Ventilator-associated pneumonia (VAP) is one of the most severe complications affecting mechanically ventilated patients. The condition is caused by microaspiration of potentially pathogenic bacteria from the upper respiratory tract into the lower respiratory tract or by bacterial pathogens from exogenous sources such as healthcare personnel, devices, aids, fluids and air. The aim of our prospective, observational study was to confirm the hypothesis that in the etiology of VAP, an important role is played by etiological agents from the upper airway bacterial microflora. At the same time, we studied the hypothesis that the vertical spread of bacterial pathogens is more frequent than their horizontal spread among patients. A total of 697 patients required mechanical ventilation for more than 48 h. The criteria for VAP were met by 47 patients. Clonality of bacterial isolates from 20 patients was determined by comparing their macrorestriction profiles obtained by pulsed-field gel electrophoresis (PFGE). Among these 20 patients, a total of 29 PFGE pulsotypes of Klebsiella spp. and Escherichia spp. strains were observed. The high variability of clones proves that there was no circulation of bacterial pathogens among hospitalized patients. Our finding confirms the development of VAP as a result of bacterial microaspiration and therefore the endogenous origin of VAP.

• Klíčová slova
• Escherichia spp., Klebsiella spp., clonality, endogenous infection, pulsed-field gel electrophoresis (PFGE), ventilator-associated pneumonia,
• Publikační typ
• časopisecké články MeSH

AIMS: To determine the presence of antibiotic-resistant faecal Escherichia coli and Enterococcus spp. in feral pigeons (Columba livia forma domestica) in the Czech Republic. METHODS AND RESULTS: Cloacal swabs of feral pigeons collected in the city of Brno in 2006 were cultivated for antibiotic-resistant E. coli. Resistance genes, class 1 and 2 integrons, and gene cassettes were detected in resistant isolates by polymerase chain reaction (PCR). The samples were also cultivated for enterococci. Species status of enterococci isolates was determined using repetitive extragenic palindromic-PCR. Resistance genes were detected in resistant enterococci by PCR. E. coli isolates were found in 203 of 247 pigeon samples. Antibiotic resistance was recorded in three (1·5%, n(E. coli) =203) isolates. Using agar containing ciprofloxacin, 12 (5%, n(samples) =247) E. coli strains resistant to ciprofloxacin were isolated. No ESBL-producing E. coli isolates were detected. A total of 143 enterococci were isolated: Ent. faecalis (36 isolates), Ent. faecium (27), Ent. durans (19), Ent. hirae (17), Ent. mundtii (17), Ent. gallinarum (12), Ent. casseliflavus (12) and Ent. columbae (3). Resistance to one to four antibiotics was detected in 45 (31%) isolates. Resistances were determined by tetK, tetL, tetM, tetO, aac(6')aph(2''), ant(4')-Ia, aph(3')-IIIa, ermB, pbp5, vanA and vanC1 genes. CONCLUSIONS: Antibiotic-resistant E. coli and Enterococcus spp. occurred in feral pigeons in various prevalences. SIGNIFICANCE AND IMPACT OF THE STUDY: Feral pigeon should be considered a risk species for spreading in the environment antimicrobial resistant E. coli and enterococci.

• MeSH
• antibakteriální látky farmakologie   MeSH
• bakteriální léková rezistence genetika   MeSH
• Columbidae mikrobiologie   MeSH
• Enterococcus účinky léků   genetika   izolace a purifikace   MeSH
• Escherichia coli účinky léků   genetika   izolace a purifikace   MeSH
• feces mikrobiologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antibakteriální látky MeSH

In the present study, the effect of Lactobacillus spp. against enteropathogenic Escherichia coli 08:K88+Ent+ under in vitro and in vivo conditions has been compared and the effect of inoculation of Lactobacillus spp. upon the colonization of both the jejunum and ileum by enteropathogenic E. coli 08:K88+Ent+ in 9 gnotobiotic pigs has been observed. Under in vitro conditions, the strain Lactobacillus spp. showed the inhibition of 2.1 +/- 0.1 mm against enteropathogenic E. coli 08:K88+Ent+. Two days after the inoculation, the enteropathogenic E. coli 08:K88+Ent+, inoculated to the control group of gnotobiotic pigs (E), colonized the mucosa of both jejunum and ileum of gnotobiotic pigs at counts of 6.41 and 6.08 log 10/cm2, respectively. In experimental group (L-E), the counts of adhered enteropathogenic E. coli in the identical sections of the small intestine, following the inoculation by Lactobacillus spp., amounted to 6.35 and 6.43 log/cm2, respectively. In both groups, numbers of E. coli in the intestinal content of both jejunum and ileum were nearly the same (group E 9.03 and 9.31 log 10/ml; group L-E 8.97 and 9.11 log 10/ml). Two to five days after E. coli inoculation, Lactobacillus spp. counts adhered to the jejunal wall ranged from 5.4 to 6.49 log 10 cm2; in the ileum they ranged from 6.05 to 6.77 log 10 cm2. In the jejunal content, the lactobacilli counts ranged from 6.81 to 8.86 log 10/ml and in the ileum from 8.5 to 898 log 10/ml. Two days after the E. coli inoculation, the concentration of lactic acid in the content of jejunum in gnotobiotic pigs of the groups E and L-E was 16.3 mmol/l and 23.6 mmol/l, respectively. The concentration of acetic acid in the jejunum of the pigs of E and L-E groups was 15.9 mmol/l and 19.6 mmol/l, respectively. Similarly, the higher concentrations of both acids were found also in the ileum of the L-E pigs. The results obtained indicate that the used strain of Lactobacillus spp. which has been preventively inoculated to gnotobiotic pigs, did not prevent the adhesion of enteropathogenic E. coli 08:K88+Ent+ to the mucosa of both jejunum and ileum also despite of the demonstrated inhibitory effect against enteropathogenic E. coli under in vitro conditions and despite good adherent ability in vivo. In both groups of animals, the diseases with pronounced clinical signs as well as losses have occurred.

• MeSH
• Escherichia coli růst a vývoj   fyziologie   MeSH
• gnotobiologické modely MeSH
• Lactobacillus růst a vývoj   fyziologie   MeSH
• prasata mikrobiologie   MeSH
• tenké střevo mikrobiologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

The application of rapid, specific, and sensitive methods for pathogen detection and quantification is very advantageous in diagnosis of human pathogens in several applications, including food analysis. The aim of this study was the evaluation of a method for the multiplexed detection and quantification of three significant foodborne pathogenic species (Escherichia coli O157, Salmonella spp., and Listeria monocytogenes). The assay combines specific DNA extraction by multiplex magnetic capture hybridization (mMCH) with multiplex real-time PCR. The amplification assay showed linearity in the range 106-10 genomic units (GU)/PCR for each co-amplified species. The sensitivity corresponded to 1 GU/PCR for E. coli O157 and L. monocytogenes, and 10 GU/PCR for Salmonella spp. The immobilization process and the hybrid capture of the MCH showed good efficiency and reproducibility for all targets, allowing the combination in equal amounts of the different nanoparticle types in mMCH. MCH and mMCH efficiencies were similar. The detection limit of the method was 10 CFU in samples with individual pathogens and 102 CFU in samples with combination of the three pathogens in unequal amounts (amount's differences of 2 or 3 log). In conclusion, this multiplex molecular platform can be applied to determine the presence of target species in food samples after culture enrichment. In this way, this method could be a time-saving and sensitive tool to be used in routine diagnosis.

• Klíčová slova
• Escherichia coli O157, Listeria monocytogenes, Multiplex magnetic capture hybridization, Multiplex real-time PCR, Quantitative detection, Salmonella spp.,
• MeSH
• Escherichia coli O157 klasifikace   genetika   MeSH
• hybridizace nukleových kyselin MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• lidé MeSH
• Listeria monocytogenes klasifikace   genetika   MeSH
• multiplexová polymerázová řetězová reakce MeSH
• Salmonella klasifikace   genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

The in-vitro activity of cefotaxime and cefoperazone were compared using clinically isolated Escherichia coli, Klebsiella spp and Pseudomonas aeruginosa. Cefotaxime was found on a weight to weight basis, to be much more active than cefoperazone. All the three species studied show the presence of cefoperazone-resistant population which were sensitive to cefotaxime. The possible mechanisms of resistance to these antibiotics were discussed.

• MeSH
• antibiotická rezistence MeSH
• cefoperazon farmakologie   MeSH
• cefotaxim farmakologie   MeSH
• Escherichia coli účinky léků   MeSH
• Klebsiella účinky léků   MeSH
• mikrobiální testy citlivosti MeSH
• Pseudomonas aeruginosa účinky léků   MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH
• Názvy látek
• cefoperazon MeSH
• cefotaxim MeSH

UNLABELLED: The spread of antimicrobial resistance from human activity derived sources to natural habitats implicates wildlife as potential vectors of antimicrobial resistance transfer. Wild birds, including corvid species can disseminate mobile genetic resistance determinants through faeces. This study aimed to determine the occurrence of plasmid-mediated quinolone resistance (PMQR) genes in Escherichia coli and Klebsiella spp. isolates obtained from winter roosting sites of American crows (Corvus brachyrhynchos) and common ravens (Corvus corax) in Canada. Faecal swabs were collected at five roosting sites across Canada. Selective media isolation and multiplex PCR screening was utilized to identify PMQR genes followed by gene sequencing, pulse-field gel electrophoresis and multilocus sequence typing to characterize isolates. Despite the low prevalence of E. coli containing PMQR (1·3%, 6/449), qnrS1, qnrB19, qnrC, oqxAB and aac(6')-Ib-cr genes were found in five sequence types (ST), including E. coli ST 131. Conversely, one isolate of Klebsiella pneumoniae contained the plasmid-mediated resistance gene qnrB19. Five different K. pneumoniae STs were identified, including two novel types. The occurrence of PMQR genes and STs of public health significance in E. coli and Klebsiella pneumoniae recovered from corvids gives further evidence of the anthropogenic derived dissemination of antimicrobial resistance determinants at the human activity-wildlife-environment interface. SIGNIFICANCE AND IMPACT OF THE STUDY: This study examined large corvids as possible vector species for the dissemination of antimicrobial resistance in indicator and pathogenic bacteria as a means to assess the anthropogenic dissemination of plasmid-mediated quinolone resistance (PMQR) genes. Although rare, PMQR genes were found among corvid populations across Canada. The clinically important Escherichia coli strain ST131 containing aac(6')-Ib-cr gene along with a four-class phenotypic antimicrobial resistance (AMR) pattern as well as one Klebsiella pneumoniae strain containing a qnrB19 gene were identified in one geographical location. Corvids are a viable vector for the circulation of PMQR genes and clinically important clones in wide-ranging environments.

• Klíčová slova
• E. coli, Klebsiella pneumoniae, PMQR genes, ST131, antimicrobial resistance,
• MeSH
• antibakteriální látky farmakologie   MeSH
• bakteriální léková rezistence genetika   MeSH
• chinolony farmakologie   MeSH
• Escherichia coli účinky léků   genetika   izolace a purifikace   MeSH
• infekce bakteriemi rodu Klebsiella farmakoterapie   mikrobiologie   MeSH
• infekce vyvolané Escherichia coli farmakoterapie   mikrobiologie   MeSH
• Klebsiella pneumoniae účinky léků   genetika   izolace a purifikace   MeSH
• lidé MeSH
• mikrobiální testy citlivosti MeSH
• multilokusová sekvenční typizace MeSH
• nemoci ptáků farmakoterapie   mikrobiologie   MeSH
• plazmidy genetika   MeSH
• průřezové studie MeSH
• vrány mikrobiologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Kanada MeSH
• Názvy látek
• antibakteriální látky MeSH
• chinolony MeSH

OBJECTIVES: The global occurrence of antibiotic resistance genes in bacteria in water environments is an increasing concern. Treated wastewater was sampled daily over a 45 day period from the outflow of a municipal wastewater treatment plant in Brno, Czech Republic, and examined for extended-spectrum β-lactamase (ESBL)-producing bacteria. METHODS: Water samples were cultivated on MacConkey agar with cefotaxime (2 mg/L) and individual colonies were examined for ESBL production. Phenotypic ESBL-positive bacteria identified as Escherichia coli or Klebsiella spp. were tested for the presence of antibiotic resistance genes, the virulence gene afa/dra and the bla(CTX-M) upstream region. Genetic relatedness was analysed by PFGE, multilocus sequence typing and plasmid analysis. RESULTS: A total of 68 ESBL-producing Enterobacteriaceae isolates were detected in 34 out of 45 wastewater samples. ESBL-producing isolates included 26 E. coli isolates, 4 Klebsiella pneumoniae isolates and 1 Klebsiella oxytoca isolate. The pandemic and multiresistant B2-O25b-ST131 clone was predominant, being detected among 19 E. coli isolates, and 17 of the B2-O25b-ST131 isolates were positive for the FIA replicon and the afa/dra operon and had an IS26 element flanking bla(CTX-M-15). Seventeen of the B2-O25b-ST131 isolates showed closely related PFGE profiles (defined by 84% band similarity) and belonged to identical clonal groups. CONCLUSIONS: The results highlight the inadequacy of the treatment process in removing multiresistant bacteria from municipal wastewater and point to a risk of transmission of clinically important multiresistant strains, such as the pandemic ST131 clone, to the environment. This is the first study demonstrating the pandemic ST131 clone in wastewater.

• MeSH
• bakteriologické techniky metody   MeSH
• beta-laktamasy metabolismus   MeSH
• čištění vody MeSH
• DNA bakterií genetika   MeSH
• DNA fingerprinting MeSH
• Escherichia coli klasifikace   enzymologie   genetika   izolace a purifikace   MeSH
• faktory virulence genetika   MeSH
• genotyp MeSH
• Klebsiella klasifikace   enzymologie   genetika   izolace a purifikace   MeSH
• kultivační média chemie   MeSH
• lidé MeSH
• mikrobiologie vody * MeSH
• multilokusová sekvenční typizace MeSH
• plazmidy analýza   MeSH
• polymerázová řetězová reakce MeSH
• pulzní gelová elektroforéza MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• beta-lactamase CTX-M-15 MeSH Prohlížeč
• beta-laktamasy MeSH
• DNA bakterií MeSH
• faktory virulence MeSH
• kultivační média MeSH

This study is focused on Escherichia spp. isolates resistant to critically important antibiotics (cefotaxime, ciprofloxacin and colistin) among Caspian gull's (Larus cachinnans) chicks nesting in the Nove Mlyny Water Reservoir, Czech Republic. The prevalence of antimicrobial resistance (AMR) in bacteria within wild birds is commonly evaluated using a single sampling event, capturing only a brief and momentary snapshot at a particular location. Therefore, the Caspian gulls in our study were sampled in May 2018 (n = 72) and May 2019 (n = 45), and a water sample was taken from the reservoir (2019). We obtained 197 isolates identified as E. coli by MALDI-TOF MS. A total of 158 representative isolates were whole-genome sequenced, 17 isolates were then reclassified to Escherichia albertii. We observed a higher (86 %; 62/72) occurrence of ESBL/AmpC-producing Escherichia spp. among gulls in 2018 compared to 38 % (17/45) in 2019 (p < 0.00001). The decrease in prevalence was linked to clonal lineage of E. coli ST11893 predominating in 2018 which carried blaCMY-2 and which was not recovered from the gulls in 2019. Oppositely, several Escherichia STs were found in gulls from both years as well as in the water sample including STs commonly recognized as internationally high-risk lineages such as ST10, ST58, ST88, ST117, ST648 or ST744. Phylogenetic analysis of E. coli from EnteroBase from countries where these particular gulls wander revealed that some STs are commonly found in various sources including humans and a portion of them is even closely related (up to 100 SNPs) to our isolates. We demonstrated that the occurrence of AMR in Escherichia can vary greatly in time in synanthropic birds and we detected both, a temporary prevalent lineage and several persistent STs. The close relatedness of isolates from gulls and isolates from EnteroBase highlights the need to further evaluate the risk connected to wandering birds.

• Klíčová slova
• AMR in wild birds, Environmental persistence of AMR bacteria, Successful Escherichia STs,
• MeSH
• antibakteriální látky * farmakologie   MeSH
• bakteriální léková rezistence MeSH
• Charadriiformes * mikrobiologie   MeSH
• Escherichia coli účinky léků   genetika   izolace a purifikace   MeSH
• Escherichia účinky léků   genetika   MeSH
• longitudinální studie MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• antibakteriální látky * MeSH

Overpopulation of domestic pigeons is considered to be one of the major problems of urban centers, as these birds are responsible for the dissemination of relevant pathogens to animal and human health. The aim of this study was to detect potentially pathogenic Escherichia coli and Salmonella spp. in domestic pigeons captured in areas near silos used for grain and feed storage, analyzing the antimicrobial sensitivity and the presence of virulence-associated genes. We evaluated 41 pigeons. From each bird, cecal contents and a pool of viscera (heart, spleen, and liver) were collected. Fifty strains of E. coli and three strains of S. Typhimurium were isolated. The antimicrobial susceptibility assay showed that 2% of the isolates of E. coli were resistant to chloramphenicol and the combination of sulfamethoxazole + trimethoprim and 4% to tetracycline, doxycycline, and sulfonamide. The three S. Typhimurium strains were sensitive to all antimicrobials tested. The pathogenicity profile demonstrated that no E. coli isolates showed a STEC compatible profile. Regarding the APEC pathotype, all genes were observed in 8% of E. coli, 6% had only the iss gene and 4% presented ompT, hlyF, and iutA genes. invA, hilA, avrA, and lpfA genes were detected in 100% of Salmonella isolates. The sitC and pefA genes were only present in one strain and the remaining genes were detected in two. In conclusion, it was found that pigeons living in the vicinity of silos are carriers of important pathogens, and control measures should be taken to minimize animal and human health risks.

• MeSH
• antibakteriální látky farmakologie   MeSH
• bakteriální infekce epidemiologie   mikrobiologie   veterinární   MeSH
• bakteriální léková rezistence účinky léků   genetika   MeSH
• bakteriální proteiny genetika   MeSH
• Columbidae mikrobiologie   MeSH
• Escherichia coli účinky léků   genetika   izolace a purifikace   patogenita   MeSH
• fylogeneze MeSH
• hospodářská zvířata mikrobiologie   MeSH
• mikrobiální testy citlivosti MeSH
• nemoci drůbeže epidemiologie   mikrobiologie   MeSH
• Salmonella účinky léků   genetika   izolace a purifikace   patogenita   MeSH
• virulence genetika   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Brazílie epidemiologie   MeSH
• Názvy látek
• antibakteriální látky MeSH
• bakteriální proteiny MeSH

RATIONALE: Tracing isotopically labeled water into proteins allows for the detection of species-specific metabolic activity in complex communities. However, a stress response may alter the newly synthesized proteins. METHODS: We traced 18-oxygen from heavy water into proteins of Escherichia coli K12 grown from permissive to retardant temperatures. All samples were analyzed using UPLC/Orbitrap Q-Exactive-MS/MS operating in positive electrospray ionization mode. RESULTS: We found that warmer temperatures resulted in significantly (P-value < 0.05) higher incorporation of 18-oxygen as seen by both substrate utilization as relative isotope abundance (RIA) and growth as labeling ratio (LR). However, the absolute number of peptides with incorporation of 18-oxygen showed no significant correlation to temperature, potentially caused by the synthesis of different proteins at low temperatures, namely, proteins related to cold stress response. CONCLUSIONS: Our results unveil the species-specific cold stress response of E. coli K12 that could be misinterpreted as general growth; this is why the quantity as RIA and LR but also the quality as absolute number of peptides with incorporation (relative abundance, RA) and their function must be considered to fully understand the activity of microbial communities.

• MeSH
• Escherichia coli K12 * chemie   metabolismus   fyziologie   MeSH
• izotopové značení metody   MeSH
• izotopy kyslíku * analýza   metabolismus   MeSH
• nízká teplota MeSH
• proteiny z Escherichia coli * analýza   chemie   metabolismus   MeSH
• reakce na chladový šok fyziologie   MeSH
• tandemová hmotnostní spektrometrie metody   MeSH
• vysokoúčinná kapalinová chromatografie metody   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• izotopy kyslíku * MeSH
• Oxygen-18 MeSH Prohlížeč
• proteiny z Escherichia coli * MeSH