Veterinary Research Forum

Abstract The rise of antimicrobial resistance (AMR) poses a severe global health challenge, with multidrug-resistant (MDR) bacteria causing millions of deaths annually and diminishing the effectiveness of traditional antibiotics. Bacteriophage-derived endolysins, such as PVP-SE1gp146 from Salmonella phage PVP-SE1, emerge as promising alternatives due to their high specificity, low toxicity, and minimal resistance development. However, their activity against Gram-negative bacteria is limited by poor outer membrane (OM) penetration. This study investigates the structural and functional impacts of fusing the polycationic PCNP peptide (KRKKRKKRK) to PVP-SE1gp146 to create engineered endolysin that enhance OM permeability while preserving enzymatic function.
Using advanced computational tools including Phyre2, GalaxyWEB, and AlphaFold2, three-dimensional models of native and engineered endolysins were generated and validated with Verify3D. Sequence analysis via BLAST identified homologs, and structural superposition with TM-align confirmed high conservation (TM-score 0.921, RMSD 1.84 Å) despite the PCNP insertion. Active sites were predicted using PrankWeb, and molecular docking with AutoDock assessed interactions with peptidoglycan components (e.g., NAM-NAG dimer, tetramer). Results revealed superior binding affinities in the native endolysin (e.g., -10.35 kcal/mol for NAM-L-alanine) compared to the engineered variant (-9.65 kcal/mol for NAM-NAG-L-alanine trimer), with positive energies for larger ligands indicating steric hindrance.
This trade-off—reduced affinity for improved OM penetration—supports enhanced bactericidal activity against MDR pathogens like Pseudomonas aeruginosa. The findings highlight the potential of rational engineering for superior antimicrobials, emphasizing the need for molecular dynamics simulations and experimental validation to optimize Artilysin design in combating AMR.

Abstract This study was conducted in West Azerbaijan province, Iran (37°27'18.022" N, 45°0'0" E) to investigate the genotyping and phylogenetic characterization of Mannheimia haemolytica in cattle and buffaloes from November 2022 to January 2024. Mannheimia haemolytica is a bacterium known to cause pasteurellosis pneumonia, a respiratory disease in ruminants, such as cattle and sheep. This is one of the main causes of economic losses in the feedlot industry. In addition to the deaths, treatment costs are also significant. The lung and nasal swab samples were collected from 378 cattle and buffaloes. The M. haemolytica was detected in 32 (8.46%) of the samples, with a notably higher isolation rate from lung tissue (56.25%; n = 18) compared to the nasal swabs (43.75%; n = 14). Interestingly, the study also revealed a seasonal pattern, with the highest isolation rates observed during January, February, and March. Multi-locus sequence typing demonstrated that all isolates belonged to sequence type 1 (ST1) within clonal complex 28. This finding is consistent with the global prevalence of ST1 in bovine isolates, indicating widespread distribution. Phylogenetic analysis revealed a strong correlation between ST1 and STs 30 and 54, highlighting the prevalence of ST1 in M. haemolytica among ruminants in West Azerbaijan, Iran. Further research is needed to investigate its potential for causing disease and its transmission pattern.