epithelium in all groups. Monoclonal antibody to H-DDDDDDD-OH and protein A-colloidal gold were used to identify regions of AP bound to bacterial cells. Initially at 0 min, M. haemolytica appeared normal and PACG label was seen at the bacterial surface. At 5 to 20 min, many organisms were distorted and contained flocculated intracellular constituents characteristic of AP cellular damage. PACG was seen in and around cells. These results indicate that AP, in pulmonary secretions, can bind and presumably inactivate organisms entering into the lower respiratory tract. This research was supported by a Cystic Fibrosis Foundation Grant.