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With the widespread use of antibiotics, antimicrobial resistance (AMR) has become a global problem that endangers public health. Despite the global high prevalence of group A Streptococcus (GAS) infections and the global widespread use of b-lactams, b-lactams remain the first-line treatment option for GAS infection. b-hemolytic streptococci maintain a persistent susceptibility to b-lactams, which is an extremely special phenomenon in the genus Streptococci, while the exact current mechanism is not known. In recent years, several studies have found that the gene encoding penicillin binding protein 2X (pbp2x) is associated with GAS with reduced-b-lactam susceptibility. The purpose of this review is to summarize the current published data on GAS penicillin binding proteins and b-lactam susceptibility, to explore the relationship between them, and to be alert to the emergence of GAS with reduced susceptibility to b-lactams. [ABSTRACT FROM AUTHOR]

With the widespread use of antibiotics, antimicrobial resistance (AMR) has become a global problem that endangers public health. Despite the global high prevalence of group A Streptococcus (GAS) infections and the global widespread use of β-lactams, β-lactams remain the first-line treatment option for GAS infection. β-hemolytic streptococci maintain a persistent susceptibility to β-lactams, which is an extremely special phenomenon in the genus Streptococci, while the exact current mechanism is not known. In recent years, several studies have found that the gene encoding penicillin binding protein 2X (pbp2x) is associated with GAS with reduced-β-lactam susceptibility. The purpose of this review is to summarize the current published data on GAS penicillin binding proteins and β-lactam susceptibility, to explore the relationship between them, and to be alert to the emergence of GAS with reduced susceptibility to β-lactams.

Theoretically, sandwich immunoassay is more sensitive and has a wider working range than that of competitive format. However, it has been thought that small molecules cannot be detected by the sandwich format due to their limited size. In the present study, we proposed a novel strategy for achieving sandwich immunoassay of β-lactams with low molecular weights. Firstly, five β-lactam antibiotics were selected to bind with penicillin binding protein (PBP)2x* to form complexes. Then, monoclonal and polyclonal antibodies against PBP2x*-β-lactams complexes were produced by animal immunization. Subsequently, the optimal pairing antibodies were utilized to establish sandwich immunoassay for detection of 18 PBP2x*-β-lactam complexes. Among them, ceftriaxone could be detected at as low as 1.65 ng/mL with working range of 1–1000 ng/mL in milk. To reveal the detection mechanism, computational chemistry and molecular recognition study were carried out. The results showed that β-lactams with a large size and complex structures maybe conducive to induce conformational changes of PBP2x*, and then exhibit greater possibility of being detected by sandwich immunoassay after combination with PBP2x*. This study provides insights for subsequent investigations of anti-metatype antibody screening and sandwich immunoassay establishment for small-molecule detection.

Infectious diseases have been one of the most important problems affecting all societies since the existence of mankind. Streptococci, one of the causes of these diseases, can be found in the normal flora of the mouth, nose, throat, skin, digestion and genital system in humans. Streptococci are an important cause of diseases in humans such as meningitis, acute otitis media, rhinosinusitis and community -acquired pneumonia. In this study, various clinical samples belonging to the disease were taken between January and September 2015. All necessary permission documents and the ethics committee approval numbered 2 and dated 13.11.2014 for clinical investigations were obtained. The collected samples were brought to the Microbiology Laboratory of Van Yüzüncü Yıl University Medical Faculty. Analyzes of determined and identified antibiotic resistance genes of isolated and identified streptococci strains were carried out in the Microbiology laboratory of Yüzüncü Yıl University Faculty of Pharmacy by PCR. The detection of pbp1a for penicillin G antibiotic, pbp2x for cefotaxime antibiotic, and gyrA resistance genes for quinolone antibiotics were targeted. In this study, the primers used in Table -2 were taken as reference. A total of 30 streptococcus strains were isolated and identified from various clinical samples including 8 sputum, 6 urine, 5 BOS, 4 blood, 4 nasopharyngeal swab, 2 ear effusion and 1 abscess. As a result of the PCR analysis, the resistance of pbp1a was found in 6 of the streptococcal isolates, pbp2x in 8, and resistance in the gyrA gene region in 5. In our study, no three resistance genes were found in 22 isolates and the presence of all three (pbp1a, pbp2x and gyrA) resistance genes in 5 isolates was determined. It is expected that the data obtained from this research will contribute to national and international knowledge accumulation. [ABSTRACT FROM AUTHOR]

With the widespread use of antibiotics, antimicrobial resistance (AMR) has become a global problem that endangers public health. Despite the global high prevalence of group A Streptococcus (GAS) infections and the global widespread use of β-lactams, β-lactams remain the first-line treatment option for GAS infection. β-hemolytic streptococci maintain a persistent susceptibility to β-lactams, which is an extremely special phenomenon in the genus Streptococci , while the exact current mechanism is not known. In recent years, several studies have found that the gene encoding penicillin binding protein 2X ( pbp2x ) is associated with GAS with reduced-β-lactam susceptibility. The purpose of this review is to summarize the current published data on GAS penicillin binding proteins and β-lactam susceptibility, to explore the relationship between them, and to be alert to the emergence of GAS with reduced susceptibility to β-lactams., Competing Interests: The authors indicated that this study was conducted without any commercial or financial relationships that could be interpreted as potential conflicts of interest., (Copyright © 2023 Yu, Guo, Zheng and Yang.)

Abstract: Mutations in the transpeptidase domain of penicillin-binding protein 2x (PBP2x) of Streptococcus pneumoniae that reduce the affinity to beta-lactams are important determinants of resistance to these antibiotics. We have now analyzed in vitro and in vivo properties of PBP2x variants from cefotaxime-resistant laboratory mutants and a clinical isolate. The patterns of two to four resistance-specific mutations present in each of the proteins, all of which are placed between 6.6 and 24 Å around the active site, fall into three categories according to their positions in the three-dimensional structure. The first PBP2x group is characterized by mutations at the end of helix α11 and carries the well-known T550A change and/or one mutation on the surface of the penicillin-binding domain in close contact with the C-terminal domain. All group I proteins display very low acylation efficiencies, ≤1700 M−1 s−1, for cefotaxime. The second class represented by PBP2x of the mutant C505 shows acylation efficiencies below 100 M−1 s−1 for both cefotaxime and benzylpenicillin and contains the mutation L403F at a critical site close to the active serine. PBP2x of the clinical isolate 669 reveals a third mutational pathway where at least the two mutations Q552E and S389L are important for resistance, and acylation efficiency is reduced for both beta-lactams to around 10,000 M−1 s−1. In each group, at least one mutation is located in close vicinity to the active site and mediates a resistance phenotype in vivo alone, whereas other mutations might exhibit secondary effects only in context with other alterations. [Copyright &y& Elsevier]

Theoretically, sandwich immunoassay is more sensitive and has a wider working range than that of competitive format. However, it has been thought that small molecules cannot be detected by the sandwich format due to their limited size. In the present study, we proposed a novel strategy for achieving sandwich immunoassay of β-lactams with low molecular weights. Firstly, five β-lactam antibiotics were selected to bind with penicillin binding protein (PBP)2x* to form complexes. Then, monoclonal and polyclonal antibodies against PBP2x*-β-lactams complexes were produced by animal immunization. Subsequently, the optimal pairing antibodies were utilized to establish sandwich immunoassay for detection of 18 PBP2x*-β-lactam complexes. Among them, ceftriaxone could be detected at as low as 1.65 ng/mL with working range of 1–1000 ng/mL in milk. To reveal the detection mechanism, computational chemistry and molecular recognition study were carried out. The results showed that β-lactams with a large size and complex structures maybe conducive to induce conformational changes of PBP2x*, and then exhibit greater possibility of being detected by sandwich immunoassay after combination with PBP2x*. This study provides insights for subsequent investigations of anti-metatype antibody screening and sandwich immunoassay establishment for small-molecule detection. [ABSTRACT FROM AUTHOR]

Since the introduction of pneumococcal conjugate vaccines, the prevalence of non-meropenem-susceptible pneumococci has been increasing in Japan. In an earlier study, we demonstrated that multidrug-resistant serotype 15A-ST63 in Japan has a specific pbp1a sequence ( pbp1a -13) that could promote meropenem resistance. To trace the origin of pbp1a , we analyzed isolates of serotype 19A-CC3111, which is the most prevalent non-meropenem-susceptible clone in Japan. We analyzed a total of 119 serotype 19A-CC3111 strains recovered in Japan using whole-genome sequencing. Of the 119 isolates, 53 (44.5%) harbored pbp1a -13, indicating that the clone may be the primary reservoir of the pbp1a type and that the pbp1a region may be horizontally transferred between different serotype strains. The single acquisition of pbp1a -13 seemed to cause only penicillin resistance and not multidrug resistance; a combination of penicillin-binding protein (PBP) recombination in the pbp2b and/or pbp2x region(s) with acquisition of pbp1a -13 caused multidrug resistance. Conserved amino acid motif analysis suggested that the pbp1a 370SXXK, pbp2b 448SXN, and pbp2x 337SXXN motifs were the candidates for amino acid substitutions increasing the MICs of meropenem, cefotaxime, and penicillin. We identified a specific clone that was correlated with multidrug resistance, although no correlation was observed between phylogenetic trees generated using core genomes and those generated with only the cps locus. All tested isolates were highly erythromycin resistant, and most harbored mefE within macrolide efflux genetic assembly (MEGA) elements and ermB within Tn 917 , which was inserted within Tn 916 and exhibited a structure identical to that of Tn 2017 ., (Copyright © 2019 American Society for Microbiology.)

The Streptococcus pneumoniae clone Hungary 19A -6 expresses unusually high levels of β-lactam resistance, which is in part due to mutations in the MurM gene, encoding a transferase involved in the synthesis of branched peptidoglycan. Moreover, it contains the allele ciaH232 , encoding the histidine kinase CiaH (M. Müller, P. Marx, R. Hakenbeck, and R. Brückner, Microbiology 157:3104-3112, 2011, https://doi.org/10.1099/mic.0.053157-0). High-level penicillin resistance primarily requires the presence of low-affinity (mosaic) penicillin binding protein (PBP) genes, as, for example, in strain Hu17, a closely related member of the Hungary 19A -6 lineage. Interestingly, strain Hu15 is β-lactam sensitive due to the absence of mosaic PBPs. This unique situation prompted us to investigate the development of cefotaxime resistance in transformation experiments with genes known to play a role in this phenotype, pbp2x , pbp1a , murM , and ciaH , and penicillin-sensitive recipient strains R6 and Hu15. Characterization of phenotypes, peptidoglycan composition, and CiaR-mediated gene expression revealed several novel aspects of penicillin resistance. The murM gene of strain Hu17 ( murM Hu17 ), which is highly similar to murM of Streptococcus mitis , induced morphological changes which were partly reversed by ciaH232. murM Hu17 conferred cefotaxime resistance only in the presence of the pbp2x o f strain Hu17 ( pbp2x Hu17 ). The ciaH232 allele contributed to a remarkable increase in cefotaxime resistance in combination with pbp2x Hu17 and pbp1a of strain Hu17 ( pbp1a Hu17 ), accompanied by higher levels of expression of CiaR-regulated genes, documenting that ciaH232 responds to PBP1a Hu17 -mediated changes in cell wall synthesis. Most importantly, the proportion of branched peptides relative to the proportion of linear muropeptides increased in cells containing mosaic PBPs, suggesting an altered enzymatic activity of these proteins., (Copyright © 2017 Schweizer et al.)

The identification of commensal streptococci species is an everlasting problem due to their ability to genetically transform. A new challenge in this respect is the recent description of Streptococcus pseudopneumoniae as a new species, which was distinguished from closely related pathogenic S. pneumoniae and commensal S. mitis by a variety of physiological and molecular biological tests. Forty-one atypical S. pneumoniae isolates have been collected at the German National Reference Center for Streptococci (GNRCS). Multilocus sequence typing (MLST) confirmed 35 isolates as the species S. pseudopneumoniae A comparison with the pbp2x sequences from 120 commensal streptococci isolated from different continents revealed that pbp2x is distinct among penicillin-susceptible S. pseudopneumoniae isolates. Four penicillin-binding protein x (PBPx) alleles of penicillin-sensitive S. mitis account for most of the diverse sequence blocks in resistant S. pseudopneumoniae , S. pneumoniae , and S. mitis , and S. infantis and S. oralis sequences were found in S. pneumoniae from Japan. PBP2x genes of the family of mosaic genes related to pbp2x in the S. pneumoniae clone Spain 23F -1 were observed in S. oralis and S. infantis as well, confirming its global distribution. Thirty-eight sites were altered within the PBP2x transpeptidase domains of penicillin-resistant strains, excluding another 37 sites present in the reference genes of sensitive strains. Specific mutational patterns were detected depending on the parental sequence blocks, in agreement with distinct mutational pathways during the development of beta-lactam resistance. The majority of the mutations clustered around the active site, whereas others are likely to affect stability or interactions with the C-terminal domain or partner proteins., (Copyright © 2017 van der Linden et al.)

Background: β-lactams resistant Streptococcus pneumoniae are an emerging problem throughout the world. Several resistance mechanisms have been reported, including expression of drug-destroying enzymes such as β-lactamases, altered drug targets such as conformational changes in PBPs, decreased bacterial permeability, and increased drug efflux., Objectives: The present study aimed to determine the relationship between the results of polymerase chain reaction identification of the Pbp1a, Pbp2b and Pbp2x genes (penicillin-binding proteins) and susceptibilities of β-lactam antibiotics against S. pneumoniae., Materials and Methods: Fifty five isolates of S. pneumoniae were obtained from clinical samples with antimicrobial tests. The susceptibilities of isolates to benzylpenicillin, imipenem, oxacillin, ceftazidime were determined. The resistance genotype was determined by the polymerase chain reaction with primers designed for the PBP genes., Results: The number of S. pneumoniae isolates resistant to benzylpenicillin, imipenem, oxacillin and ceftazidime were 94.5%, 100%, 100%, and 21.8%, respectively. Analysis of mutation in the genes for pbp showed that 85% of isolates had mutations in pbp2x, pbp2b and pbp1a. Susceptibility to benzylpenicillin was decreased once the number of mutated pbp genes in S. pneumonia increased. According to the results of this study, S. pneumoniae isolates showed reduced susceptibility due to accumulation of resistance genes., Conclusions: We suggest that studies should be performed to evaluate changes in Minimum Inhibitory Concentration (MIC) values as well as genetic mutations in order to determine prevalence of S. pneumoniae resistance against antimicrobial agents.