Veterinary Research Forum

Abstract The rise of antimicrobial resistance poses a severe global health challenge, with multidrug-resistant 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 nonapeptide, PCNP 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 Basic Local Alignment Search Tool identified homologs, and structural superposition with template modelin-align (TM-align) confirmed high conservation (TM-score 0.921, Root Mean Square Deviation 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., N-acetylmuramic acid - N-acetylglucosamine dimer, tetramer). Results revealed superior binding affinities in the native endolysin (e.g., – 10.35 kcal mol^-1 for N-acetylmuramic acid -L-alanine) compared to the engineered variant (– 9.65 kcal mol^-1 for N-acetylmuramic acid - N-acetylglucosamine -L-alanine trimer), with positive energies for larger ligands indicating steric hindrance. This trade-off reduced affinity for improved OM penetration supports potential enhancement of OM penetration while preserving bactericidal function 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 antimicrobial resistance.

Abstract Cholestasis, a multi-faceted liver complication arising from an imbalance in bile production and secretion, can lead to damage in extra-hepatic organs, including the reproductive system. This study aimed to investigate the therapeutic potential of Livergol^®, a silymarin-based drug, in the reproductive complications induced by experimental cholestasis in mature male rats. Twenty adults male Wistar rats were randomly divided into four groups (n = 5), including group 1 (control) remained untreated, group 2 (sham) underwent laparotomy and bile duct manipulation without ligation, group 3 (cholestasis) underwent bile duct ligation (BDL) to induce experimental cholestasis, and group 4 (cholestasis + Livergol^®) received oral Livergol^® (300 mg kg^-1) for 4 weeks post-BDL. After the study period, testicular histopathology was evaluated using Johnsen's score and the Cosentino criterion. Epididymal sperm parameters, including count, viability, and motility, were also assessed. The BDL-induced cholestasis caused significant testicular damage, evidenced by a decreased Johnsen's score and an increased Cosentino criterion. It also led to a significant reduction in sperm count, viability, and motility. Treatment with Livergol^® significantly ameliorated these effects, leading to a higher Johnsen's score, a lower Cosentino criterion, and marked improvements in all evaluated sperm parameters compared to the untreated cholestatic group. Oral administration of Livergol^® effectively attenuates testicular damage and improves spermatogenesis and sperm quality in rats with experimental cholestasis. These findings suggest Livergol^® as a promising protective agent against cholestasis-induced reproductive toxicity, likely due to the anti-oxidant properties of its active component, silymarin.