Version 2: Molecular characterization and transmission pattern of tetracycline resistance gene in tigecycline and carbapenem resistance klebsiella pneumoniae isolates at a tertiary care hospital India

Background:

The increasing prevalence of tigecycline and carbapenem-resistant K. pneumoniae (CRKP) poses a serious challenge, especially in resource-limited settings. Its ability to exchange resistance genes with other bacteria accelerates the spread of multidrug resistance. While carbapenems and tetracyclines have been used effectively against K. pneumoniae, resistance to these agents is now rising globally, narrowing available treatment options.

Objective:

The study aimed to determine the phenotypic and genotypic prevalence of carbapenem and tetracycline resistance K. pneumoniae isolates along with transferability pattern of carbapenem and tetracycline resistance gene in these isolates. 

Methodology

Clinical isolates from pus and respiratory samples were identified using biochemical tests and MALDI-TOF MS. AST was performed by the Kirby-Bauer disc diffusion method, and MICs were determined by BMD method. PCR was performed to detect carbapenemase (bla_NDM, bla_OXA-48, bla_KPC) and tetracycline resistance genes [tet(A), tet(B), tet(K), tet(M), tet(S)], followed by Sanger sequencing for validation. Conjugation assays assessed gene transferability. 

Results

Out of 152 carbapenem resistant K. pneumoniae isolates, 31 (20.4%) were found to be resistant to tigecycline. All tigecycline-resistant isolates (31/31; 100%) exhibited complete resistance to ceftazidime, ciprofloxacin, and omadacycline. Additionally, resistance to amikacin and cefoperazone-sulbactam (SCF) was observed in 87.1% (27/31) and 77.4% (24/31) of the isolates. Resistance to minocycline and colistin was detected in 51.6% (16/31) and 29.0% (9/31) of the isolates, respectively. 

PCR analysis revealed that 51.6% (16/31) of the isolates carried the bla_OXA-48 gene, and 29.0% (9/31) carried the blaNDM gene. None of the isolates harbored the bla_KPC gene. With respect to tetracycline resistance determinants, the tet(A) gene was detected in 12.9% (4/31) of the isolates, and the tet(B) gene in 3.2% (1/31), while tet(K), tet(M), tet(S) and bla_KPC were not detected in any isolate. 

Conjugation assays demonstrated that plasmids carrying bla_NDM and bla_OXA-48 were transferable to a recipient strain, indicating their potential for horizontal gene transfer. In contrast, plasmids harboring tet(A) and tet(B) genes were not transferable under the experimental conditions. 

Conclusion

Tigecycline-resistant K. pneumoniae isolates showed high multidrug resistance, with transferable bla_NDM and bla_OXA-48 genes. In contrast, Chromosome and plasmid borne tetracycline resistance genes tet(A) and tet(B) were non-transferable, indicating limited horizontal spread.