scholarly journals PBP 4 Mediates High-Level Resistance to New-Generation Cephalosporins in Staphylococcus aureus

2016 ◽  
Vol 60 (7) ◽  
pp. 3934-3941 ◽  
Author(s):  
Liana C. Chan ◽  
Aubre Gilbert ◽  
Li Basuino ◽  
Thaina M. da Costa ◽  
Stephanie M. Hamilton ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Core Genome ◽  
Binding Protein ◽  
Penicillin Binding Protein ◽  
Methicillin Resistant ◽  
Content Type ◽  
Resistant Strains ◽  
High Level ◽  
New Generation ◽  
Level Resistance

ABSTRACTStaphylococcus aureusis an important cause of both hospital- and community-associated methicillin-resistantS. aureus(MRSA) infections worldwide. β-Lactam antibiotics are the drugs of choice to treatS. aureusinfections, but resistance to these and other antibiotics make treatment problematic. High-level β-lactam resistance ofS. aureushas always been attributed to the horizontally acquired penicillin binding protein 2a (PBP 2a) encoded by themecAgene. Here, we show thatS. aureuscan also express high-level resistance to β-lactams, including new-generation broad-spectrum cephalosporins that are active against methicillin-resistant strains, through an uncanonical core genome-encoded penicillin binding protein, PBP 4, a nonessential enzyme previously considered not to be important for staphylococcal β-lactam resistance. Our results show that PBP 4 can mediate high-level resistance to β-lactams.

scholarly journals PBP4 Mediates β-Lactam Resistance by Altered Function

2017 ◽  
Vol 61 (11) ◽  
Author(s):  
Som S. Chatterjee ◽  
Liang Chen ◽  
Aubre Gilbert ◽  
Thaina M. da Costa ◽  
Vinod Nair ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Binding Protein ◽  
Missense Mutations ◽  
Penicillin Binding Protein ◽  
Content Type ◽  
High Level ◽  
Promoter Mutations ◽  
Level Resistance

ABSTRACT Penicillin binding protein 4 (PBP4) can provide high-level β-lactam resistance in Staphylococcus aureus. A series of missense and promoter mutations associated with pbp4 were detected in strains that displayed high-level resistance. We show here that the missense mutations facilitate the β-lactam resistance mediated by PBP4 and the promoter mutations lead to overexpression of pbp4. Our results also suggest a cooperative interplay among PBPs for β-lactam resistance.

scholarly journals Microtiter Plate-Based Assay for Inhibitors of Penicillin-Binding Protein 2a from Methicillin-Resistant Staphylococcus aureus

2011 ◽  
Vol 55 (6) ◽  
pp. 2783-2787 ◽  
Author(s):  
Sudheer Bobba ◽  
V. K. Chaithanya Ponnaluri ◽  
Mridul Mukherji ◽  
William G. Gutheil
Keyword(s):  
Staphylococcus Aureus ◽  
Binding Protein ◽  
Binding Constants ◽  
Public Health Problem ◽  
Microtiter Plate ◽  
Penicillin Binding Protein ◽  
Methicillin Resistant ◽  
Content Type ◽  
High Level

ABSTRACTPenicillin-binding protein 2a (PBP2a), the molecular determinant for high-level β-lactam resistance in methicillin-resistantStaphylococcus aureus(MRSA), is intrinsically resistant to most β-lactam antibiotics. The development and characterization of new inhibitors targeting PBP2a would benefit from an effective and convenient assay for inhibitor binding. This study was directed toward the development of a fluorescently detected β-lactam binding assay for PBP2a from MRSA. Biotinylated ampicillin and biotinylated cephalexin were tested as tagging reagents for fluorescence detection by using a streptavidin-horseradish peroxidase conjugate. Both bound surprisingly well to PBP2a, with binding constants of 1.6 ± 0.4 μM and 13.6 ± 0.8 μM, respectively. Two forms of the assay were developed, a one-step direct competition form of the assay and a two-step indirect competition form of the assay, and both forms of the assay gave comparable results. This assay was then used to characterize PBP2a binding to ceftobiprole, which gave results consistent with previous studies of ceftobiprole-PBP2a binding. This assay was also demonstrated for screening for PBP2a inhibitors by screening a set of 13 randomly selected β-lactams for PBP2a inhibition at 750 μM. Meropenem was observed to give substantial inhibition in this screen, and a follow-up titration experiment determined its apparentKito be 480 ± 70 μM. The availability of convenient and sensitive microtiter-plate based assays for the screening and characterization of PBP2a inhibitors is expected to facilitate the discovery and development of new PBP2a inhibitors for use in combating the serious public health problem posed by MRSA.

scholarly journals High-Level Resistance of Staphylococcus aureus to β-Lactam Antibiotics Mediated by Penicillin-Binding Protein 4 (PBP4)

2017 ◽  
Vol 61 (6) ◽  
Author(s):  
Stephanie M. Hamilton ◽  
J. Andrew N. Alexander ◽  
Eun Ju Choo ◽  
Li Basuino ◽  
Thaina M. da Costa ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Binding Protein ◽  
Penicillin Binding Protein ◽  
Content Type ◽  
Native Promoter ◽  
Lactam Antibiotic ◽  
High Level ◽  
Level Resistance

ABSTRACT Penicillin-binding protein 4 (PBP4), a nonessential, low-molecular-weight penicillin-binding protein of Staphylococcus aureus, has been implicated in low-level resistance to β-lactam antibiotics, although the mechanism is unknown. Mutations in PBP4 and its promoter were identified in a laboratory-generated mutant strain, CRB, which expresses high-level resistance to β-lactams, including resistance to the new-generation cephalosporins active against methicillin-resistant strains of S. aureus. These mutations did not appreciably alter the β-lactam antibiotic binding affinity of purified recombinant mutant PBP4 compared to that of wild-type PBP4. Compared to the susceptible parent strain, COLnex, the CRB strain produces a highly cross-linked cell wall peptidoglycan, indicative of increased transpeptidase activity. The pbp4 promoter mutation of CRB was associated with greatly increased amounts of PBP4 in membranes compared to those in the COLnex parent. Replacement of the native promoter of COLnex with the mutant promoter of CRB resulted in increased amounts of PBP4 in membranes and a highly cross-linked cell wall. PBP4 can be repurposed to provide essential transpeptidase activity in vivo and confer high-level resistance to β-lactam antibiotics, such as ceftobiprole and ceftaroline.

scholarly journals β-Lactam Antibiotics Targeting PBP1 Selectively Enhance Daptomycin Activity against Methicillin-Resistant Staphylococcus aureus

2013 ◽  
Vol 57 (10) ◽  
pp. 5005-5012 ◽  
Author(s):  
Andrew D. Berti ◽  
George Sakoulas ◽  
Victor Nizet ◽  
Ryan Tewhey ◽  
Warren E. Rose
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Division ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Binding Protein ◽  
Membrane Insertion ◽  
Penicillin Binding Protein ◽  
Compensatory Response ◽  
Binding Profile ◽  
Methicillin Resistant ◽  
Content Type

ABSTRACTThe activity of daptomycin (DAP) against methicillin-resistantStaphylococcus aureus(MRSA) is enhanced in the presence of subinhibitory concentrations of antistaphylococcal β-lactam antibiotics by an undefined mechanism. Given the variability in the penicillin-binding protein (PBP)-binding profiles of different β-lactam antibiotics, the purpose of this study was to examine the relative enhancement of DAP activity against MRSA by different β-lactam antibiotics to determine if a specific PBP-binding profile is associated with the ability to enhance the anti-MRSA activity of DAP. We determined that both broad- and narrow-spectrum β-lactam antibiotics known to exhibit PBP1 binding demonstrated potent enhancement of DAP anti-MRSA activity, whereas β-lactam antibiotics with minimal PBP1 binding (cefoxitin, ceftriaxone, cefaclor, and cefotaxime) were less effective. We suspect that PBP1 disruption by β-lactam antibiotics affects pathways of cell division inS. aureusthat may be a compensatory response to DAP membrane insertion, resulting in DAP hypersusceptibility.

scholarly journals Genetic Diversity amongStaphylococcus aureusIsolates Showing Oxacillin and/or Cefoxitin Resistance Not Linked to the Presence ofmecGenes

2018 ◽  
Vol 62 (7) ◽  
pp. e00091-18 ◽  
Author(s):  
M. Angeles Argudín ◽  
S. Roisin ◽  
L. Nienhaus ◽  
M. Dodémont ◽  
R. de Mendonça ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Staphylococcus Aureus ◽  
Binding Protein ◽  
Missense Mutations ◽  
Penicillin Binding Protein ◽  
Methicillin Resistant ◽  
Content Type

ABSTRACTMethicillin-resistantStaphylococcus aureusisolates lackingmecgenes (n= 32), collected from Belgian hospitals, were characterized for their β-lactamase production and the presence of mutations inpbpgenes, thepbp4promoter, and genes involved in penicillin-binding protein 4 overproduction (gdpPandyjbH). Twelve isolates were β-lactamase hyperproducers (BHPs), while 12 non-BHP isolates might produce an incomplete GdpP protein. Most isolates showed nucleotide missense mutations inpbpgenes. A few isolates also showed mutations in thepbp4promoter.

scholarly journals Antibiotic Resistance as a Stress Response: Recovery of High-Level Oxacillin Resistance in Methicillin-Resistant Staphylococcus aureus “Auxiliary” (fem) Mutants by Induction of the Stringent Stress Response

2017 ◽  
Vol 61 (8) ◽  
Author(s):  
Choon Keun Kim ◽  
Catarina Milheiriço ◽  
Hermínia de Lencastre ◽  
Alexander Tomasz
Keyword(s):  
Staphylococcus Aureus ◽  
Antibiotic Resistance ◽  
Stress Response ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Penicillin Binding Protein ◽  
Methicillin Resistant ◽  
Content Type ◽  
Oxacillin Resistance ◽  
Response Recovery ◽  
High Level

ABSTRACT Studies with methicillin-resistant Staphylococcus aureus (MRSA) strain COL have shown that the optimal resistance phenotype requires not only mecA but also a large number of “auxiliary genes” identified by Tn551 mutagenesis. The majority of auxiliary mutants showed greatly increased levels of oxacillin resistance when grown in the presence of sub-MICs of mupirocin, suggesting that the mechanism of reduced resistance in the auxiliary mutants involved the interruption of a stringent stress response, causing reduced production of penicillin-binding protein 2A (PBP 2A).

scholarly journals PBP2a Mutations Causing High-Level Ceftaroline Resistance in Clinical Methicillin-Resistant Staphylococcus aureus Isolates

2014 ◽  
Vol 58 (11) ◽  
pp. 6668-6674 ◽  
Author(s):  
S. Wesley Long ◽  
Randall J. Olsen ◽  
Shrenik C. Mehta ◽  
Timothy Palzkill ◽  
Patricia L. Cernoch ◽  
...  
Keyword(s):  
United States ◽  
Staphylococcus Aureus ◽  
Binding Pocket ◽  
The United States ◽  
Penicillin Binding Protein ◽  
Methicillin Resistant ◽  
Content Type ◽  
Mrsa Strains ◽  
Biochemical Analyses ◽  
High Level

ABSTRACTCeftaroline is the first member of a novel class of cephalosporins approved for use in the United States. Although prior studies have identified eight ceftaroline-resistant methicillin-resistantStaphylococcus aureus(MRSA) isolates in Europe and Asia with MICs ranging from 4 to 8 mg/liter, high-level resistance to ceftaroline (>32 mg/liter) has not been described in MRSA strains isolated in the United States. We isolated a ceftaroline-resistant (MIC > 32 mg/liter) MRSA strain from the blood of a cystic fibrosis patient and five MRSA strains from the respiratory tract of this patient. Whole-genome sequencing identified two amino acid-altering mutations uniquely present in the ceftaroline-binding pocket of the transpeptidase region of penicillin-binding protein 2a (PBP2a) in ceftaroline-resistant isolates. Biochemical analyses and the study of isogenic mutant strains confirmed that these changes caused ceftaroline resistance. Thus, we identified the molecular mechanism of ceftaroline resistance in the first MRSA strain with high-level ceftaroline resistance isolated in the United States.

scholarly journals Ceftobiprole- and Ceftaroline-Resistant Methicillin-Resistant Staphylococcus aureus

2015 ◽  
Vol 59 (5) ◽  
pp. 2960-2963 ◽  
Author(s):  
Liana C. Chan ◽  
Li Basuino ◽  
Binh Diep ◽  
Stephanie Hamilton ◽  
Som S. Chatterjee ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Methicillin Resistant ◽  
Content Type ◽  
Low Level ◽  
Mrsa Strains ◽  
High Level ◽  
Level Resistance

ABSTRACTThe role ofmecAmutations in conferring resistance to ceftobiprole and ceftaroline, cephalosporins with anti-methicillin-resistantStaphylococcus aureus(MRSA) activity, was determined with MRSA strains COL and SF8300. The SF8300 ceftaroline-passaged mutant carried a singlemecAmutation, E447K (E-to-K change at position 447), and expressed low-level resistance. This mutation in COL conferred high-level resistance to ceftobiprole but only low-level resistance to ceftaroline. The COL ceftaroline-passaged mutant, which expressed high-level resistance to ceftobiprole and ceftaroline, had mutations inpbp2,pbp4, andgdpPbut notmecA.

scholarly journals Ceftaroline Resistance by Clone-Specific Polymorphism in Penicillin-Binding Protein 2a of Methicillin-Resistant Staphylococcus aureus

2018 ◽  
Vol 62 (9) ◽  
Author(s):  
Hyukmin Lee ◽  
Eun-Jeong Yoon ◽  
Dokyun Kim ◽  
Jung Wook Kim ◽  
Kwang-Jun Lee ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Amino Acid ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Binding Protein ◽  
Amino Acid Substitutions ◽  
Penicillin Binding Protein ◽  
Methicillin Resistant ◽  
Content Type ◽  
Cefoxitin Disk ◽  
Cefoxitin Disk Diffusion

ABSTRACT A total of 281 nonduplicated Staphylococcus aureus blood isolates were collected from January to May 2017 from eight hospitals in South Korea to investigate the epidemiological traits of ceftaroline resistance in methicillin-resistant S. aureus (MRSA). Cefoxitin-disk diffusion tests and the mecA gene PCR revealed that 56.6% (159/281) of the S. aureus isolates were MRSA, and most belonged to ST5 (50.3%, 80/281) and ST72 (41.5%, 66/281). Of the MRSA isolates, 44.0% (70/159) were nonsusceptible to ceftaroline (MIC ≥ 2 mg/liter), whereas all of the methicillin-susceptible S. aureus isolates were susceptible to the drug. Eight amino acid substitutions in penicillin-binding protein 2a (PBP2a), including four (L357I, E447K, I563T, and S649A) in the penicillin-binding domain (PBD) and four (N104K, V117I, N146K, and A228V) in the non-PBD (nPBD) of PBP2a, were associated with ceftaroline resistance. The accumulation of substitutions in PBP2a resulted in the elevation of ceftaroline MICs: one substitution at 1 to 2 mg/liter, two or three substitutions at 2 to 4 mg/liter, and five substitutions at 4 or 16 mg/liter. Ceftaroline resistance in MRSA might be the result of clone-specific PBP2a polymorphism, along with substitutions both in PBD and nPBD, and the elevated ceftaroline MICs were associated with the substitution sites and accumulation of substitutions.

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