Characteristics of Klebsiella spp. from sheep flocks in northern China

Background Klebsiella spp. are Gram-negative opportunistic pathogens that affect both humans and animals. However, data on their antimicrobial resistance and virulence characteristics in sheep, particularly in China, remain limited. Methods In 2024, 421 sheep fecal samples were collected from six regions in northern China. Klebsiella spp. were isolated and identified, followed by antimicrobial susceptibility testing and PCR screening for virulence-associated genes. The pathogenic potential of selected isolates was evaluated using the Galleria mellonella infection model. Whole-genome sequencing was performed on 26 selected isolates to characterize their antimicrobial resistance and virulence determinants. Results Klebsiella spp. were isolated from 120 samples, corresponding to an overall detection rate of 28.5%, including 85 Klebsiella pneumoniae isolates (70.8%, 85/120). All isolates were resistant to sulfamethoxazole, with high resistance to tetracycline (76.7%), florfenicol (42.5%), and ciprofloxacin (41.7%). Resistance to tigecycline (10.0%) and polymyxin B (5.8%) was also observed at lower levels. Overall, 64.2% of the isolates exhibited multidrug resistance. PCR screening revealed virulence-associated genes, including iucA (10.8%), rmpA (8.3%), peg -344 (0.8%), and iroB (3.3%). Based on their virulence-associated gene profiles, 14 isolates were selected for further analysis, of which 11 exhibited high pathogenic potential in the Galleria mellonella infection model. Whole-genome sequencing revealed diverse virulence and antimicrobial resistance determinants. No plasmid-mediated resistance determinants related to polymyxin B (e.g., mcr-like genes) or recognized tigecycline resistance determinants were identified in the genomes. In the isolates analyzed in this study, mutations in lipid A modification and regulatory genes, including arnT and pmrA / pmrB , may have contributed to polymyxin B resistance, while mutations in efflux pump-associated genes, including oqxA / oqxB, acrA , and acrR , may have contributed to tigecycline resistance. Conclusion Multidrug resistance and pathogenic potential coexist among Klebsiella spp. isolated from healthy sheep in northern China. These findings suggest that sheep-associated Klebsiella may serve as reservoirs of clinically relevant antimicrobial resistance and virulence determinants, highlighting the need for continued surveillance and prudent antimicrobial use in livestock.