Veterinary Research Forum

Abstract Pasteurella multocida and Mannheimia haemolytica are significant bacteria of respiratory tract infections in cattle. Identifying whether these bacteria are commensal or pathogenic is crucial for understanding their role in predisposing calves to respiratory tract infections. This study aimed to determine the prevalence and pathogenic characteristics of P. multocida and M. haemolytica in nasal swab samples of clinically healthy calves in Konya, Turkey, and to assess antibiotic resistance profiles. Using conventional culture methods, 5 samples (1.67%) from 4 different farms were positive for P. multocida and 3 samples (1%) were positive for M. haemolytica. PCR of nasal swab samples yielded P. multocida positivity in 17 (5.67%) samples and M. haemolytica positivity in 13 (4.33%), including all culture-positive samples. While all P. multocida isolates were atypical for the capsule, 18% of the PCR-positive samples were L2, 70% L3A, and 12% L8 serotypes. The ompA, omp87, hghA, and tbpA genes were the most prevalent, while the presence of other virulence genes varied. High rates of ampicillin/cloxacillin (100%) and streptomycin (80%) resistance were observed among the isolates. The rates of serotypes A1 and A6 in M. haemolytica PCR-positive samples were 53.85% and 46.15%, respectively. The ssa-1 gene was detected in 33.33% of the isolates. Moderate resistance to trimethoprim/sulfamethoxazole, streptomycin, and neomycin (66.67%) was recorded. The findings revealed the presence of P. multocida and M. haemolytica with various virulence characteristics and antimicrobial resistance profiles in the nasal microbiota of healthy calves, providing novel insights that could inform improved herd management, disease control, and treatment strategies.

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 Pathogenic Escherichia coli is one of the world’s most important zoonotic foodborne pathogens and poses a serious threat to public health. We examined the prevalence, virulence genes, and antibiotic resistance profile of Shiga toxin (Stx)-producing E. coli (STEC) isolated from broiler chickens in the Kafr El-Sheikh governorate, Egypt. A total of 410 samples (230 cloacal swabs, 180 internal organs) were collected to isolate E. coli. A total of 29 (7.07%) E. coli isolates were recovered and identified, and 18 of them harbored Stx genes (stx). Out of 18 isolates, five (17.24%) carried the stx1 gene, five (17.24%) carried the stx2 gene, four (13.79%) carried both stx1 and stx2 genes, and four (13.79%) carried stx1, stx2, and eaeA genes. Overall, complete antibiotic resistance was observed against amoxicillin, ampicillin, cefpodoxime, and cefoperazone; high resistance was observed against ampicillin/sulbactam, nalidixic acid, cefuroxime, aztreonam, ciprofloxacin, ceftriaxone, chloramphenicol, sulfamethoxazole/ trimethoprim, and ceftazidime; moderate resistance against gentamicin; low resistance against cefoxitin; lower resistance was detected against norfloxacin, cefotetan, and amikacin; and the lowest resistance against imipenem. All E. coli isolates demonstrated multidrug resistance against at least four antibiotic classes. Out of 29 E. coli isolates, STEC accounted for 18 isolates, of which the O78, O26:H11, O128:H2, O1:H7, O119:H6, and O91:H21 serogroups were predominant. All E. coli isolates were multidrug resistant and therefore pose a potential public health concern as these virulent, resistant strains may spread to humans. Thus, high levels of hygiene and biosecurity are required by chicken handlers to decrease the danger of infection spreading to humans.