• Ruina Bao, Faculty of Veterinary Medicine, University of Calgary
• Zhi Ma, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou, China
• Kim Stanford, Department of Biological Sciences, University of Lethbridge, AB, Canada
• Tim A. McAllister, Agriculture and Agri-Food Canada, Lethbridge T1J 4B1, AB, Canada
• Yan D. Niu, Faculty of Veterinary Medicine, University of Calgary,

Bovine respiratory disease (BRD) is the leading cause of morbidity and mortality in cattle raised in North America. Upon feedlot arrival, cattle are usually subject to metaphylactic use of antibiotics to prevent and treat bacterial cases of BRD. Vaccination offers limited duration of immunity against M. haemolytica, the major bacterial causative agent of BRD. Emerging resistance to antibiotics has resulted in an urgent need for novel strategies to control members of the BRD bacterial complex.

The inhibitory effects of three antimicrobial peptides were evaluated against multi-drug resistant M. haemolytica isolates using microbroth dilution. Isolates from Alberta feedlots were screened for biofilm formation using the crystal violet assay. The most robust biofilm former, Mh330, was co-cultured with bovine turbinate (BT) cells to mimic infection conditions before an antimicrobial peptide, WK2, was added to inhibit attachment. Cytotoxicity of WK2 against bovine turbinate cells was assessed by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. To determine the inhibitory mechanism, the membrane integrity of Mh330 after peptide treatment was examined using NPN (1-N-phenylnapthylamine), and ONPG (o-Nitrophenyl β-D-galactopyranoside) assays.

Although all three peptides displayed inhibitory activity against M. haemolytica, WK2, a beta sheet peptide was bactericidal against strong biofilm former, isolate Mh330, at 8-16 μg/mL and decreased adhesion to BT cells by 3 logs at 32 μg/mL without cytotoxic effects. The NPN and ONPG assays determined that both inner and outer membranes were permeabilized.

Conclusion WK2 may be a viable alternative to antibiotics for the control of M. haemolytica related cases of BRD.