BACKGROUND: Data from surveillance on antibiotic resistance have shown an increasing prevalence of non-enzymatic resistance (β-lactamase-negative ampicillin-resistant) to β-lactam antibiotics among H. influenzae strains in the Czech Republic. Aminopenicillins are recommended agents for non-invasive Haemophilus influenzae infections. The phenomenon of non-enzymatic resistance to β-lactams is complicated by the fact that the phenotypic detection of PBP3 with specific amino acid substitutions (rPBP3) is challenging, since rPBP3 isolates have repeatedly been demonstrated to be split by the epidemiological cut-off values (ECOFF) for aminopenicillins defined by EUCAST. OBJECTIVES: We sought to determine whether the penicillin disc has sufficient detection ability to predict the non-enzymatic mechanism; whether other antibiotics can be used for detection; and what is the agreement between the broth microdilution and disc diffusion methods. METHODS: We undertook susceptibility testing of selected antibiotics according to EUCAST of 153 rPBP3 strains, and sequencing of the ftsI gene to determination amino acid substitutions. RESULTS: For a selected set of rPBP strains: (i) the detection capability for penicillin, ampicillin, cefuroxime and amoxicillin/clavulanate was found to be 91.5%, 94.4%, 89.5% and 70.6%, respectively; (ii) the categorical agreement between the disc diffusion method and the MIC for ampicillin and cefuroxime was 71.1% and 83.8%, respectively. CONCLUSIONS: We observed better recognition of rPBP3 strains by the ampicillin disc than by the penicillin disc. There is frequently a discrepancy in the interpretation of susceptibility results between the methods used.

• MeSH
• antibakteriální látky * farmakologie   MeSH
• beta-laktamová rezistence * genetika   MeSH
• beta-laktamy * farmakologie   MeSH
• Haemophilus influenzae * genetika   účinky léků   MeSH
• hemofilové infekce mikrobiologie   MeSH
• lidé MeSH
• mikrobiální testy citlivosti * MeSH
• proteiny vázající penicilin * genetika   MeSH
• sekvenční analýza DNA MeSH
• substituce aminokyselin * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• antibakteriální látky * MeSH
• beta-laktamy * MeSH
• proteiny vázající penicilin * MeSH

This report describes mutations in genes responsible for cell deformities in haemophili under beta-lactam pressure in vitro. Light and transmission electron microscopy confirmed a hypothesis regarding changes in the shape of haemophili that had become more filamentous in the presence of ampicillin (2 mg/L) and cefuroxime (8 mg/L) after 30 days of serial passage. Short-axis size increased by 28% (from 0.767 to 1.06 µm) and long-axis length increased by 54% (from 1 to 2.175 µm). Additionally, whole-genome sequencing analysis (Illumina platform, software PROKKA) revealed a variety of mutations in genes responsible for cell morphology in isolates examined in this study: ftsI (A1576 → C; G1154 → C; T986 → C; G1684 → C), mreB (C476 → T), mreC (A5 → G), mrdA (A1148 → G; C179 → T; G1613 → T), mrdB (T668 → G), mltC (C1016 → T) and rodA (T668 → G). The results of this study indicate that shifts in bacterial shape could play a role in the adaptation of haemophili to a new niche created by beta-lactams as a strategy of antibiotic therapy survival.

• Klíčová slova
• Beta-lactams, Haemophili, Whole-genome sequencing,
• MeSH
• ampicilin farmakologie   MeSH
• antibakteriální látky * farmakologie   MeSH
• beta-laktamy * farmakologie   MeSH
• cefuroxim farmakologie   MeSH
• genom bakteriální * MeSH
• Haemophilus influenzae * genetika   účinky léků   MeSH
• lidé MeSH
• mikrobiální testy citlivosti MeSH
• mutace MeSH
• sekvenování celého genomu * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• ampicilin MeSH
• antibakteriální látky * MeSH
• beta-laktamy * MeSH
• cefuroxim MeSH

BACKGROUND: Aminopenicillins are recommended agents for non-invasive Haemophilus influenzae infections. One of the mechanisms of resistance to β-lactams is the alteration of the transpeptidase region of penicillin binding protein 3 (PBP3) which is caused by mutations in the ftsI gene. It was shown that exposure to beta-lactams has a stimulating effect on increase of prevalence of H. influenzae strains with the non-enzymatic mechanism of resistance. OBJECTIVES: The aim of our study was to compare the mutational potential of ampicillin and cefuroxime in H. influenzae strains, determination of minimum inhibitory concentration and the evolution of mutations over time, focusing on amino acid substitutions in PBP3. METHODS: 30 days of serial passaging of strains in liquid broth containing increasing concentrations of ampicillin or cefuroxime was followed by whole-genome sequencing. RESULTS: On average, cefuroxime increased the minimum inhibitory concentration more than ampicillin. The minimum inhibitory concentration was increased by a maximum of 32 fold. Substitutions in the PBP3 started to appear after 15 days of passaging. In PBP3, cefuroxime caused different substitutions than ampicillin. CONCLUSIONS: Our experiment observed differences in mutation selection by ampicillin and cefuroxime. Selection pressure of antibiotics in vitro generated substitutions that do not occur in clinical strains in the Czech Republic.

• Klíčová slova
• Ampicillin, Cefuroxime, Haemophilus, PBP3, WGS,
• MeSH
• ampicilin * farmakologie   MeSH
• antibakteriální látky * farmakologie   MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• cefuroxim * farmakologie   MeSH
• Haemophilus influenzae * genetika   účinky léků   MeSH
• hemofilové infekce mikrobiologie   MeSH
• lidé MeSH
• mikrobiální testy citlivosti * MeSH
• molekulární evoluce MeSH
• mutace * MeSH
• proteiny vázající penicilin * genetika   metabolismus   MeSH
• sekvenování celého genomu MeSH
• selekce (genetika) MeSH
• sériové pasážování MeSH
• substituce aminokyselin * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• ampicilin * MeSH
• antibakteriální látky * MeSH
• bakteriální proteiny MeSH
• cefuroxim * MeSH
• proteiny vázající penicilin * MeSH

Enteroaggregative Escherichia coli (EAEC) strains including those of serogroup O111 are important causes of diarrhea in children. In the Czech Republic, no information is available on the etiological role of EAEC in pediatric diarrhea due to the lack of their targeted surveillance. To fill this gap, we determined the proportion of EAEC among E. coli O111 isolates from children with gastrointestinal disorders ≤ 2 years of age submitted to the National Reference Laboratory for E. coli and Shigella during 2013-2022. EAEC accounted for 177 of 384 (46.1 %) E. coli O111 isolates, being the second most frequent E. coli O111 pathotype. Most of them (75.7 %) were typical EAEC that carried aggR, usually with aaiC and aatA marker genes; the remaining 24.3 % were atypical EAEC that lacked aggR but carried aaiC and/or aatA. Whole genome sequencing of 11 typical and two atypical EAEC O111 strains demonstrated differences in serotypes, sequence types (ST), virulence gene profiles, and the core genomes between these two groups. Typical EAEC O111:H21/ST40 strains resembled by their virulence profiles including the presence of the aggregative adherence fimbriae V (AAF/V)-encoding cluster to such strains from other countries and clustered with them in the core genome multilocus sequence typing (cgMLST). Atypical EAEC O111:H12/ST10 strains lacked virulence genes of typical EAEC and differed from them in cgMLST. All tested EAEC O111 strains displayed stacked-brick aggregative adherence to human intestinal epithelial cells. The AAF/V-encoding cluster was located on a plasmid of 95,749 bp or 93,286 bp (pAAO111) which also carried aggR, aap, aar, sepA, and aat cluster. EAEC O111 strains were resistant to antibiotics, in particular to aminopenicillins and cephalosporins; 88.3 % produced AmpC β-lactamase, and 4.1 % extended spectrum β-lactamase. We conclude that EAEC are frequent among E. coli O111 strains isolated from children with gastrointestinal disorders in the Czech Republic. To reliably assess the etiological role of EAEC in pediatric diarrhea, a serotype-independent, PCR-based pathotype surveillance system needs to be implemented in the future.

• Klíčová slova
• Czech Republic, Enteroaggregative Escherichia coli O111, PAA(O111) plasmid, Pediatric diarrhea, Surveillance, Whole genome sequencing,
• MeSH
• antibakteriální látky farmakologie   MeSH
• Escherichia coli * genetika   izolace a purifikace   patogenita   klasifikace   MeSH
• faktory virulence genetika   MeSH
• gastrointestinální nemoci mikrobiologie   MeSH
• genom bakteriální MeSH
• infekce vyvolané Escherichia coli * mikrobiologie   epidemiologie   MeSH
• kojenec MeSH
• lidé MeSH
• multilokusová sekvenční typizace MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• proteiny z Escherichia coli genetika   MeSH
• průjem * mikrobiologie   MeSH
• sekvenování celého genomu * MeSH
• séroskupina MeSH
• trans-aktivátory MeSH
• Check Tag
• kojenec MeSH
• lidé MeSH
• mužské pohlaví MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Česká republika epidemiologie   MeSH
• Názvy látek
• AggR protein, E coli MeSH Prohlížeč
• antibakteriální látky MeSH
• faktory virulence MeSH
• proteiny z Escherichia coli MeSH
• trans-aktivátory MeSH

The objective of this study was to assess the susceptibility of cefiderocol against multidrug-resistant carbapenemase-producing and nonproducing bacteria. The panel comprised 182 isolates of the order Enterobacterales, and 40 strains of Pseudomonas aeruginosa. Antimicrobial susceptibility testing has been performed using broth microdilution method according to the European Committee on Antimicrobial Susceptibility Testing recommendations. Mass spectrometry matrix-assisted laser desorption/ionization-time of flight mass spectrometry and carbapenemase-producing test were used to verify the presence of carbapenemases in clinical isolates. The genetic expression of single carbapenemases (blaKPC, blaOXA-48, blaNDM, blaVIM, blaIMP, blaGES) was determined by real-time polymerase chain reaction. Cefiderocol exhibited a good activity against the majority of strains tested in this study. Altogether, growth of 81.9% (n = 149) strains of the order Enterobacterales and 77.5% (n = 31) of P. aeruginosa isolates were inhibited at minimal inhibitory concentration (MIC) ≤2 mg/L. Values MIC50/MIC90 were 0.5/8 mg/L for enterobacteria, and 1/8 mg/L for P. aeruginosa. One isolate (Klebsiella pneumoniae) harboring two carbapenemases (blaOXA-48, blaNDM) had cefiderocol MIC 0.5 mg/L. In enterobacteria resistant to cefiderocol, blaNDM carbapenemase prevailed (43.3%, n = 29), followed by blaOXA-48 (31.3%, n = 21) and blaKPC (4.5%, n = 3). blaIMP (n = 8) and blaVIM (n = 1) metallo-β-lactamases dominated in cefiderocol-resistant P. aeruginosa (n = 9) isolates. Very good susceptibility (100%) to this drug showed blaGES-positive strains of P. aeruginosa (n = 8) and isolates resistant to meropenem without confirmed carbapenemase gene (n = 10). In this study, cefiderocol demonstrated potent activity against important nosocomial pathogens, therefore, therapeutic options of this drug against multidrug-resistant bacteria should be considered.

• Klíčová slova
• Enterobacterales, Pseudomonas aeruginosa, carbapenemase, cefiderocol,
• MeSH
• antibakteriální látky * farmakologie   MeSH
• beta-laktamasy genetika   metabolismus   MeSH
• cefiderokol MeSH
• Enterobacteriaceae genetika   metabolismus   MeSH
• karbapenemy * farmakologie   MeSH
• mikrobiální testy citlivosti MeSH
• Pseudomonas aeruginosa MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antibakteriální látky * MeSH
• beta-laktamasy MeSH
• karbapenemy * MeSH

The occurrence of colistin resistance has increased rapidly among Enterobacterales around the world. We performed a national survey of plasmid-mediated colistin resistance in human clinical isolates through a retrospective analysis of samples from 2009 to 2017 and a prospective sampling in 2018-2020. The aim of this study was to identify and characterize isolates with mcr genes from various regions of the Czech Republic using whole genome sequencing (WGS). Of all 1932 colistin-resistant isolates analyzed, 73 (3.8%) were positive for mcr genes. Most isolates carried mcr-1 (48/73) and were identified as Escherichia coli (n = 44) and Klebsiella pneumoniae (n = 4) of various sequence types (ST). Twenty-five isolates, including Enterobacter spp. (n = 24) and Citrobacter freundii (n = 1) carrying the mcr-9 gene were detected; three of them (Enterobacter kobei ST54) co-harbored the mcr-4 and mcr-9 genes. Multi-drug resistance phenotype was a common feature of mcr isolates and 14% (10/73) isolates also co-harbored clinically important beta-lactamases, including two isolates with carbapenemases KPC-2 and OXA-48. Phylogenetic analysis of E. coli ST744, the dominant genotype in this study, with the global collection showed Czech isolates belonged to two major clades, one containing isolates from Europe, while the second composed of isolates from diverse geographical areas. The mcr-1 gene was carried by IncX4 (34/73, 47%), IncHI2/ST4 (6/73, 8%) and IncI2 (8/73, 11%) plasmid groups. Small plasmids belonging to the ColE10 group were associated with mcr-4 in three isolates, while mcr-9 was carried by IncHI2/ST1 plasmids (4/73, 5%) or the chromosome (18/73, 25%). We showed an overall low level of occurrence of mcr genes in colistin-resistant bacteria from human clinical samples in the Czech Republic.

• Klíčová slova
• Enterobacterales, antibiotic resistance, human, mcr, plasmids,
• Publikační typ
• časopisecké články MeSH

The ribotyping of Clostridioides difficile is one of the basic methods of molecular epidemiology for monitoring the spread of C. difficile infections. In the Czech Republic, this procedure is mainly available in university hospitals. The introduction of ribotyping in a tertiary health care facility such as Liberec Regional Hospital not only increases safety in the facility but also supports regional professional development. In our study, 556 stool samples collected between June 2017 and June 2018 were used for C. difficile infection screening, followed by cultivation, toxinotyping, and ribotyping of positive samples. The toxinotyping of 96 samples revealed that 44.8% of typed strains could produce toxins A and B encoded by tcdA and tcdB, respectively. The ribotyping of the same samples revealed two epidemic peaks, caused by the regionally most prevalent ribotype 176 (n = 30, 31.3). C. difficile infection incidence ranged between 5.5 and 4.2 cases per 10,000 patient-bed days. Molecular diagnostics and molecular epidemiology are the two most developing parts of clinical laboratories. The correct applications of molecular methods help ensure greater safety in hospitals.

• Klíčová slova
• Clonal spreading, Clostridioides difficile, Health care facility, Ribotyping, Toxinotyping,
• MeSH
• bakteriální toxiny * MeSH
• Clostridioides difficile * genetika   MeSH
• Clostridioides MeSH
• klostridiové infekce * diagnóza   epidemiologie   MeSH
• lidé MeSH
• nemocnice univerzitní MeSH
• poskytování zdravotní péče MeSH
• ribotypizace MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální toxiny * MeSH

Multinational surveillance programmes for methicillin-resistant Staphylococcus aureus (MRSA) are dependent on national structures for data collection. This study aimed to capture the diversity of national MRSA surveillance programmes and to propose a framework for harmonisation of MRSA surveillance. The International Society of Antimicrobial Chemotherapy (ISAC) MRSA Working Group conducted a structured survey on MRSA surveillance programmes and organised a webinar to discuss the programmes' strengths and challenges as well as guidelines for harmonisation. Completed surveys represented 24 MRSA surveillance programmes in 16 countries. Several countries reported separate epidemiological and microbiological surveillance. Informing clinicians and national policy-makers were the most common purposes of surveillance. Surveillance of bloodstream infections (BSIs) was present in all programmes. Other invasive infections were often included. Three countries reported active surveillance of MRSA carriage. Methodology and reporting of antimicrobial susceptibility, virulence factors, molecular genotyping and epidemiological metadata varied greatly. Current MRSA surveillance programmes rely upon heterogeneous data collection systems, which hampers international epidemiological monitoring and research. To harmonise MRSA surveillance, we suggest improving the integration of microbiological and epidemiological data, implementation of central biobanks for MRSA isolate collection, and inclusion of a representative sample of skin and soft-tissue infection cases in addition to all BSI cases.

• Klíčová slova
• Antimicrobial resistance, Epidemiology, Monitoring, Staphylococcus aureus,
• MeSH
• antibakteriální látky farmakologie   terapeutické užití   MeSH
• epidemiologické monitorování MeSH
• infekce měkkých tkání * farmakoterapie   MeSH
• lidé MeSH
• methicilin rezistentní Staphylococcus aureus * MeSH
• stafylokokové infekce * farmakoterapie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antibakteriální látky MeSH

The isolation of Planococcus glaciei (designed strain CNCTC 7660) from blood of a patient with appendicitis is reported. Species P. glaciei (type strain CGMCC 1.6846 T) was for the first time identified as an environmental bacterium acquired from a glacier in China in 2009. To reveal the identity of the isolate CNCTC 7660, the 16S rDNA sequencing and the whole genome sequencing (Illumina MiSeq, Oxford Nanopore) were performed. The level of 16S rDNA gene sequencing similarity between CNCTC 7660 and CGMCC 1.6846 T was 99.55%. Phylogenetic analysis and average nucleotide analysis (ANI) based on the whole genome sequencing confirmed that the isolate CNCTC 7660 and CGMCC1.6846 T had ANI value above the taxonomic threshold for belonging to the same species (95%). The G + C content of CNCTC 7660 DNA was 46.8% (mol/mol). Except for the growth temperature, strains CGMCC1.6846 T and CNCTC 7660 were distinguished also biochemically. Due to the lack of information about the pathogenicity of P. glaciei, the possibility that it exerts pathogenicity in persons is suggested. But for understanding the nature of this species, further cases are needed.

• MeSH
• DNA bakterií genetika   MeSH
• fylogeneze MeSH
• hybridizace nukleových kyselin MeSH
• lidé MeSH
• mastné kyseliny * analýza   MeSH
• RNA ribozomální 16S genetika   MeSH
• sekvenční analýza DNA MeSH
• techniky typizace bakterií MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• DNA bakterií MeSH
• mastné kyseliny * MeSH
• RNA ribozomální 16S MeSH

• Publikační typ
• tisková chyba MeSH