Objective—To evaluate antimicrobial activity of bovine bactericidal permeability–increasing protein (bBPI)–derived synthetic peptides against mastitis-causing gram-negative bacteria.
Sample Population—Bacterial isolates from the milk of cows with clinical mastitis.
Procedures—3 peptides were synthesized with sequences corresponding to amino acids 65 to 99 (bBPI65–99) or 142 to 169 (bBPI142–169) or the combination of amino acids 90 to 99 and 148 to 161 (bBPI90–99,148–161) of bBPI. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of these peptides against bacterial isolates from cows with mastitis were determined by use of a standardized broth microdilution assay. The ability of these peptides to retain their antimicrobial activity in serum and milk was also evaluated. Finally, bacterial lipopolysaccharide (LPS)-neutralizing activity of these peptides was assayed with the Limulus amebocyte lysate test.
Results—Of the 3 peptides tested, bBPI90–99,148–161 had the widest spectrum of antimicrobial activity, with MIC and MBC values ranging from 16 to 64 Mg/mL against Escherichia coli, Klebsiella pneumoniae, and Enterobacter spp and from 64 to 128 Mg/mL against Pseudomonas aeruginosa. None of the peptides had any growth-inhibitory effect on Serratia marcescens. The antimicrobial activity of bBPI90–99,148–161 was inhibited in milk, but preserved in serum. Finally, bBPI142–169 and bBPI90–99,148–161 completely neutralized LPS.
Conclusions and Clinical Relevance—bBPI90–99,148–161 is a potent neutralizer of the highly proinflammatory molecule bacterial LPS and has antimicrobial activity against a variety of gram-negative bacteria. The ability of bBPI90–99,148–161 to retain antimicrobial activity in serum suggests a potential therapeutic application for this peptide in the management of gram-negative septicemia.
Objective—To evaluate the effects of cis–urocanic acid (cis-UCA) on mammary gland (MG) inflammation and injury associated with Escherichia coli–induced mastitis in dairy cows.
Animals—12 lactating dairy cows (36 MGs).
Procedures—At 7-week intervals, a different MG in each cow was experimentally inoculated with E coli. At 6-hour intervals from 6 to 36 hours after inoculation, the inoculated MG in each cow was infused with 40 mL of saline (0.9% NaCl) solution, 12.5mM cis-UCA, or 25mM cis-UCA (4 cows/group); ultimately, each cow received each treatment. Immediately prior to and at various time points after inoculation and treatment, milk samples were collected. Bacterial CFUs, somatic cell counts (SCCs), N-acetyl-beta-D-glucosaminidase (NAGase) and lactate dehydrogenase (LDH) activities, and concentrations of bovine serum albumin, tumor necrosis factor-α, and cis-UCA were quantified in each milk sample.
Results—Compared with findings in saline solution–treated MGs, NAGase and LDH activities in milk samples from cis-UCA–treated MGs were lower. Cis-UCA had no effect on milk SCCs and milk concentrations of bovine serum albumin and tumor necrosis factor-α. Furthermore, cis-UCA had no adverse effect on bacterial clearance; CFUs of E coli in MGs treated with saline solution or cis-UCA were equivalent.
Conclusions and Clinical Relevance—In cows, milk NAGase and LDH activities were both lower in E coli–infected MGs infused with cis-UCA than in those infused with saline solution, which suggests that cis-UCA reduced mastitis-associated tissue damage. Furthermore, these data indicated that therapeutic concentrations of cis-UCA in milk can be achieved via intramammary infusion.