Submitted to: American Association of Veterinary Laboratory Diagnosticians
Publication Type: Abstract Only
Publication Acceptance Date: 7/15/2017
Publication Date: N/A
Interpretive Summary: Bovine respiratory disease complex (or pneumonia), which is commonly known as shipping fever, is causing severe economic losses to the beef and dairy cattle industry in the United States. Bacterial pathogens associated with cattle respiratory diseases are normal inhabitants of the upper respiratory tract. Due to the emergence of antibiotic-resistant bacteria, there is a need for the identification and development of reagents for an alternative to antibiotics. In this regard, we tested several small proteins (NK-lysins) for their killing activities against bacterial pathogens causing respiratory diseases in cattle. Small proteins were able to kill these bacteria very efficiently. Further studies are needed to assess whether small proteins tested in this study can be used for prophylaxis or treat cattle against bacteria causing respiratory diseases.
Technical Abstract: Bovine NK-lysins, which are functionally and structurally similar to human granulysin and porcine NK-lysin, are predominantly found in the granules of cytotoxic T-lymphocytes and NK-cells. Although antimicrobial activity of bovine NK-lysin has been assessed for several bacterial pathogens, not all the important bacterial pathogens that are involved in the bovine respiratory disease complex have been studied. Therefore the objective of the present study was to evaluate the antimicrobial activity of bovine NK-lysin-derived peptides on bovine respiratory pathogen Histophilus somni. Four, 30-mer peptides corresponding to the functional region of NK-lysin helices 2 and 3 were synthesized and assessed for antibacterial activity on four bovine pneumonic H. somni isolates. Although there were some differences in the efficiency of bactericidal activity among the NK-lysin peptides at lower concentrations (2 - 5 µM), all four peptides effectively killed most H. somni isolates at higher concentrations (10 - 30 µM) as determined by a bacterial killing assay. Confocal microscopic and flow cytometric analysis of Live/Dead Baclight stained H. somni (which were preincubated with NK-lysin peptides) were consistent with the killing assay findings and suggest NK-lysin peptides are bactericidal for H. somni. Among the four peptides, NK2A-derived peptide consistently showed the highest antimicrobial activity against all four H. somni isolates. Electron microscopic examination of H. somni following incubation with NK-lysin revealed extensive cell membrane damage, protrusions of outer membranes, and cytoplasmic content leakage. Taken together, the findings from this study clearly demonstrate the antimicrobial activity of all four bovine NK-lysin-derived peptides against bovine H. somni isolates.