RESUMEN
Methicillin-resistant Staphylococcus aureus (MRSA), a Gram-positive bacterial pathogen, continues to pose a serious threat to the current public health system in our society. The high level of resistance to ß-lactam antibiotics in MRSA is attributed to the expression of penicillin-binding protein 2a (PBP2a), which catalyzes cell wall cross-linking. According to numerous research reports, the activity of the PBP2a protein is known to be regulated by an allosteric site distinct from the active site where cell wall cross-linking occurs. Here, we conducted a screening of 113 compounds containing a 1,3,4-oxadiazole core to design new covalent inhibitors targeting the allosteric site of PBP2a and establish their structural-activity relationship. The stereochemically selective synthesis of sulfonyl oxadiazole compounds identified in the initial screening resulted in a maximum eightfold enhancement in cell inhibition activity. The sulfonyl oxadiazole-based compounds formulated as PEG-based ointments, with low toxicity test results on human cells (CC 50 : >78µM), demonstrated potent antimicrobial effects not only in a mouse skin wound infection model but also against oxacillin-resistant clinical isolate MRSA (IC 50 ≈ 1µM), as evidenced by the results. Furthermore, additional studies utilizing LC-MS/MS and in-silico approaches clearly support the allosteric site covalent binding mechanism through the nucleophilic aromatic substitution (S N Ar) reaction, as well as its association with the closure of the major active site of PBP2a.
RESUMEN
Background: Elizabethkingia anophelis is an emerging Gram-negative nonlactose fermenter in the health care setting, where it causes life-threatening infections in immunocompromised patients. We aimed to characterize the molecular mechanisms of antimicrobial resistance and evaluate the utility of contemporary antibiotics with the intent to offer targeted therapy against an uncommonly encountered pathogen. Methods: Whole-genome sequencing (WGS) was conducted to accurately identify isolate species and elucidate the determinants of ß-lactam resistance. Antimicrobial susceptibility testing was performed using broth microdilution and disk diffusion assays. To assess the functional contribution of the major metallo-ß-lactamase (MBL) encoding genes to the resistance profile, bla BlaB was cloned into pBCSK(-) phagemid vector and transformed into Escherichia coli DH10B. Results: WGS identified the organism as E. anophelis. MBL genes bla BlaB-1 and bla GOB-26 were identified, in addition to bla CME-2, which encodes for an extended-spectrum ß-lactamase (ESBL). Plasmids were not detected. The isolate was nonsusceptible to all commonly available ß-lactams, carbapenems, newer ß-lactam ß-lactamase inhibitor combinations, and to the combination of aztreonam (ATM) with ceftazidime-avibactam (CAZ-AVI). Susceptibility to the novel siderophore cephalosporin cefiderocol was determined. A BlaB-1 transformant E. coli DH10B isolate was obtained and demonstrated increased minimum inhibitory concentrations to cephalosporins, carbapenems, and CAZ-AVI, but not ATM. Conclusions: Using WGS, we accurately identified and characterized an extensively drug-resistant E. anophelis in an immunocompromised patient. Rapid evaluation of the genetic background can guide accurate susceptibility testing to better inform antimicrobial therapy selection.
RESUMEN
Treatment of multidrug-resistant Gram-negative bacterial pathogens represents a critical clinical need. Here, we report a novel γ-lactam pyrazolidinone that targets penicillin-binding proteins (PBPs) and incorporates a siderophore moiety to facilitate uptake into the periplasm. The MIC values of γ-lactam YU253434, 1, are reported along with the finding that 1 is resistant to hydrolysis by all four classes of ß-lactamases. The druglike characteristics and mouse PK data are described along with the X-ray crystal structure of 1 binding to its target PBP3.
