In Vitro Microbiological Characterization of a Novel Azalide, Two Triamilides and an Azalide Ketal against Bovine and Porcine Respiratory Pathogens

Several novel 15-membered-ring macrolide agents (azalide 1, triamilides 2 and 3, and the azalide 3, 6-ketal 4) were identified as potential antibacterial agents against Mannheimia (formerly named as Pasteurella) haemolytica, Pasteurella multocida, Haemophilus somnus and Actinobacillus pleuropneumoniae, important etiological agents of bovine and porcine respiratory disease. Compound 3 is the major component of the antibiotic tulathromycin. Antibacterial activity against tilmicosin-resistant P. multocida field isolates was also tested. In vitro MIC 50/90 analysis revealed that the four newly synthesized compounds were more potent than tilmicosin against M. haemolytica (4-8×), P. multocida (8-16×), A. pleuropneumoniae (4×), H. somnus (2× and 16×), and tilmicosin-resistant P. multocida (32×). In time-kill kinetic studies, all four novel compounds and tilmicosin showed bactericidal activity against M. haemolytica, P. multocida and A. pleuropneumoniae at both 4× and 8× MIC. A functional assay using genetically defined mutants revealed that all four novel compounds were poorer substrates for the efflux pump, AcrA/B system, than tilmicosin. A pH study using LPS mutants indicated that the enhanced in vitro potency of the triamilides, particularly compound 3 was mainly due to better penetration of the molecule through the outer membrane. The third amine group at the C-4" position of the triamilde molecules contributed to this increased membrane penetration by increasing overall basicity. These studies indicate that the four novel compounds have potential as antibacterial agents against bovine and porcine respiratory disease.