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Cefotaximase-Munich (CTX-M) extended-spectrum beta-lactamases (ESBLs) are commonly associated with Gram-negative, hospital-acquired infections worldwide. Several beta-lactamase inhibitors, such as clavulanate, are used to inhibit the activity of these enzymes. To understand the mechanism of CTX-M-15 activity, we have determined the crystal structures of CTX-M-15 in complex with two specific classes of beta-lactam compounds, desfuroylceftiofur (DFC) and ampicillin, and an inhibitor, clavulanic acid. The crystal structures revealed that Ser70 and five other residues (Lys73, Tyr105, Glu166, Ser130, and Ser237) participate in catalysis and binding of those compounds. Based on analysis of steady-state kinetics, thermodynamic data, and molecular docking to both wild-type and S70A mutant structures, we determined that CTX-M-15 has a similar affinity for all beta-lactam compounds (ceftiofur, nitrocefin, DFC, and ampicillin), but with lower affinity for clavulanic acid. A catalytic mechanism for tested β-lactams and two-step inhibition mechanism of clavulanic acid were proposed. CTX-M-15 showed a higher activity toward DFC and nitrocefin, but significantly lower activity toward ampicillin and ceftiofur. The interaction between CTX-M-15 and both ampicillin and ceftiofur displayed a higher entropic but lower enthalpic effect, compared with DFC and nitrocefin. DFC, a metabolite of ceftiofur, displayed lower entropy and higher enthalpy than ceftiofur. This finding suggests that compounds containing amine moiety (e.g., ampicillin) and the furfural moiety (e.g., ceftiofur) could hinder the hydrolytic activity of CTX-M-15. 

Antibiotic use and bacterial transmission are responsible for the emergence, spread and persistence of antimicrobial-resistant (AR) bacteria, but their relative contribution likely differs across varying socio-economic, cultural, and ecological contexts. To better understand this interaction in a multi-cultural and resource-limited context, we examine the distribution of antimicrobial-resistant enteric bacteria from three ethnic groups in Tanzania. Household-level data (n = 425) was collected and bacteria isolated from people, livestock, dogs, wildlife and water sources (n = 62,376 isolates). The relative prevalence of different resistance phenotypes is similar across all sources. Multi-locus tandem repeat analysis (n = 719) and whole-genome sequencing (n = 816) ofEscherichia colidemonstrate no evidence for host-population subdivision. Multivariate models show no evidence that veterinary antibiotic use increased the odds of detecting AR bacteria, whereas there is a strong association with livelihood factors related to bacterial transmission, demonstrating that to be effective, interventions need to accommodate different cultural practices and resource limitations.

 

Biofilm formation causes prolonged wound infections due to the dense biofilm structure, differential gene regulation to combat stress, and production of extracellular polymeric substances.Acinetobacter baumannii,Staphylococcus aureus, andPseudomonas aeruginosaare three difficult-to-treat biofilm-forming bacteria frequently found in wound infections. This work describes a novel wound dressing in the form of an electrochemical scaffold (e-scaffold) that generates controlled, low concentrations of hypochlorous acid (HOCl) suitable for killing biofilm communities without substantially damaging host tissue. Production of HOCl near the e-scaffold surface was verified by measuring its concentration using needle-type microelectrodes. E-scaffolds producing 17, 10 and 7 mM HOCl completely eradicatedS. aureus,A. baumannii, andP. aeruginosabiofilms after 3 hours, 2 hours, and 1 hour, respectively. Cytotoxicity and histopathological assessment showed no discernible harm to host tissues when e-scaffolds were applied to explant biofilms. The described strategy may provide a novel antibiotic-free strategy for treating persistent biofilm-associated infections, such as wound infections.