Clinical isolates of Streptococcus mitis/oralis-related species with reduced carbapenem susceptibility, harboring amino acid substitutions in penicillin-binding proteins in Japan.

Streptococcus mitis/oralis group isolates with reduced carbapenem susceptibility have been reported, but its isolation rate in Japan is unknown. We collected 356 clinical α-hemolytic streptococcal isolates and identified 142 of them as S. mitis/oralis using partial sodA sequencing. The rate of meropenem non-susceptibility was 17.6% (25/142). All 25 carbapenem-non-susceptible isolates harbored amino acid substitutions in/near the conserved motifs in PBP1A, PBP2B, and PBP2X. Carbapenem non-susceptibility is common among S. mitis/oralis group isolates in Japan.

Hydroxyhexylitaconic acids as potent IMP-type metallo-ß-lactamase inhibitors for controlling carbapenem resistance in Enterobacterales.

Metallo-ß-lactamases (MBLs) represent one of the main causes of carbapenem resistance in the order Enterobacterales. To combat MBL-producing carbapenem-resistant Enterobacterales, the development of MBL inhibitors can restore carbapenem efficacy for such resistant bacteria. Microbial natural products are a promising source of attractive seed compounds for the development of antimicrobial agents. Here, we report that hydroxyhexylitaconic acids (HHIAs) produced by a member of the genus Aspergillus can suppress carbapenem resistance conferred by MBLs, particularly IMP (imipenemase)-type MBLs. HHIAs were found to be competitive inhibitors with micromolar orders of magnitude against IMP-1 and showed weak inhibitory activity toward VIM-2, while no inhibitory activity against NDM-1 was observed despite the high dosage. The elongated methylene chains of HHIAs seem to play a crucial role in exerting inhibitory activity because itaconic acid, a structural analog without long methylene chains, did not show inhibitory activity against IMP-1. The addition of HHIAs restored meropenem and imipenem efficacy to satisfactory clinical levels against IMP-type MBL-producing Escherichia coli and Klebsiella pneumoniae clinical isolates. Unlike EDTA and Aspergillomarasmine A, HHIAs did not cause the loss of zinc ions from the active site, resulting in the structural instability of MBLs. X-ray crystallography and in silico docking simulation analyses revealed that two neighboring carboxylates of HHIAs coordinated with two zinc ions in the active sites of VIM-2 and IMP-1, which formed a key interaction observed in MBL inhibitors. Our results indicated that HHIAs are promising for initiating the design of potent inhibitors of IMP-type MBLs.IMPORTANCEThe number and type of metallo-ß-lactamase (MΒL) are increasing over time. Carbapenem resistance conferred by MΒL is a significant threat to our antibiotic regimen, and the development of MΒL inhibitors is urgently required to restore carbapenem efficacy. Microbial natural products have served as important sources for developing antimicrobial agents targeting pathogenic bacteria since the discovery of antibiotics in the mid-20th century. MΒL inhibitors derived from microbial natural products are still rare compared to those derived from chemical compound libraries. Hydroxyhexylitaconic acids (HHIAs) produced by members of the genus Aspergillus have potent inhibitory activity against clinically relevant IMP-type MBL. HHIAs may be good lead compounds for the development of MBL inhibitors applicable for controlling carbapenem resistance in IMP-type MBL-producing Enterobacterales.

ß-Lactam Susceptibility of Streptococcus dysgalactiae subsp. equisimilis.

All clinical isolates of Streptococcus dysgalactiae subsp. equisimilis (SDSE) are considered susceptible to ß-lactams, the first-line drugs used for SDSE infections. However, penicillin-non-susceptible SDSE has been reported from Denmark. In this study, we attempted to detect ß-lactam-non-susceptible clinical isolates of SDSE in Japan. One hundred and fifty clinical isolates of S. dysgalactiae were collected in 2018, and species identification was performed using Rapid ID Strep API. The minimum inhibitory concentrations (MIC) of six ß-lactams (penicillin G, oxacillin, ceftizoxime, ceftibuten, cefoxitin, and cefaclor) were determined for 85 clinical isolates of SDSE using the agar dilution method standardized by the Clinical Laboratory Standards Institute. For the 85 isolates identified as SDSE, the MIC ranges of penicillin G, oxacillin, ceftizoxime, ceftibuten, cefoxitin, and cefaclor were 0.007-0.06, 0.03-0.12, 0.015-0.06, 0.25-2, 0.12-2, and 0.06-0.5 µg/mL, respectively. None of the clinical isolates were non-susceptible to penicillin G, indicating that all 85 clinical isolates of SDSE were susceptible to ß-lactams. Our findings indicate that almost all clinical isolates of SDSE in several prefectures of Japan remain susceptible to ß-lactams. Nevertheless, there remains a need for continuous and careful monitoring of drug susceptibility among clinical isolates of SDSE in Japan.

Improved disk diffusion method for simple detection of group B streptococci with reduced penicillin susceptibility (PRGBS).

We used 73 group B Streptococcus with reduced penicillin susceptibility (PRGBS) isolates and determined more rational cutoff values of previously developed disk diffusion method for detecting PRGBS using oxacillin, ceftizoxime, and ceftibuten disks. Using the novel cutoff values, the three disks showed high sensitivity and specificity, which were above 90.0%.

Difference in the Inhibitory Effect of Thiol Compounds and Demetallation Rates from the Zn(II) Active Site of Metallo-ß-lactamases (IMP-1 and IMP-6) Associated with a Single Amino Acid Substitution.

Gram-negative bacteria producing metallo-ß-lactamases (MBLs) have become a considerable threat to public health. MBLs including the IMP, VIM, and NDM types are Zn(II) enzymes that hydrolyze the ß-lactam ring present in a broad range of antibiotics, such as N-benzylpenicillin, meropenem, and imipenem. Among IMPs, IMP-1 and IMP-6 differ in a single amino acid substitution at position 262, where serine in IMP-1 is replaced by glycine in IMP-6, conferring a change in substrate specificity. To investigate how this mutation influences enzyme function, we examined lactamase inhibition by thiol compounds. Ethyl 3-mercaptopropionate acted as a competitive inhibitor of IMP-1, but a noncompetitive inhibitor of IMP-6. A comparison of the crystal structures previously reported for IMP-1 (PDB code: 5EV6) and IMP-6 (PDB code: 6LVJ) revealed a hydrogen bond between the side chain of Ser262 and Cys221 in IMP-1 but the absence of hydrogen bond in IMP-6, which affects the Zn2 coordination sphere in its active site. We investigated the demetallation rates of IMP-1 and IMP-6 in the presence of chelating agent ethylenediaminetetraacetic acid (EDTA) and found that the demetallation reactions had fast and slow phases with a first-order rate constant (kfast = 1.76 h-1, kslow = 0.108 h-1 for IMP-1, and kfast = 14.0 h-1 and kslow = 1.66 h-1 for IMP-6). The difference in the flexibility of the Zn2 coordination sphere between IMP-1 and IMP-6 may influence the demetallation rate, the catalytic efficiency against ß-lactam antibiotics, and the inhibitory effect of thiol compounds.

Bird's-eye MApping of plasmids (BeMAp) for visualization and comparison of genomic structures of different plasmids by mapping antimicrobial resistance genes on spreadsheets.

Effective classification and visualization of multiple antimicrobial resistance plasmids can be challenging, and few tools to analyze similarities among plasmids depending on the location of genes are available. We created a new plasmid mapping program called Bird's-eye MApping of plasmids (BeMAp) to map antimicrobial resistance genes across multiple plasmids onto a spreadsheet and visualize their similarities based on gene types, locations, alignments, and organization. We analyzed plasmids containing various antimicrobial resistance genes, together with genes coding for IMP-type metallo-ß-lactamases. Moreover, the mapping of plasmids with antimicrobial resistance genes and Incompatibility (Inc) groups showed that clustered plasmids with a similar organization of antimicrobial resistance genes were not always classified into the same Inc groups, indicating that the program displays multiple plasmids regardless of the Inc group classification. Our results showed that this calculation protocol and mapping strategy could provide a valuable tool for the practical and convenient visualization and comparison of the genomic structure of multiple plasmids in parallel.

Comparative Metabolomics Reveals a Bifunctional Antibacterial Conjugate from Combined-Culture of Streptomyces hygroscopicus HOK021 and Tsukamurella pulmonis TP-B0596.

To investigate the potential for secondary metabolite biosynthesis by Streptomyces species, we employed a coculture method to discover natural bioactive products and identified specific antibacterial activity from a combined-culture of Streptomyces hygroscopicus HOK021 and Tsukamurella pulmonis TP-B0596. Molecular networking using ultrahigh performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (UPLC-QTOF-MS/MS) data revealed a specific clade of metabolites in this combined-culture that were not detected in both monocultures. Using the chemical profiles, a previously unidentified conjugate between FabF inhibitor and catechol-type siderophore was successfully identified and named harundomycin A. Harundomycin A was a conjugate between the 2,4-dihydroxy-3-aminobenzoate moiety of platensimycin and N,N'-bis(2,3-dihydroxybenzoyl)-O-seryl-cysteine (bisDHBA-Ser-Cys) with a thioester linkage. Along with the production of harundomycin A, platensimycin, its thiocarboxylic acid form thioplatensimycin, enterobactin, and its degradation product N,N'-bis(2,3-dihydroxybenzoyl)-O-l-seryl-dehydroalanine (bisDHBA-Ser-Dha) were also induced in the combined-culture. Genomic data of S. hygroscopicus HOK021 and T. pulmonis TP-B0596 indicated that strain HOK021 possessed biosynthetic gene clusters for both platensimycin and enterobactin, and thereby revealed that T. pulmonis stimulates HOK021 and acts as an inducer of both of these metabolites. Although the harundomycin A was modified by bulky bisDHBA-Ser-Cys, responsible for the binding to the target molecule FabF, it showed a similar antibacterial spectrum to platensimycin, including against methicillin-resistant Staphylococcus aureus and vancomycin-resistant enterococci, suggesting that the pharmacophore is platensimycin. Additionally, Chrome Azurol S assay showed that harundomycin A possesses ferric iron-chelating activity comparable to that of enterobactin. Our study demonstrated the transformation of existing natural products to bifunctional molecules driven by bacterial interaction.

PCR-based ORF typing of Klebsiella pneumoniae for rapid identification of global clones and transmission events.

AIMS: Klebsiella pneumoniae is a major cause of healthcare-associated infections. In this study, we aimed to develop a rapid and simple genotyping method that can characterize strains causing nosocomial infections. METHODS AND RESULTS: The PCR-based open reading frame (ORF) typing (POT) method consists of two multiplex PCR reactions that were designed to detect 25 ORFs specific to bacterial genetic lineages, species, antimicrobial-resistant genes (blaCTX-M group-1 , blaCTX-M group-9 , blaIMP and blaKPC ), a capsular K1-specific gene and a virulence factor gene (rmpA/A2). The electrophoresis results are then digitized. A total of 192 strains (136 clinical and 8 reference strains of K. pneumoniae, 33 clinical and 1 reference strains of K. variicola and 14 clinical strains of K. quasipneumoniae) were classified into 95, 26 and 11 POT values, respectively. The distribution patterns of ORFs among K. pneumoniae correlated well with multilocus sequence typing (MLST). Furthermore, closely related species could be distinguished and key antimicrobial resistance and hypervirulence genes were identified as part of POT. CONCLUSIONS: The POT method was developed and validated for K. pneumoniae. In comparison to MLST, the POT method is a rapid and easy genotyping method for monitoring transmission events by K. pneumoniae in clinical microbiology laboratories. SIGNIFICANCE AND IMPACT OF THE STUDY: The POT method supplies clear and informative molecular typing results for K. pneumoniae. The method would facilitate molecular epidemiological analysis in infection control and hospital epidemiology investigations.

Genomic Traits Associated with Virulence and Antimicrobial Resistance of Invasive Group B Streptococcus Isolates with Reduced Penicillin Susceptibility from Elderly Adults.

This study aimed to investigate genomic traits underlying the antimicrobial resistance and virulence of multidrug-resistant (MDR) group B streptococci with reduced penicillin susceptibility (PRGBS) recovered from elderly patients with bloodstream infections, which remain poorly characterized. The pangenome was found to be open, with the predicted pan- and core genome sizes being 3,531 and 1,694 genes, respectively. Accessory and unique genes were enriched for the Clusters of Orthologous Groups (COG) categories L, Replication, recombination, and repair, and K, Transcription. All MDR PRGBS isolates retained a core virulence gene repertoire (bibA, fbsA/-B/-C, cspA, cfb, hylB, scpB, lmb, and the cyl operon), supporting an invasive ability similar to that of the other invasive GBS, penicillin-susceptible GBS (PSGBS), and noninvasive PRGBS isolates. The putative sequence type 1 (ST1)-specific AlpST-1 virulence gene was also retained among the serotype Ia/ST1 PRGBS isolates. In addition to tet(M) and erm(B), mef(A)-msr(D) elements or the high-level gentamicin resistance gene aac(6')-aph(2″), which are both rare in PSGBS, were detected among those MDR PRGBS isolates. In the core single-nucleotide polymorphism (SNP) phylogenetic tree, all invasive ST1 PRGBS isolates with serotypes Ia and III were placed together in a clade with a recombination rate of 3.97, which was 36 times higher than the value found for a clade formed by serotype V/ST1 PSGBS isolates derived mostly from human blood. ST1 has been the predominant sequence type among the PRGBS isolates in Japan, and serotypes Ia and III have been very rare among the ST1 PSGBS isolates. Thus, these lineages that mostly consisted of serotypes Ia/ST1 and III/ST1 PRGBS could possibly emerge through recombination within the ST1 populations. IMPORTANCE Streptococcus agalactiae, or group B Streptococcus (GBS), is recognized as the leading cause of neonatal invasive infections. However, an increasing incidence of invasive GBS infections among nonpregnant adults, particularly the elderly and those with underlying diseases, has been observed. There is a trend toward the increasing occurrence of penicillin nonsusceptibility among GBS clinical isolates, from 4.8% in 2008 to 5.8% in 2020 in Japan. Also, in the United States, the frequency of adult invasive GBS isolates suggestive of ß-lactam nonsusceptibility increased from 0.7% in 2015 to 1.0% in 2016. In adults, mortality has been significantly higher among patients with bacteremia than among those without bacteremia. Our study revealed that invasive GBS with reduced penicillin susceptibility (PRGBS) isolates harbor major virulence and resistance genes known among GBS, highlighting the need for large population-based genomic surveillance studies to better understand the clinical relevance of invasive PRGBS isolates.

Dissecting the clonality of I1 plasmids using ORF-based binarized structure network analysis of plasmids (OSNAp).

OBJECTIVES: We aimed to elucidate the relationship among blaCTX-M-carrying plasmids and their transmission between humans and domestic animals. METHODS: Phylogenetic relationship of 90 I1 plasmids harboring blaCTX-M genes encoding extended-spectrum ß-lactamase (ESBL) was analyzed using the ORF-based binarized structure network analysis of plasmids (OSNAp). RESULTS: The majority of plasmids carrying blaCTX-M-1 or blaCTX-M-8 belonged to a single lineage, respectively, and were primarily associated with domestic animals especially chickens. On the other hand, plasmids carrying blaCTX-M-14 or blaCTX-M-15, identified from both humans and domestic animals, were distributed in two or more lineages. CONCLUSION: OSNAp has revealed the phylogenetic relationships and diversity of plasmids carrying blaCTX-M more distinctly than pMLST. The findings suggest that circulation of I1 plasmids between humans and animals may contribute to their diversity.

Molecular Epidemiology of Enterobacter cloacae Complex Isolates with Reduced Carbapenem Susceptibility Recovered by Blood Culture.

The Enterobacter cloacae complex (ECC) is one of the most common causes of bacteremia and leads to poor clinical outcomes. The aim of this study was to clarify the antimicrobial susceptibility profiles and genetic backgrounds of non-carbapenemase-producing reduced-carbapenem-susceptible (RCS) ECC blood isolates in Japan using agar dilution antimicrobial susceptibility testing, whole-genome sequencing, and quantitative polymerase chain reaction for ampC, ompC, and ompF transcripts. Forty-two ECC blood isolates were categorized into RCS and carbapenem-susceptible groups based on the minimum inhibitory concentration of imipenem. The RCS ECC blood isolates belonged to distinct species and sequence types and produced varying class C ß-lactamases. The E. roggenkampii, E. asburiae, and E. bugandensis isolates belonged only to the RCS group. Some E. hormaechei ssp. steigerwaltii isolates from the RCS group exhibited AmpC overexpression caused by amino acid substitutions in AmpD and AmpR along with ompF downregulation. These findings suggest that non-carbapenemase-producing RCS ECC blood isolates are genetically diverse.

Isolation of Group A Streptococci with Reduced In Vitro ß-Lactam Susceptibility Harboring Amino Acid Substitutions in Penicillin-Binding Proteins in Japan.

Streptococcus pyogenes (group A Streptococcus [GAS]) has long been regarded as being susceptible to ß-lactams. However, amino acid substitutions in penicillin-binding protein 2X (PBP2X) conferring reduced in vitro ß-lactam susceptibility have been indicated since 2019 in the United States and Iceland. Here, we report the first isolation of Streptococcus pyogenes possessing the PBP2X substitution conferring reduced in vitro ß-lactam susceptibility in Asia; however, the MICs were below the susceptible breakpoint of the CLSI.

Crystal Structures of Metallo-ß-Lactamase (IMP-1) and Its D120E Mutant in Complexes with Citrate and the Inhibitory Effect of the Benzyl Group in Citrate Monobenzyl Ester.

The emergence and rapid spread of carbapenem-resistant pathogens producing metallo-ß-lactamases such as IMP-1 and NDM-1 have been of great concern in the global clinical setting. The X-ray crystal structures of IMP-1 from Serratia marcescens and its single mutant, D120E, in complexes with citrate were determined at resolutions of 2.00 and 1.85 Å, respectively. Two crystal structures indicate that a single mutation at position 120 caused a structural change around Zn1, where the geometry changes from a tetrahedron in the native IMP-1 to a square pyramid in D120E. Based on these two complex structures, the authors synthesized citrate monobenzyl ester 1 to evaluate the structural requirement for the inhibitory activity against IMP-1 and compared the inhibitory activities with nonsubstituted citrate. The introduction of a benzyl group into citrate enhanced the inhibitory activity in comparison to citrate (IC50 > 5 mM).

Practical Agar-Based Disk Diffusion Tests Using Sulfamoyl Heteroarylcarboxylic Acids for Identification of Subclass B1 Metallo-ß-Lactamase-Producing Enterobacterales.

The worldwide distribution of carbapenemase-producing Enterobacterales (CPE) is a serious public health concern as they exhibit carbapenem resistance, thus limiting the choice of antimicrobials for treating CPE infections. Combination treatment with a ß-lactam and one of the newly approved ß-lactamase inhibitors, such as avibactam, relebactam, or vaborbactam, provides a valuable tool to cope with CPE; however, these inhibitors are active only against serine-type carbapenemases and not against metallo-ß-lactamases (MßLs). Therefore, it is important to readily differentiate carbapenemases produced by CPE by using simple and reliable methods in order to choose an appropriate treatment. Here, we developed three practical agar-based disk diffusion tests (double-disk synergy test [DDST], disk potentiation test, and modified carbapenem inactivation method [mCIM]) to discriminate the production of subclass B1 MßLs, such as IMP-, NDM-, and VIM-type MßLs, from the other carbapenemases, especially serine-type carbapenemases. This was accomplished using B1 MßL-specific sulfamoyl heteroarylcarboxylic acid inhibitors, 2,5-dimethyl-4-sulfamoylfuran-3-carboxylic acid (SFC) and 2,5-diethyl-1-methyl-4-sulfamoylpyrrole-3-carboxylic acid (SPC), originally developed by us. The DDST and mCIM using SFC and SPC revealed high sensitivity (95.3%) and specificity (100%) in detecting B1 MßL-producing Enterobacterales. In the disk potentiation test, the sensitivities using SFC and SPC were 89.1% and 93.8%, respectively, whereas the specificities for both were 100%. These methods are simple and inexpensive and have a high accuracy rate. These methods would therefore be of immense assistance in the specific detection and discrimination of B1 MßL-producing Enterobacterales in clinical microbiology laboratories and would lead to better prevention against infection with such multidrug-resistant bacteria in clinical settings.

Presence of Colistin- and Tigecycline-Resistant Klebsiella pneumoniae ST29 in Municipal Wastewater Influents in Japan.

The aim of this study was to investigate the presence of colistin- and/or tigecycline-resistant Klebsiella spp. in influents from four wastewater treatment plants (WWTPs), which partly reflect the gut microbiome of human populations. Colistin- and tigecycline-resistant Klebsiella pneumoniae isolates (K30/ST29) were detected four times from the WWTP A during a period of 3 months. Disruptions of the mgrB and ramR genes by ISEc68 and ISKpn21, respectively, were identified in those four isolates. They also shared the IncL/M 86,197-bp plasmids carrying a blaCTX-M-3 and Tn1548-associated armA [IS26-IntI1-dfrA12-gucF-aadA2-qacEΔ1-sul1-ISCR1-ISEc28-armA-ISEc29-msr(E)-mph(E)-IS26]. Those isolates formed a distinct cluster within wgMLST clusters of ST29 K30 public reference strains of human origin and were unique due to harboring of Tn21-like mercury resistance operon transposons in addition to silver, copper, and arsenic resistance determinants. Five K. pneumoniae strains with different STs and 1 Klebsiella quasipneumoniae strain, exhibiting colistin resistance, were detected in WWTPs B, C, and D. For these isolates, disruptions of mgrB by ISEc68 (three isolates) or ISEcl1 (one isolate), insertion of IS2 in the mgrB promoter region (one isolate), and inactivation of MgrB by a nonsense mutation (one isolate) were identified. Close monitoring of these mcr-negative colistin- and/or tigecycline-resistant bacteria in wastewater influents is imperative to avoid further limiting of treatment options.

Genomic characterisation and epidemiology of nosocomial Serratia marcescens isolates resistant to ceftazidime and their plasmids mediating rare blaTEM-61.

OBJECTIVES: We determined the whole DNA sequences of plasmids carrying a rare extended-spectrum ß-lactamase gene (blaTEM-61) to precisely understand the spread of resistance among nosocomial Serratia marcescens populations. METHODS: Twenty non-duplicate ceftazidime-resistant S. marcescens nosocomial isolates (ceftazidime MICs, 32 to >128 mg/L) collected over 1 year were pulsotyped and nucleotide sequences of the blaTEM-61 gene and its promoter region were determined. Twelve representative isolates were analysed by whole-genome sequencing. RESULTS: The 20 isolates comprised two distinct pulsotypes: I (14 isolates) and II (6 isolates). They all contained the blaTEM-61 gene. A polymorphism in the repeat number of a 15-nucleotide sequence (5'-ATGTCATGATAATAA-3') was found in the promoter region of blaTEM-61; two, three and four repeat units were found in 6, 12 and 2 isolates, respectively. Single nucleotide polymorphism (SNP)-based phylogenetic analysis of 12 isolates revealed that 7 isolates of pulsotype I (12-44 SNP differences) and 5 isolates of pulsotype II (15-55 SNP differences) formed two distinct clusters of genotypes 1 and 2, respectively. All 12 isolates harboured a plasmid carrying the Tn1-blaTEM-61 element, although they were slightly different in size (78 883 bp, 78 898 bp and 78 913 bp) owing to differences in the number of 15-bp repetitive sequences. A 42 542-bp broad-host-range plasmid carrying the Tn1-blaTEM-61 element was also found in one of the isolates. CONCLUSIONS: We characterised a plasmid-encoded novel Tn1-blaTEM-61 element and transposon-dependent mechanisms underlying the propagation of antibiotic resistance, together with repeated new polymorphic 15-bp units in the promoter of blaTEM-61.

Ruptured Emphysematous Prostatic Abscess Caused by K1-ST23 Hypervirulent Klebsiella pneumoniae Presenting as Brain Abscesses: A Case Report and Literature Review.

Emphysematous prostatic abscess (EPA) is an extremely rare but potentially fatal urinary tract infection (UTI). Here, we describe a case (a 69-year-old male with prediabetes) of ruptured EPA caused by a hypervirulent Klebsiella pneumoniae (hvKp) K1-ST23 strain, presenting as motor aphasia. Our patient presented with ruptured EPA concurrent with various severe systemic pyogenic complications (e.g., urethro-prostatic fistula, ascending UTIs, epididymal and scrotal abscesses, and liver, lung, and brain abscesses). Whole-body computed tomography (CT) and next-generation sequencing (NGS) were useful for the detection of ruptured EPA and its systemic complications, and for identification of K1-ST23 hvKp strains, respectively. Subsequently, the infections were successfully treated with aggressive antimicrobial therapy and multiple surgical procedures. This case highlights the significance of awareness of this rare entity, the clinical importance of CT for the early diagnosis of EPA and the detection of its systemic complications in view of hvKp being an important causative organism of severe community-acquired UTI, and the usefulness of NGS to identify hvKp strains.

Daptomycin Susceptibility of Group B Streptococcus.

We previously reported the emergence and high prevalence of group B streptococci (GBS) with reduced penicillin susceptibility (PRGBS) clinical isolates in Japan. PRGBS tend to be non-susceptible to macrolides and fluoroquinolones. In our previous study, we found that the minimum inhibitory concentration (MIC) of daptomycin for one clinical isolate of GBS was above the susceptible breakpoint settled by the Clinical and Laboratory Standards Institute (CLSI). This suggests the possibility of the unrecognized spread of daptomycin-non-susceptible clinical GBS isolates in Japan. This study aimed to analyze the daptomycin susceptibility in 1,046 clinical GBS isolates that were recovered after the approval of daptomycin in Japan. MICs of daptomycin for the 1,046 clinical isolates were determined by the microdilution method recommended by the CLSI. The MIC range was 0.12-1 µg/mL, and the MIC50 and MIC90 were 0.5 µg/mL and 1 µg/mL, respectively. All the GBS isolates evaluated in this study were susceptible to daptomycin. Therefore, at present, daptomycin might be considered as a new option to treat GBS infections, especially multidrug-resistant PRGBS infections.

Aminoglycoside Resistance: Updates with a Focus on Acquired 16S Ribosomal RNA Methyltransferases.

The clinical usefulness of aminoglycosides has been revisited as an effective choice against ß-lactam-resistant and fluoroquinolone-resistant gram-negative bacterial infections. Plazomicin, a next-generation aminoglycoside, was introduced for the treatment of complicated urinary tract infections and acute pyelonephritis. In contrast, bacteria have resisted aminoglycosides, including plazomicin, by producing 16S ribosomal RNA (rRNA) methyltransferases (MTases) that confer high-level and broad-range aminoglycoside resistance. Aminoglycoside-resistant 16S rRNA MTase-producing gram-negative pathogens are widespread in various settings and are becoming a grave concern. This article provides up-to-date information with a focus on aminoglycoside-resistant 16S rRNA MTases.