PBP4-mediated β-lactam resistance among clinical strains of <i>Staphylococcus aureus</i>

Satishkumar, Nidhi; Alexander, J. Andrew N.; Poon, Raymond; Buggeln, Emma; Argudín, Maria Ángeles; Strynadka, N.C.J.; Chatterjee, Som S.

doi:10.1093/jac/dkab201

Cited by 7 publications

(18 citation statements)

References 14 publications

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“…The wild-type strain as well as the strains with promoter-associated mutations all had similar growth patterns in absence of β-lactams (Figure 1f). Taken together, these results suggested that promoterassociated mutations led to PBP4 overexpression and β-lactam resistance in MRSA background, similar to what was previously demonstrated in MSSA background strains (7,13).…”

Section: Pbp4 Promoter-associated Mutations Led To Increased Pbp4 Exp...supporting

confidence: 88%

“…These studies were performed by introducing promoter-associated mutations into strains devoid of mecA . Using allelic replacement, we created isogenic mutants by introducing three different, previously detected mutations in the pbp4 promoter region of SF8300 (13, 17). Of these, two mutations were detected in laboratory-generated resistant strains as a result of a passaging experiment.…”

Section: Resultsmentioning

confidence: 99%

“…Similarly, insertion of mutations detected in the strain CRT (a thymine insertion 377 bp upstream the start codon and a 90 bp deletion 275 bp upstream the start codon) into the pbp4 -promoter region of SF8300 produced SF8300 P pbp4 * (CRT). The third mutation was one detected in a clinically-isolated strain (10, 13), namely Strain 1, and consisted of a T to A substitution 266 bp upstream the start codon. The introduction of this mutation in SF8300 gave rise to SF8300 P pbp4 * (Strain 1) (Figure 1a) .…”

Section: Resultsmentioning

confidence: 99%

“…pbp4 promoter-associated mutations that were detected in a laboratory passaged, resistant strain that was devoid of mecA (namely CRB) and in a resistant strain containing mecA (namely CRT) were introduced in SF8300 (6). Further, a pbp4 promoter-associated mutation detected in a β-lactam resistant, clinical isolate (namely Strain 1) was also used in this study (13). Using these promoter mutants, we demonstrated that pbp4 promoter-associated mutations leads to PBP4 overexpression and increased β-lactam resistance in MRSA strains in a mechanism similar to what was previously shown in MSSA strains (7, 13).…”

Section: Introductionmentioning

confidence: 99%

“…Further, a pbp4 promoter-associated mutation detected in a β-lactam resistant, clinical isolate (namely Strain 1) was also used in this study (13). Using these promoter mutants, we demonstrated that pbp4 promoter-associated mutations leads to PBP4 overexpression and increased β-lactam resistance in MRSA strains in a mechanism similar to what was previously shown in MSSA strains (7, 13).…”

Section: Introductionmentioning

confidence: 99%

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A non-classical mechanism of β-lactam resistance in Methicillin-Resistant Staphylococcus aureus (MRSA) and its effect on virulence

Satishkumar

Lai

Mukkayyan

et al. 2022

Preprint

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Methicillin-Resistant Staphylococcus aureus (MRSA) are pathogenic bacteria that are infamously resistant to β-lactam antibiotics, a property attributed to the mecA gene. Recent studies have reported that mutations associated with the promoter region of pbp4 demonstrated high levels of β-lactam resistance, suggesting the role of PBP4 as an important non-mecA mediator of β-lactam resistance. The pbp4 promoter-associated mutations have been detected in strains with or without mecA. Our previous studies that were carried out in strains devoid of mecA described that pbp4 promoter-associated mutations lead to PBP4 overexpression and β-lactam resistance. In this study, by introducing various pbp4 promoter-associated mutations in the genome of an MRSA strain, we demonstrate that PBP4 overexpression can supplement mecA-associated resistance in S. aureus and can lead to increased β-lactam resistance. The promoter and regulatory region of pbp4 is shared with a divergently transcribed gene, abcA, which encodes for a multidrug exporter. We demonstrate that the promoter mutations caused an upregulation of pbp4 and downregulation of abcA, confirming that the resistant phenotype is associated with PBP4 overexpression only. PBP4 has also been associated with staphylococcal pathogenesis, however, its exact role remains unclear. Using a C. elegans model, we demonstrate that strains having increased PBP4 expression are less virulent compared to wild-type strains, suggesting that β-lactam resistance mediated via PBP4 likely comes at the cost of virulence.ImportanceOur study demonstrates the ability of PBP4 to be an important mediator of β-lactam resistance in not only Methicillin-susceptible Staphylococcus aureus (MSSA) background strains as previously demonstrated, but also in MRSA strains. When present together, PBP2a and PBP4 overexpression can produce increased levels of β-lactam resistance, causing complications in treatment. Thus, this study suggests the importance of monitoring PBP4-associated resistance in clinical settings, as well as understanding the mechanistic basis of associated resistance, so that treatments targeting PBP4 may be developed. This study also demonstrates that S. aureus strains with increased PBP4 expression are less pathogenic, providing important hints about the role of PBP4 in S. aureus resistance and pathogenesis.

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Section: Pbp4 Promoter-associated Mutations Led To Increased Pbp4 Exp...supporting

confidence: 88%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

A non-classical mechanism of β-lactam resistance in Methicillin-Resistant Staphylococcus aureus (MRSA) and its effect on virulence

Satishkumar

Lai

Mukkayyan

et al. 2022

Preprint

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Adaptive and maladaptive consequences of deregulation in a bacterial gene regulatory network

Vinchhi

Yelpure

Balachandran

et al. 2023

Preprint

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The archetypal PhoQP two-component system from Enterobacteria regulates crucial pathways like magnesium homeostasis in Escherichia coli and virulence factor expression in Salmonella enterica. Previously we had reported that a laboratory strain of E. coli rapidly accumulated loss-of-function mutations in the mgrB gene, a negative feedback regulator of PhoQP, when evolved in the presence of the antibiotic trimethoprim (Patel and Matange, eLife 2021). Hyperactive PhoQP enhanced the expression of dihydrofolate reductase, target of trimethoprim, resulting in antibiotic tolerance. Here we ask, firstly, how important are mutations in mgrB for the evolution of trimethoprim resistance? We report that MgrB loss, though itself only mildly beneficial, facilitates the evolution of resistance by enhancing the fixation probability of trimethoprim-resistant bacteria under selection. Secondly, we investigate why negative feedback may be needed in the PhoQP system. We show that under drug-free conditions MgrB is required to mitigate the fitness costs of pervasive gene dysregulation by hyperactive PhoQP. Using RNA-seq transcriptomics and genetic analyses, we demonstrate that PhoQP-hyperactivation perturbs the balance of RpoS and RpoD regulated transcriptional programs, and spontaneous mutations in rpoS rectify this imbalance. We propose that relative adaptive and maladaptive consequences of deregulation explain the evolution of negative feedback in bacterial gene regulatory networks.

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Four chromosomally clustered class-C β-lactamases (blaAmpH1/2/3/4) control the cephalosporin hydrolysis inMycobacterium tuberculosisand similar genetic loci appeared as pseudogenes inMycobacterium leprae

Chakraborty

2023

Preprint

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Mycobacterium tuberculosis (Mt) chromosomal ten PBPs and one class-A beta-lactamase (blaC) were implicated in multi-drug resistance against penicillin, cephalosporin and carbapenem drugs. The E. coli Class-A PBP1A and PBP1B proteins referred as PonA1 and PonA2 in Mt with 34 percent homologies whereas Class-B PBP2 referred as PbpA and PbpB sub-class proteins with 27 percent homologies. The class-C PBPs were, PBP4, MecA_N, DacB1, DacB2 and AmpH1-AmpH4. During database search, we found that many Mt Pbp4 had no homology to E. coli PBP4 and such enzyme were found to be AmpH class with AmpC, ACC, DHA and GES beta-lactamases similarities although one of such enzymes suggested as AmpH but the other referred as DacB or PBP5 in the literature. We investigated the Mt whole genomes (accession nos. AL123456, CP001642, CP054013, CP001641, CP025597) to get authentic PBP4 in Mt strain FDAARGOS_757 genome (protein id. AUP69687, 34% homology to E. coli PBP4) which was designated as conserved protein in chromosomes of Mt strains H37Rv, H37Rv-1, GG-36-11 and CCDC5180 making confusion in data analysis. Similarly, we BLASTP homology searched with plasmid-mediated 24 beta-lactamases suggested that four blaAmpH genes, (AmpH1, AmpH2, AmpH3 and AmpH4) predominantly control the degradation of cephalosporins in M. tuberculosis but such genes were found as pseudogenes in M. leprae. The Mt AmpH1/2/3/4 enzymes had better homologies with V. parahaemolyticus and Yersinia pekkanenii AmpH enzyme as well as with E. coli AmpH enzyme. The DacB1 (PBP6) and DacB2 (PBP7) enzymes had blaTEM similarity but no AmpH similarity indicating blaC and DacB1/B2 controlled the Penicillin hydrolysis in Mt. The blaC enzyme had 30% homology to S. aureus blaZ beta-lactamase but such enzyme was also missing in M. leprae. Homology search suggested that carbapenem hydrolysis by blaOXA-23/51-like PBPA/B enzymes and blaOXA-58 related MecA_N domain beta-lactamase which has 21 percent similarity to S. aureus mecA enzyme. The PonA1/A2 had no homology to 24 classes of beta-lactamases but still were popular enzymes implicated for better penicillin hydrolysis. We designed primers for Mt PBPs to check transcription of individual genes by RT-PCR as well as to check chromosomal locus by BLASTN search after WGS. The E. coli genome had no similarities to blaAmpH1/2/3/4 genes but a 17nt (CCTTGGTGCCGTCGACC) sequence found in pKEC-a3c plasmid of C. fruendii with homology to blaAmpH4 gene (nt. 1236-1252) and shared a homology with IS1182-like ISKpn6 transposase of plasmids. The oligonucleotides selected many Mt chromosomes and located conserved blaAmpH4 protein in all cases. However, D435E and F495V mutations in the Mt strain 5521 blaAmpH4 were evident and frameshift deletions located in Mt stain FDAARGOS_756 blaAmpH4 gene with no protein was made. Thus, mutations, deletions and rearrangements mediated by IS-elements were the driving force to make new AmpH genes in M. tuberculosis.

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PBP4-mediated β-lactam resistance among clinical strains of Staphylococcus aureus

Cited by 7 publications

References 14 publications

A non-classical mechanism of β-lactam resistance in Methicillin-Resistant Staphylococcus aureus (MRSA) and its effect on virulence

A non-classical mechanism of β-lactam resistance in Methicillin-Resistant Staphylococcus aureus (MRSA) and its effect on virulence

Adaptive and maladaptive consequences of deregulation in a bacterial gene regulatory network

Four chromosomally clustered class-C β-lactamases (blaAmpH1/2/3/4) control the cephalosporin hydrolysis inMycobacterium tuberculosisand similar genetic loci appeared as pseudogenes inMycobacterium leprae

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