scholarly journals Antimicrobial Resistance and Molecular Typing of Neisseria gonorrhoeae Isolates in Kyoto and Osaka, Japan, 2010 to 2012: Intensified Surveillance after Identification of the First Strain (H041) with High-Level Ceftriaxone Resistance

2013 ◽  
Vol 57 (11) ◽  
pp. 5225-5232 ◽  
Author(s):  
Ken Shimuta ◽  
Magnus Unemo ◽  
Shu-ichi Nakayama ◽  
Tomoko Morita-Ishihara ◽  
Misato Dorin ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Neisseria Gonorrhoeae ◽  
Molecular Typing ◽  
Multilocus Sequence Typing ◽  
Binding Protein ◽  
Sporadic Case ◽  
Penicillin G ◽  
Penicillin Binding Protein ◽  
Content Type ◽  
High Level

ABSTRACTIn 2009, the first high-level ceftriaxone-resistantNeisseria gonorrhoeaestrain (H041) was isolated in Kyoto, Japan. The present study describes an intensified surveillance (antimicrobial resistance and molecular typing) ofNeisseria gonorrhoeaeisolates in Kyoto and its neighboring prefecture Osaka, Japan, in 2010 to 2012, which was initiated after the identification of H041. From April 2010 to March 2012, 193N. gonorrhoeaeisolates were collected and the MICs (μg/ml) to six antimicrobials, including ceftriaxone, were determined. All isolates showed susceptibility to ceftriaxone and cefixime (MIC values, <0.5 μg/ml), and spectinomycin. The rates of resistance (intermediate susceptibility) to azithromycin, penicillin G, and ciprofloxacin were 3.6% (19.7%), 24.4% (71.0%), and 78.2% (0.5%), respectively. Multilocus sequence typing (MLST) showed that 40.9%, 19.2%, and 17.1% of isolates belonged to ST1901, ST7359, and ST7363, respectively. Furthermore,N. gonorrhoeaemultiantigen sequence typing (NG-MAST) revealed that 12 (63%) of the 19 isolates with decreased susceptibility to ceftriaxone (MIC > 0.064 μg/ml) were of ST1407. NG-MAST ST1407 was also the most prevalent ST (16.1%; 31 of 193 isolates). In those NG-MAST ST1407 strains, several mosaic typepenAalleles were found, including SF-A type (penicillin binding protein 2 allele XXXIV) and its derivatives. These were confirmed using transformation of thepenAmosaic alleles as critical determinants for enhanced cefixime and ceftriaxone MICs. The intensified surveillance in Kyoto and Osaka, Japan, did not identify any dissemination of the high-level ceftriaxone-resistantN. gonorrhoeaestrain H041, suggesting that H041 might have caused only a sporadic case and has not spread further.

scholarly journals Effect of Variants of Penicillin-Binding Protein 2 on Cephalosporin and Carbapenem Susceptibilities in Neisseria gonorrhoeae

2015 ◽  
Vol 59 (8) ◽  
pp. 5003-5006 ◽  
Author(s):  
Amrita Bharat ◽  
Walter Demczuk ◽  
Irene Martin ◽  
Michael R. Mulvey
Keyword(s):  
Antimicrobial Resistance ◽  
Neisseria Gonorrhoeae ◽  
Binding Protein ◽  
Penicillin Binding Protein ◽  
Wild Type ◽  
Molecular Surveillance ◽  
Content Type ◽  
Extended Spectrum ◽  
The Relationship

ABSTRACTTo characterize the relationship between penicillin-binding protein 2 (PBP2/penA) and susceptibility to extended-spectrum cephalosporins (ESCs) and carbapenem antibiotics, we compared 17 PBP2 variants inNeisseria gonorrhoeae. Nonmosaic and mosaic variants of PBP2 caused decreased susceptibility to ESCs and, to a lesser extent, to carbapenems. An A501P substitution in mosaic XXXIV_A501P conferred decreased susceptibility to ESCs but restored carbapenem susceptibility to wild-type levels. These results could aid the molecular surveillance of antimicrobial resistance to these agents.

scholarly journals Crystal Structures of Penicillin-Binding Protein D2 fromListeria monocytogenesand Structural Basis for Antibiotic Specificity

2018 ◽  
Vol 62 (9) ◽  
Author(s):  
Jae-Hee Jeong ◽  
Hyung Jin Cha ◽  
Yeon-Gil Kim
Keyword(s):  
Crystal Structures ◽  
Bacterial Infections ◽  
Binding Protein ◽  
Bacterial Pathogen ◽  
Penicillin G ◽  
Structural Basis ◽  
Side Chains ◽  
Penicillin Binding Protein ◽  
Content Type ◽  
Highly Correlated

ABSTRACTβ-Lactam antibiotics that inhibit penicillin-binding proteins (PBPs) have been widely used in the treatment of bacterial infections. However, the molecular basis underlying the different inhibitory potencies of β-lactams against specific PBPs is not fully understood. Here, we present the crystal structures of penicillin-binding protein D2 (PBPD2) fromListeria monocytogenes, a Gram-positive foodborne bacterial pathogen that causes listeriosis in humans. The acylated structures in complex with four antibiotics (penicillin G, ampicillin, cefotaxime, and cefuroxime) revealed that the β-lactam core structures were recognized by a common set of residues; however, the R1 side chains of each antibiotic participate in different interactions with PBPD2. In addition, the structural complementarities between the side chains of β-lactams and the enzyme were found to be highly correlated with the relative reactivities of penam or cephem antibiotics against PBPD2. Our study provides the structural basis for the inhibition of PBPD2 by clinically important β-lactam antibiotics that are commonly used in listeriosis treatment. Our findings imply that the modification of β-lactam side chains based on structural complementarity could be useful for the development of potent inhibitors against β-lactam-resistant PBPs.

scholarly journals A Novel Mechanism of High-Level, Broad-Spectrum Antibiotic Resistance Caused by a Single Base Pair Change in Neisseria gonorrhoeae

mBio ◽  
2011 ◽  
Vol 2 (5) ◽  
Author(s):  
Elizabeth A. Ohneck ◽  
Yaramah M. Zalucki ◽  
Paul J. T. Johnson ◽  
Vijaya Dhulipala ◽  
Daniel Golparian ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Antimicrobial Resistance ◽  
Neisseria Gonorrhoeae ◽  
Base Pair ◽  
Health Concern ◽  
Content Type ◽  
Single Base ◽  
Single Base Pair ◽  
Resistant Strains ◽  
High Level

ABSTRACTThe MtrC-MtrD-MtrE multidrug efflux pump ofNeisseria gonorrhoeaeconfers resistance to a diverse array of antimicrobial agents by transporting these toxic compounds out of the gonococcus. Frequently in gonococcal strains, the expression of themtrCDEoperon is differentially regulated by both a repressor, MtrR, and an activator, MtrA. ThemtrRgene lies 250 bp upstream of and is transcribed divergently from themtrCDEoperon. Previous research has shown that mutations in themtrRcoding region and in themtrR-mtrCDEintergenic region increase levels of gonococcal antibiotic resistance andin vivofitness. Recently, a C-to-T transition mutation 120 bp upstream of themtrCstart codon, termedmtr120, was identified in strain MS11 and shown to be sufficient to confer high levels of antimicrobial resistance when introduced into strain FA19. Here we report that this mutation results in a consensus −10 element and that its presence generates a novel promoter formtrCDEtranscription. This newly generated promoter was found to be stronger than the wild-type promoter and does not appear to be subject to MtrR repression or MtrA activation. Although rare, themtr120mutation was identified in an additional clinical isolate during sequence analysis of antibiotic-resistant strains cultured from patients with gonococcal infections. We propose thatcis-acting mutations can develop in gonococci that significantly alter the regulation of themtrCDEoperon and result in increased resistance to antimicrobials.IMPORTANCEGonorrhea is the second most prevalent sexually transmitted bacterial infection and a worldwide public health concern. As there is currently no vaccine againstNeisseria gonorrhoeae, appropriate diagnostics and subsequent antibiotic therapy remain the primary means of infection control. However, the effectiveness of antibiotic treatment is constantly challenged by the emergence of resistant strains, mandating a thorough understanding of resistance mechanisms to aid in the development of new antimicrobial therapies and genetic methods for antimicrobial resistance testing. This study was undertaken to characterize a novel mechanism of antibiotic resistance regulation inN. gonorrhoeae. Here we show that a single base pair mutation generates a second, stronger promoter formtrCDEtranscription that acts independently of the known efflux system regulators and results in high-level antimicrobial resistance.

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 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 New Ceftriaxone- and Multidrug-Resistant Neisseria gonorrhoeae Strain with a Novel MosaicpenAGene Isolated in Japan

2016 ◽  
Vol 60 (7) ◽  
pp. 4339-4341 ◽  
Author(s):  
Shu-ichi Nakayama ◽  
Ken Shimuta ◽  
Kei-ichi Furubayashi ◽  
Takuya Kawahata ◽  
Magnus Unemo ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Binding Protein ◽  
Multidrug Resistant ◽  
Terminal Region ◽  
Penicillin Binding Protein ◽  
Content Type ◽  
Gonorrhoeae Strain ◽  
Resistant Strains

ABSTRACTWe have characterized in detail a new ceftriaxone- and multidrug-resistantNeisseria gonorrhoeaestrain (FC428) isolated in Japan in 2015. FC428 differed from previous ceftriaxone-resistant strains and contained a novel mosaicpenAallele encoding a new mosaic penicillin-binding protein 2 (PBP 2). However, the resistance-determining 3′-terminal region ofpenAwas almost identical to the regions of two previously reported ceftriaxone-resistant strains from Australia and Japan, indicating that both ceftriaxone-resistant strains and conserved ceftriaxone resistance-determining PBP 2 regions might spread.

scholarly journals Whole-Genome Phylogenomic Heterogeneity of Neisseria gonorrhoeae Isolates with Decreased Cephalosporin Susceptibility Collected in Canada between 1989 and 2013

2014 ◽  
Vol 53 (1) ◽  
pp. 191-200 ◽  
Author(s):  
Walter Demczuk ◽  
Tarah Lynch ◽  
Irene Martin ◽  
Gary Van Domselaar ◽  
Morag Graham ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Large Scale ◽  
Epidemiological Studies ◽  
23S Rrna ◽  
Genome Comparison ◽  
Whole Genome ◽  
Content Type ◽  
Genomic Epidemiology ◽  
High Level ◽  
Sequence Types

A large-scale, whole-genome comparison of CanadianNeisseria gonorrhoeaeisolates with high-level cephalosporin MICs was used to demonstrate a genomic epidemiology approach to investigate strain relatedness and dynamics. Although current typing methods have been very successful in tracing short-chain transmission of gonorrheal disease, investigating the temporal evolutionary relationships and geographical dissemination of highly clonal lineages requires enhanced resolution only available through whole-genome sequencing (WGS). Phylogenomic cluster analysis grouped 169 Canadian strains into 12 distinct clades. While someN. gonorrhoeaemultiantigen sequence types (NG-MAST) agreed with specific phylogenomic clades or subclades, other sequence types (ST) and closely related groups of ST were widely distributed among clades. Decreased susceptibility to extended-spectrum cephalosporins (ESC-DS) emerged among a group of diverse strains in Canada during the 1990s with a variety of nonmosaicpenAalleles, followed in 2000/2001 with thepenAmosaic X allele and then in 2007 with ST1407 strains with thepenAmosaic XXXIV allele. Five genetically distinct ESC-DS lineages were associated withpenAmosaic X, XXXV, and XXXIV alleles and nonmosaic XII and XIII alleles. ESC-DS with coresistance to azithromycin was observed in 5 strains with 23S rRNA C2599T or A2143G mutations. As the costs associated with WGS decline and analysis tools are streamlined, WGS can provide a more thorough understanding of strain dynamics, facilitate epidemiological studies to better resolve social networks, and improve surveillance to optimize treatment for gonorrheal infections.

scholarly journals Mosaic-Like Structure of Penicillin-Binding Protein 2 Gene (penA) in Clinical Isolates of Neisseria gonorrhoeae with Reduced Susceptibility to Cefixime

2002 ◽  
Vol 46 (12) ◽  
pp. 3744-3749 ◽  
Author(s):  
Satoshi Ameyama ◽  
Shoichi Onodera ◽  
Masahiro Takahata ◽  
Shinzaburo Minami ◽  
Nobuko Maki ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Neisseria Gonorrhoeae ◽  
Neisseria Meningitidis ◽  
Binding Protein ◽  
Clinical Isolates ◽  
Penicillin Binding Protein ◽  
Gene Sequences ◽  
Neisseria Flavescens ◽  
Neisseria Perflava

ABSTRACT Neisseria gonorrhoeae strains with reduced susceptibility to cefixime (MICs, 0.25 to 0.5 μg/ml) were isolated from male urethritis patients in Tokyo, Japan, in 2000 and 2001. The resistance to cephems including cefixime and penicillin was transferred to a susceptible recipient, N. gonorrhoeae ATCC 19424, by transformation of the penicillin-binding protein 2 gene (penA) that had been amplified by PCR from a strain with reduced susceptibility to cefixime (MIC, 0.5 μg/ml). The sequences of penA in the strains with reduced susceptibilities to cefixime were different from those of other susceptible isolates and did not correspond to the reported N. gonorrhoeae penA gene sequences. Some regions in the transpeptidase-encoding domain in this penA gene were similar to those in the penA genes of Neisseria perflava (N. sicca), Neisseria cinerea, Neisseria flavescens, and Neisseria meningitidis. These results showed that a mosaic-like structure in the penA gene conferred reductions in the levels of susceptibility of N. gonorrhoeae to cephems and penicillin in a manner similar to that found for N. meningitidis and Streptococcus pneumoniae.

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