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All detectable high-molecular-mass penicillin-binding proteins are modified in a high-level beta-lactam-resistant
A Amoroso1, D Demares, M Mollerach
1Laboratorio de resistencia microbiana, Cátedra de Microbiología. Departamento de Microbiología, Inmunología y Biotecnología, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina.
Abstract:
All detectable high-molecular-mass penicillin-binding proteins (HMM PBPs) are altered in a clinical isolate of Streptococcus mitis for which the beta-lactam MICs are increased from those previously reported in our region (cefotaxime MIC, 64 microg/ml). These proteins were hardly detected at concentrations that saturate all PBPs in clinical isolates and showed, after densitometric analysis, 50-fold-lower radiotracer binding. Resistance was related to mosaic structure in all HMM PBP-coding genes, where critical region replacement was complemented not only by substitutions already reported for the closely related Streptococcus pneumoniae but also by other specific replacements that are presumably close to the active-site serine. Mosaic structure was also presumed in a pbp1a-sensitive strain used for comparison, confirming that these structures do not unambiguously imply, by themselves, detectable critical changes in the kinetic properties of these proteins.
Insights
High-molecular-mass penicillin-binding proteins (HMM PBPs) in Streptococcus mitis show significantly reduced radiotracer binding, indicating altered function. This resistance is linked to mosaic gene structures in HMM PBPs.
Area of Science:
- Microbiology
- Molecular Biology
- Antimicrobial Resistance
Background:
- Penicillin-binding proteins (PBPs) are crucial targets for beta-lactam antibiotics.
- Clinical isolates of Streptococcus mitis can develop resistance to beta-lactam antibiotics.
- Previous studies have reported lower beta-lactam minimum inhibitory concentrations (MICs) for S. mitis in the region.
Purpose of the Study:
- To investigate the alterations in high-molecular-mass penicillin-binding proteins (HMM PBPs) in a clinical isolate of Streptococcus mitis with increased beta-lactam MICs.
- To determine the relationship between PBP alterations and antimicrobial resistance.
- To analyze the genetic basis of observed PBP changes.
Main Methods:
- Analysis of high-molecular-mass penicillin-binding proteins (HMM PBPs) in a clinical isolate of Streptococcus mitis.
- Radiotracer binding assays to quantify PBP activity.
- Densitometric analysis of protein binding.
- Sequencing and analysis of PBP-coding genes to identify mosaic structures and substitutions.
Main Results:
- Detectable high-molecular-mass penicillin-binding proteins (HMM PBPs) were significantly reduced in the resistant S. mitis isolate.
- Radiotracer binding to HMM PBPs was 50-fold lower compared to susceptible strains.
- Resistance correlated with mosaic structures in all HMM PBP-coding genes, including novel substitutions.
- Mosaic structures alone did not definitively indicate critical kinetic property changes.
Conclusions:
- Alterations in HMM PBPs, characterized by reduced binding and mosaic gene structures, contribute to beta-lactam resistance in this Streptococcus mitis clinical isolate.
- The identified genetic alterations in HMM PBPs provide insight into the mechanisms of antimicrobial resistance.
- Further research is needed to fully elucidate the functional impact of these mosaic structures on PBP activity.