Identification of Antimicrobial Peptides in Dry Secretions of Dairy Cows and Their In Vitro Efficacy Against Mastitis Pathogens
Creators: Hasanllari, Barjam
Publisher: name: University of Alberta Library
Year: 2024

<p>The dairy industry heavily relies on the health of dairy cows for optimal milk production, but mastitis, characterized by udder inflammation, poses significant challenges. This study aimed to identify the peptide signatures related to SCM and different time points throughout the dry-off period in dry secretions of dairy cows to understand how peptide composition influences mastitis susceptibility and mammary gland physiology. In addition, this study aimed to shed light on the mastitis therapeutic potential of known AMPs and new potential AMPs deriving from dry secretions. A combination of liquid chromatography and mass spectrometry (LC-MS) analyses was used for 32 peptide samples extracted from the dry secretions of dairy cows collected on days 2 and 21 of the dry-off period. These samples were initially categorized into H and SCM quarters based on SCC. Distinct patterns in peptide composition were found between H and SCM cows, with temporal variations observed during the dry-off period. In total, 1,235 peptides were detected among the four groups (SCM-D21, SCM-D2, H-D21, and H-D2), sourced from 59 distinct proteins, predominantly deriving from β-casein. Comparing SCM and H groups at day 2 (SCM-D2 vs. H-D2) showed 158 differentially expressed peptides, and SCM-D21 vs. H-D21 revealed 138 differentially expressed peptides. Within each health condition, comparing time points (SCM-D21 vs. SCM-D2 and H-D21 vs. H-D2) revealed significant peptide expression changes, with 635 peptides differing in SCM-D21 vs. SCM-D2 and 651 peptides in H-D21 vs. H-D2. Functional analysis revealed 56 peptides with a total of 17 functions, such as antimicrobial, immunomodulatory, and cytomodulatory functions, shedding light on their roles in mastitis pathogenesis and udder physiology. The comparative analyses also revealed changes in the levels of these functional peptides across the 4 different groups, indicating their potential involvement in regulating immune responses, oxidative stress, inflammatory processes, and other biological processes during subclinical mastitis and the dry-off period. Furthermore, using disc diffusion assay and 96 well MIC assay, five known AMPs (cathelicidin LL37-002, cathelicidin 37-005, anti-infective peptide, indolicidin, lactoferricin) were evaluated for their efficacy against mastitis-causing pathogens, including S. aureus, S. uberis, S. agalactiae, S. dysgalactiae, E. coli, S. haemolyticus, S. simulans, S. chromogenes, and K. pneumoniae. Indolicidin emerged as a potent candidate, demonstrating broad-spectrum inhibitory effects, except against E. coli. Synergistic effects were observed when combining indolicidin with the other known AMPs against E. coli. However, new potential AMPs identified in dry secretions (CHTL-1A, CHTL-1B, CHTL-1C, CHTL-1D, CHTL-1E, CHTL-1F, CHTL-2A, CHTL-2B, CHTL-2C, CHTL-2D, CHTL-2E, CHTL-5A, CHTL-6A), tested against S. aureus and E. coli using 96 well MIC assay, showed no antimicrobial activity. These findings underscore the complexity of AMP interactions and highlight the ongoing need for innovative therapeutic strategies in mastitis management. Overall, this research contributes valuable insights into mastitis detection, management, and dairy cow health monitoring, offering promising avenues for enhancing milk quality and industry sustainability.</p>