Evolution of mutations in the ftsI gene leading to amino acid substitutions in PBP3 in Haemophilus influenzae strains under the selective pressure of ampicillin and cefuroxime.

BACKGROUND: Aminopenicillins are recommended agents for non-invasive Haemophilus influenzae infections. One of the mechanisms of resistance to ß-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.

Enteroaggregative Escherichia coli: Frequent, yet underdiagnosed pathotype among E. coli O111 strains isolated from children with gastrointestinal disorders in the Czech Republic.

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 ß-lactamase, and 4.1 % extended spectrum ß-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.

Staphylococcus aureus bloodstream infections: diverging trends of meticillin-resistant and meticillin-susceptible isolates, EU/EEA, 2005 to 2018.

BackgroundInvasive infections caused by Staphylococcus aureus have high clinical and epidemiological relevance. It is therefore important to monitor the S. aureus trends using suitable methods.AimThe study aimed to describe the trends of bloodstream infections (BSI) caused by meticillin-resistant S. aureus (MRSA) and meticillin-susceptible S. aureus (MSSA) in the European Union (EU) and the European Economic Area (EEA).MethodsAnnual data on S. aureus BSI from 2005 to 2018 were obtained from the European Antimicrobial Resistance Surveillance Network (EARS-Net). Trends of BSI were assessed at the EU/EEA level by adjusting for blood culture set rate (number of blood culture sets per 1,000 days of hospitalisation) and stratification by patient characteristics.ResultsConsidering a fixed cohort of laboratories consistently reporting data over the entire study period, MRSA percentages among S. aureus BSI decreased from 30.2% in 2005 to 16.3% in 2018. Concurrently, the total number of BSI caused by S. aureus increased by 57%, MSSA BSI increased by 84% and MRSA BSI decreased by 31%. All these trends were statistically significant (p < 0.001).ConclusionsThe results indicate an increasing health burden of MSSA BSI in the EU/EEA despite a significant decrease in the MRSA percentage. These findings highlight the importance of monitoring antimicrobial resistance trends by assessing not only resistance percentages but also the incidence of infections. Further research is needed on the factors associated with the observed trends and on their attributable risk.

Whole genome sequencing of macrolide resistant Streptococcus pneumoniae serotype 19A sequence type 416.

BACKGROUND: The resistance of Streptococcus pneumoniae to macrolides is becoming an increasingly important issue and thus it is important to understand the genetics related to adaptation of this species to the widespread use of antibiotics in Europe. The 58 isolates of S. pneumoniae belonging to sequence type (ST) 416 and serotype 19A and to several different phenotypes originated from Italy, Portugal and Czech Republic were thus sequenced on Illumina MiSeq. The aim of the study was to describe genetical origine of isolates, investigate their macrolide resistance and suggest reasons for spread of ST416 in the Czech Republic. RESULTS: Investigation of genes associated with serotype determined serotype switch between 15B and 19A serotypes and core genome multilocus sequence typing (cgMLST) confirmed the origine of concerned isolates in Netherlands15B-37 clone. Inspected genomes proved variability of genes associated with the macrolide resistance even within closely genetically relative isolates. CONCLUSIONS: Participation of 19A/ST416 on the spread of Netherlands15B-37 is accompanied by serotype switch between 19A and 15B serotypes and with acquisition of genes involved in macrolide resistance to the clone that was originally macrolide susceptible. There is evident tendency to interchanging and modifications of these and surrounding genes, that could lead to accelerate spreading of this sequence type in regions with high macrolide consumption.

Antibiotic administration reduces the rate of intraamniotic inflammation in preterm prelabor rupture of the membranes.

BACKGROUND: Preterm prelabor rupture of the membranes (PPROM) is frequently complicated by intraamniotic inflammatory processes such as intraamniotic infection and sterile intraamniotic inflammation. Antibiotic therapy is recommended to patients with PPROM to prolong the interval between this complication and delivery (latency period), reduce the risk of clinical chorioamnionitis, and improve neonatal outcome. However, there is a lack of information regarding whether the administration of antibiotics can reduce the intensity of the intraamniotic inflammatory response or eradicate microorganisms in patients with PPROM. OBJECTIVE: The first aim of the study was to determine whether antimicrobial agents can reduce the magnitude of the intraamniotic inflammatory response in patients with PPROM by assessing the concentrations of interleukin-6 in amniotic fluid before and after antibiotic treatment. The second aim was to determine whether treatment with intravenous clarithromycin changes the microbial load of Ureaplasma spp DNA in amniotic fluid. STUDY DESIGN: A retrospective cohort study included patients who had (1) a singleton gestation, (2) PPROM between 24+0 and 33+6 weeks, (3) a transabdominal amniocentesis at the time of admission, and (4) intravenous antibiotic treatment (clarithromycin for patients with intraamniotic inflammation and benzylpenicillin/clindamycin in the cases of allergy in patients without intraamniotic inflammation) for 7 days. Follow-up amniocenteses (7th day after admission) were performed in the subset of patients with a latency period lasting longer than 7 days. Concentrations of interleukin-6 were measured in the samples of amniotic fluid with a bedside test, and the presence of microbial invasion of the amniotic cavity was assessed with culture and molecular microbiological methods. Intraamniotic inflammation was defined as a bedside interleukin-6 concentration ≥745 pg/mL in the samples of amniotic fluid. Intraamniotic infection was defined as the presence of both microbial invasion of the amniotic cavity and intraamniotic inflammation; sterile intraamniotic inflammation was defined as the presence of intraamniotic inflammation without microbial invasion of the amniotic cavity. RESULTS: A total of 270 patients with PPROM were included in this study: 207 patients delivered within 7 days and 63 patients delivered after 7 days of admission. Of the 63 patients who delivered after 7 days following the initial amniocentesis, 40 underwent a follow-up amniocentesis. Patients with intraamniotic infection (n = 7) and sterile intraamniotic inflammation (n = 7) were treated with intravenous clarithromycin. Patients without either microbial invasion of the amniotic cavity or intraamniotic inflammation (n = 26) were treated with benzylpenicillin or clindamycin. Treatment with clarithromycin decreased the interleukin-6 concentration in amniotic fluid at the follow-up amniocentesis compared to the initial amniocentesis in patients with intraamniotic infection (follow-up: median, 295 pg/mL, interquartile range [IQR], 72-673 vs initial: median, 2973 pg/mL, IQR, 1750-6296; P = .02) and in those with sterile intraamniotic inflammation (follow-up: median, 221 pg/mL, IQR 118-366 pg/mL vs initial: median, 1446 pg/mL, IQR, 1300-2941; P = .02). Samples of amniotic fluid with Ureaplasma spp DNA had a lower microbial load at the time of follow-up amniocentesis compared to the initial amniocentesis (follow-up: median, 1.8 × 104 copies DNA/mL, 2.9 × 104 to 6.7 × 108 vs initial: median, 4.7 × 107 copies DNA/mL, interquartile range, 2.9 × 103 to 3.6 × 107; P = .03). CONCLUSION: Intravenous therapy with clarithromycin was associated with a reduction in the intensity of the intraamniotic inflammatory response in patients with PPROM with either intraamniotic infection or sterile intraamniotic inflammation. Moreover, treatment with clarithromycin was related to a reduction in the load of Ureaplasma spp DNA in the amniotic fluid of patients with PPROM <34 weeks of gestation.

Lactobacilli-dominated cervical microbiota in women with preterm prelabor rupture of membranes.

BACKGROUND: To determine the association between microbial invasion of the amniotic cavity (MIAC) and the presence of Lactobacillus crispatus- or Lactobacillus iners-dominated cervical microbiota in pregnancies with preterm prelabor rupture of membrane. Next, to assess the relationship between the presence of L. crispatus- or L. iners-dominated cervical microbiota and short-term neonatal morbidity. METHOD: A total of 311 women were included. Cervical samples were obtained using a Dacron polyester swab and amniotic fluid samples were obtained by transabdominal amniocentesis. Bacterial DNA, L. crispatus, and L. iners in the cervical samples were assessed by PCR. Cervical microbiota was assigned as L. crispatus- or L. iners-dominated when the relative abundance of L. crispatus or L. iners was ≥50% of the whole cervical microbiota, respectively. RESULTS: Women with MIAC showed a lower rate of L. crispatus-dominated cervical microbiota (21% vs. 39%; p = 0.003) than those without MIAC. Lactobacillus crispatus-dominated cervical microbiota was associated with a lower rate of early-onset sepsis (0% vs. 5%; p = 0.02). CONCLUSIONS: The presence of L. crispatus-dominated cervical microbiota in women with preterm prelabor rupture of membrane was associated with a lower risk of intra-amniotic complications and subsequent development of early-onset sepsis of newborns.

Methicillin-resistant Staphylococcus aureus in veterinary professionals in 2017 in the Czech Republic.

BACKGROUND: Cases of colonization or infection caused by Methicillin-resistant Staphylococcus aureus (MRSA) are frequently reported in people who work with animals, including veterinary personnel. The aim of this study was to determine the prevalence of MRSA colonization among veterinary professionals. A total of 134 nasal swabs from healthy attendees of a veterinary conference held in the Czech Republic were tested for presence of MRSA. The stains were further genotypically and phenotypically characterized. RESULTS: Nine isolated MRSA strains were characterized with sequence type (ST), spa type (t) and Staphylococcal Cassette Chromosome mec type. Five different genotypes were described, including ST398-t011-IV (n = 5), ST398-t2330-IV (n = 1), ST398-t034-V (n = 1), ST225-t003-II (n = 1) and ST4894-t011-IV (n = 1). The carriage of the animal MRSA strain was confirmed in 8 cases, characteristics of one strain corresponded to the possible nosocomial origin. Among animal strains were described three spa types (t011, t034, t2330) belonging into one dominating clonal complex spa-CC11. CONCLUSION: According to our results, the prevalence of nasal carriage of MRSA in veterinary personnel is 6.72%. Although we described an increase compared to the results of previous study (year 2008), the prevalence in the Czech Republic is still remaining lower than reported from neighboring countries. Our results also indicate that healthcare - associated MRSA strains are still not spread among animals.

Improved laboratory diagnostics of Streptococcus pneumoniae in respiratory tract samples through qPCR.

The aim of this study was to test the detection performance of the cpsA, lytA and ply genes through qPCR in the identification of Streptococcus pneumoniae in respiratory tract samples. Specificity was tested on a panel of 128 streptococci and other bacteria DNA samples. The qPCR assay was tested on a total of 51 respiratory tract samples from patients with community-acquired pneumonia (CAP). The specificity of the cpsA, lytA and ply genes was 100%, 100%, and 86%, respectively. The quantitative assessment, based on lytA, determined a cutoff value of ~2x104, 4x102 and 4x102 DNA copies per 1 mL of valid sputum, tracheal aspirate and bronchial aspirate samples, respectively. The results from the present study suggest that qPCR detection of all three genes would be optimal in the accurate detection of Streptococcus pneumoniae.

Complete Nucleotide Sequences of mcr-4.3-Carrying Plasmids in Acinetobacter baumannii Sequence Type 345 of Human and Food Origin from the Czech Republic, the First Case in Europe.

Here, we describe two plasmids carrying mcr-4.3 in two Acinetobacter baumannii strains isolated from imported food and a clinical sample. The comparative analysis of these plasmids, with two other plasmids reported in the NCBI database, highlighted the common origin of the plasmidic structure carrying mcr-4.3 This is the first case of the mcr-4.3 gene in a A. baumannii strain isolated from a clinical case in Europe. We hypothesize that food import is initiating the spread in Czech Republic.

Strong correlation between the rates of intrinsically antibiotic-resistant species and the rates of acquired resistance in Gram-negative species causing bacteraemia, EU/EEA, 2016.

BackgroundAntibiotic resistance, either intrinsic or acquired, is a major obstacle for treating bacterial infections.AimOur objective was to compare the country-specific species distribution of the four Gram-negative species Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter species and the proportions of selected acquired resistance traits within these species.MethodWe used data reported for 2016 to the European Antimicrobial Resistance Surveillance Network (EARS-Net) by 30 countries in the European Union and European Economic Area.ResultsThe country-specific species distribution varied considerably. While E. coli accounted for 31.9% to 81.0% (median: 69.0%) of all reported isolates, the two most common intrinsically resistant species P. aeruginosa and Acinetobacter spp. combined (PSEACI) accounted for 5.5% to 39.2% of isolates (median: 10.1%). Similarly, large national differences were noted for the percentages of acquired non-susceptibility to third-generation cephalosporins, carbapenems and fluoroquinolones. There was a strong positive rank correlation between the country-specific percentages of PSEACI and the percentages of non-susceptibility to the above antibiotics in all four species (rho > 0.75 for 10 of the 11 pairs of variables tested).ConclusionCountries with the highest proportion of P. aeruginosa and Acinetobacter spp. were also those where the rates of acquired non-susceptibility in all four studied species were highest. The differences are probably related to national differences in antibiotic consumption and infection prevention and control routines.

Pentraxin 3 in Noninvasively Obtained Cervical Fluid Samples from Pregnancies Complicated by Preterm Prelabor Rupture of Membranes.

PROBLEM: To determine the changes of pentraxin 3 (PTX3) level in noninvasively obtained cervical fluid samples from women with preterm prelabor rupture of membranes (PPROM) based on the presence of microbial invasion of the amniotic cavity (MIAC) and/or intra-amniotic inflammation (IAI), and intra-amniotic infection (the presence of both MIAC and IAI). METHODS OF STUDY: A total of 160 women with PPROM were included. Cervical fluid samples were obtained using a Dacron polyester swab and amniotic fluid samples were obtained by transabdominal amniocentesis. Cervical fluid PTX3 levels were assessed using enzyme-linked immunosorbent assay. RESULTS: PTX3 was found in all the cervical fluid samples and its levels were higher in women with MIAC, IAI, and intra-amniotic infection than in women without these conditions. When the women were categorized into four subgroups based on the presence of MIAC and/or IAI, women with intra-amniotic infection had higher cervical fluid PTX3 levels than those with sterile IAI (IAI alone), colonization (MIAC alone), or no MIAC or IAI. A cervical fluid PTX3 level of 11 ng/mL was the best value for identifying the presence of intra-amniotic infection in women with PPROM. CONCLUSIONS: PTX3 is a constituent of cervical fluid of women with PPROM. Cervical fluid PTX3 level reflects the situation in the intra-amniotic compartments of women with PPROM. Cervical fluid PTX3 is a potential marker for the noninvasive identification of intra-amniotic infection in PPROM.

Characterization of KPC-Encoding Plasmids from Enterobacteriaceae Isolated in a Czech Hospital.

Ten Enterobacteriaceae isolates collected in a Czech hospital carried blaKPC-positive plasmids of different sizes (∼30, ∼45, and ∼80 kb). Sequencing revealed three types of plasmids (A to C) with the Tn4401a transposon. Type A plasmids comprised an IncR backbone and a KPC-2-encoding multidrug resistance (MDR) region. Type B plasmids were derivatives of type A plasmids carrying an IncN3-like segment, while type C plasmids were IncP6 plasmids sharing the same KPC-2-encoding MDR region with type A and B plasmids.

Late preterm prelabor rupture of fetal membranes: fetal inflammatory response and neonatal outcome.

BackgroundTo characterize the influence of microbial invasion of the amniotic cavity (MIAC) and/or intra-amniotic inflammation (IAI) on the intensity of the fetal inflammatory response and the association between the presence of the fetal inflammatory response syndrome (FIRS) and short-term neonatal morbidity in the preterm prelabor rupture of membranes (PPROM) between the gestational ages of 34 and 37 weeks.MethodsOne hundred and fifty-nine women were included in the study. The umbilical cord blood interleukin (IL)-6 concentrations were determined using enzyme-linked immunosorbent assay kits. FIRS was defined based on the umbilical cord blood IL-6 concentration and the presence of funisitis and/or chorionic plate vasculitis.ResultsWomen with both MIAC and IAI had the highest median umbilical cord blood IL-6 concentrations and highest rates of FIRS. Women with FIRS had the higher rates of early-onset sepsis and intraventricular hemorrhage grades I and II when FIRS was characterized based on the umbilical cord blood IL-6 concentrations and the histopathological findings.ConclusionThe presence of both MIAC and IAI was associated with a higher fetal inflammatory response and a higher rate of FIRS. Different aspects of short-term neonatal morbidity were related to FIRS when defined by umbilical cord blood IL-6 concentrations and the histopathology of the placenta.

Amniotic fluid cell-free DNA in preterm prelabor rupture of membranes.

INTRODUCTION: We evaluated the levels of cell-free nuclear DNA (nDNA) and cell-free mitochondrial DNA (mtDNA) in the amniotic fluid supernatant from pregnancies complicated by preterm prelabor rupture of membranes (PPROM) based on evidence of microbial invasion of the amniotic cavity (MIAC) and/or intra-amniotic inflammation (IAI). MATERIAL AND METHODS: A total of 155 women with PPROM were included in this study. Amniotic fluid samples were obtained by transabdominal amniocentesis. The levels of cell-free nDNA and mtDNA in the amniotic fluid supernatant were assessed and quantified by real-time polymerase chain reaction. RESULTS: The levels of cell-free nDNA and mtDNA were higher in women with MIAC and IAI than in women without these conditions (nDNA: with MIAC: median 3.9 × 104 genome equivalent [GE]/mL vs without MIAC: median 1.2 × 104  GE/mL, with IAI: median: 5.3 × 104  GE/mL vs without IAI: median 1.2 × 104  GE/mL; mtDNA: with MIAC: median 9.2 × 105  GE/mL vs without MIAC: median 2.5 × 105  GE/mL, with IAI: median 1.1 × 106  GE/mL vs without IAI: median 2.5 × 105 ; all P values ≤ 0.01). Women with the microbial-associated IAI showed the highest levels of cell-free nDNA and mtDNA. CONCLUSIONS: Cell-free nDNA and mtDNA are constituents of the amniotic fluid supernatant from PPROM pregnancies. Both cell-free nDNA and mtDNA are involved in the intra-amniotic inflammatory response in women with PPROM.

Molecular Characterization of Carbapenemase-Producing Pseudomonas aeruginosa of Czech Origin and Evidence for Clonal Spread of Extensively Resistant Sequence Type 357 Expressing IMP-7 Metallo-ß-Lactamase.

The objective of this study was to perform molecular surveillance for assessing the spread of carbapenemase-producing Pseudomonas aeruginosa in Czech hospitals. One hundred thirty-six carbapenemase-producing isolates were recovered from 22 hospitals located throughout the country. Sequence type 357 (ST357) dominated (n = 120) among carbapenemase producers. One hundred seventeen isolates produced IMP-type (IMP-7 [n = 116] and IMP-1 [n = 1]) metallo-ß-lactamases (MßLs), 15 produced the VIM-2 MßL, and the remaining isolates expressed the GES-5 enzyme. The blaIMP-like genes were located in three main integron types, with In-p110-like being the most prevalent (n = 115). The two other IMP-encoding integrons (In1392 and In1393) have not been described previously. blaVIM-2-carrying integrons included In59-like, In56, and a novel element (In1391). blaGES-5 was carried by In717. Sequencing data showed that In-p110-like was associated with a Tn4380-like transposon inserted in genomic island LESGI-3 in the P. aeruginosa chromosome. The other integrons were also integrated into the P. aeruginosa chromosome. These findings indicated the clonal spread of ST357 P. aeruginosa, carrying the IMP-7-encoding integron In-p110, in Czech hospitals. Additionally, the sporadic emergence of P. aeruginosa producing different carbapenemase types, associated with divergent or novel integrons, punctuated the ongoing evolution of these bacteria.

Molecular Characterization of OXA-48-Like-Producing Enterobacteriaceae in the Czech Republic and Evidence for Horizontal Transfer of pOXA-48-Like Plasmids.

The aim of this study was to characterize the first cases and outbreaks of OXA-48-like-producing Enterobacteriaceae recovered from hospital settings in the Czech Republic. From 2013 to 2015, 22 Klebsiella pneumoniae isolates, 3 Escherichia coli isolates, and 1 Enterobacter cloacae isolate producing OXA-48-like carbapenemases were isolated from 20 patients. Four of the patients were colonized or infected by two or three different OXA-48-like producers. The K. pneumoniae isolates were classified into nine sequence types (STs), with ST101 being predominant (n = 8). The E. coli isolates were of different STs, while the E. cloacae isolate belonged to ST109. Twenty-four isolates carried blaOXA-48, while two isolates carried blaOXA-181 or blaOXA-232 Almost all isolates (n = 22) carried blaOXA-48-positive plasmids of a similar size (∼60 kb), except the two isolates producing OXA-181 or OXA-232. In an ST45 K. pneumoniae isolate and an ST38 E. coli isolate, S1 nuclease profiling plus hybridization indicated a chromosomal location of blaOXA-48 Sequencing showed that the majority of blaOXA-48-carrying plasmids exhibited high degrees of identity with the pOXA-48-like plasmid pE71T. Additionally, two novel pE71T derivatives, pOXA-48_30715 and pOXA-48_30891, were observed. The blaOXA-181-carrying plasmid was identical to the IncX3 plasmid pOXA181_EC14828, while the blaOXA-232-carrying plasmid was a ColE2-type plasmid, being a novel derivative of pOXA-232. Finally, sequencing data showed that the ST45 K. pneumoniae and ST38 E. coli isolates harbored the IS1R-based composite transposon Tn6237 containing blaOXA-48 integrated into their chromosomes. These findings underlined that the horizontal transfer of pOXA-48-like plasmids has played a major role in the dissemination of blaOXA-48 in the Czech Republic. In combination with the difficulties with their detection, OXA-48 producers constitute an important public threat.

The susceptibility to fosfomycin of Gram-negative bacteria isolates from urinary tract infection in the Czech Republic: data from a unicentric study.

BACKGROUND: Against a background of rapid increase of ß-lactamase-producing or multi-resistant pathogenic bacteria and the resulting lack of effective antibiotic treatment, some older antibiotics have been tested for new therapeutic uses. One of these is fosfomycin, to which according to studies these resistant bacteria are very sensitive. Our study was designed because there is no data on the fosfomycin susceptibility rate in the Czech Republic. METHOD: In this study from January 2013 to June 2014 3295 unique isolates of Gram-negative bacteria which had caused urinary tract infections were examined. The antibiotic susceptibility was measured by disk diffusion test. Both EUCAST and CLSI guidelines criteria (for fosfomycin only) were used for the antibiotic susceptibility evaluation. RESULTS: The most frequently tested bacterial isolates were Escherichia coli (51.3%, n = 1703), Klebsiella pneumoniae (19.4%, n = 643) and Proteus spp. (11.8%, n = 392). Among all isolates 29.0% (n = 963) were resistant to fluoroquinolones, 11.3% (n = 374) produced extended spectrum ß-lactamase and 4.2% (n = 141) produced AmpC ß-lactamase. The overall in vitro susceptibility was significantly higher for fosfomycin compared to the other tested per-oral antibiotics (nitrofurantoin, ampicillin, co-trimoxazole, ciprofloxacin and cefuroxime) against all tested Gram-negative rod isolates (excluding Morganella morgani and Acinetobacter spp. isolates). Fosfomycin also remained highly active against those isolates with extended spectrum ß-lactamase (ESBL) production (95.8% in Escherichia coli isolates and 85.3% in Klebsiella pneumoniae isolates), unlike other tested per-oral antibiotics, which showed significant (p < 0.0001) susceptibility decrease. CONCLUSION: We have confirmed in the Czech Republic the very high susceptibility to fosfomycin trometamol of urinary tract infection pathogens, particularly Gram-negative rods including those producing ß-lactamase.

[Antimicrobial activity of ceftolozane/tazobactam against Enterobacteriaceae and Pseudomonas aeruginosa in the Czech Republic in 2016].

OBJECTIVE: Ceftolozane/tazobactam is an antibiotic effective against Gram-negative bacteria(including Pseudomonas aeruginosa).The study aimed at determining the effectiveness of the novel antibiotic in the Czech Republic. MATERIAL AND METHODS: The effectiveness of the antibiotic was studied in 16 Czech laboratories in 822 Enterobacteriaceae isolates (including AmpC and ESBL producers) and P. aeruginosa causing complicated intraabdominal or urinary tract infections. Minimum inhibitory concentrations were determined using the Etest. With the exception of Citrobacter freundii and Enterobacter cloacae, ceftozolane/tazobactam proved to be very effective against Enterobacteriaceae; no P. aeruginosa strain was resistant to the antibiotic. CONCLUSION: The results confirmed good activity of ceftozolane/tazobactam in vitro against Enterobacteriaceae (Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca, Proteus mirabilis) and P. aeruginosa in the Czech Republic.

[Sharing bacterial microbiota between owners and their pets (dogs, cats)].

INTRODUCTION: The microbiological aspect of a relationship between pets (dogs/cats) and their owners is mainly concerned with the incidence of shared bacterial species, in particular potential pathogens. Given the great popularity of sharing homes with pets (dogs/cats) in the Czech Republic, there is an increased possibility of communication between microbiota of the two macroorganisms (pet and owner). The aim of the study was to determine the biodiversity of shared bacteria and possibility of exchange of genes of resistance to antimicrobial agents between potential pathogens based on the close relationship between pets and humans. METHODS: A total of 103 samples were collected from 20 pairs (20 owners, 16 dogs and 4 cats). All owners completed a questionnaire with their pets' veterinarians. In owners, swabs were collected from the nasal mucosa, armpit and interdigital spaces of the foot. In pets, swabs were obtained from the external auditory meatus and nasal mucosa. In individuals with skin lesions, samples were also collected from the affected areas. Bacterial species were identified by culture and matrix-assisted laser desorption/ionization - time of flight (MALDI-TOF) mass spectrometry. In shared species, susceptibility to antibiotics was tested by the disk diffusion method. Statistical methods were used to correlate the closeness of relationship with the number of shared bacterial species and to correlate previous antimicrobial therapy with shared resistance of the common bacteria. RESULTS: Analysis of the questionnaires showed that 65 % of owners who participated in the study kept more pets at home than only the tested one. In the previous year, 5 % of pets and 5 % of owners received antimicrobial therapy. As many as 45 % of dogs or cats slept in their owners' beds and 80 % rested on a sofa together with their owners. Also, 45 % owners had their faces licked by pets. Eighty percent of pets were fed with several types of food (dry food and cooked food). Further, 70 % of pets lived permanently with their owners in the same household. A total of 76 bacterial species of 33 genera were identified. The most frequently isolated species (29 samples) was S. intermedius. Seventeen bacterial species occurring in both humans and animals were found and identified. At least one bacterial species was shared by 11 pairs and two shared species were found in two pairs. The shared species were S. intermedius, E. coli, E. faecalis, A. lwoffii, P. putida and S. aureus. Antimicrobial susceptibility was tested in the shared species. Common antimicrobial resistance was found in four pairs. In one pair, shared E. faecalis showed identical resistance to co-trimoxazole; in another pair, S. intermedius was resistant to gentamycin, erythromycin, clindamycin and co-trimoxazole. The third resistant bacterial species was E. coli; in one pair, it showed borderline resistance to colistin; in the second case, it was fully resistant to this antimicrobial agent. The other pairs with shared bacteria did not show any common resistance. CONCLUSION: The study results showed that there was an association between closeness of the human-pet relationship and the prevalence of shared bacterial species. Pairs with a close relationship were 37.5 % more likely to share bacteria than pairs with a less close relationship. The study suggests that antimicrobial therapy in at least one pair member may increase the risk of shared bacterial resistance.

A genomic portrait of the emergence, evolution, and global spread of a methicillin-resistant Staphylococcus aureus pandemic.

The widespread use of antibiotics in association with high-density clinical care has driven the emergence of drug-resistant bacteria that are adapted to thrive in hospitalized patients. Of particular concern are globally disseminated methicillin-resistant Staphylococcus aureus (MRSA) clones that cause outbreaks and epidemics associated with health care. The most rapidly spreading and tenacious health-care-associated clone in Europe currently is EMRSA-15, which was first detected in the UK in the early 1990s and subsequently spread throughout Europe and beyond. Using phylogenomic methods to analyze the genome sequences for 193 S. aureus isolates, we were able to show that the current pandemic population of EMRSA-15 descends from a health-care-associated MRSA epidemic that spread throughout England in the 1980s, which had itself previously emerged from a primarily community-associated methicillin-sensitive population. The emergence of fluoroquinolone resistance in this EMRSA-15 subclone in the English Midlands during the mid-1980s appears to have played a key role in triggering pandemic spread, and occurred shortly after the first clinical trials of this drug. Genome-based coalescence analysis estimated that the population of this subclone over the last 20 yr has grown four times faster than its progenitor. Using comparative genomic analysis we identified the molecular genetic basis of 99.8% of the antimicrobial resistance phenotypes of the isolates, highlighting the potential of pathogen genome sequencing as a diagnostic tool. We document the genetic changes associated with adaptation to the hospital environment and with increasing drug resistance over time, and how MRSA evolution likely has been influenced by country-specific drug use regimens.